Preface

Oxidative stress plays a crucial role in the pathophysiology of various diseases when there is a disruption of the intracellular redox balance and the homeostatic balance between cellular oxidants and antioxidants. Reactive oxygen species (ROS), although essential for normal physiologic processes, are deleterious when produced in excess. Free radicals include not only ROS but also reactive nitrogen species (RNS) such as nitric oxide and peroxynitrite similarly leading to nitrosative stress. These molecules react with molecular targets including proteins, lipids, and nucleic acids contributing to mitochondrial injury and cellular dysfunction.

This book intends to provide the readers with an extensive overview of the novel ap‐ proaches and prospects based on oxidative and nitrosative stress in the pathophysiology of various diseases and in the current treatment strategies with antioxidants, and it is the one to which the authors have made significant contributions in all chapters.

I believe that this book will provide a good grounding in pointing the way to new disciplines that will contribute to the evolution of strategies for creating, analyzing, and presenting the medical information in the future stimulating our colleagues at all multidisciplinary levels.

> **Prof. Dr. Pinar Atukeren** Istanbul University Cerrahpasa Medical Faculty Department of Medical Biochemistry Turkey

**Chapter 1**

Provisional chapter

**Oxidative Stress: Noxious but Also Vital**

DOI: 10.5772/intechopen.73394

Oxidative Stress: Noxious but Also Vital

Margarete Dulce Bagatini,

Margarete Dulce Bagatini,

Carla Santos de Oliveira,

Carla Santos de Oliveira,

Cintia dos Santos Moser,

Abstract

Graciele Almeida de Oliveira,

Jeandre Augusto dos Santos Jaques,

Jeandre Augusto dos Santos Jaques,

Micheli Mainardi Pillat, Aline Mânica,

Aline Mânica, Cintia dos Santos Moser,

http://dx.doi.org/10.5772/intechopen.73394

cycle, apoptosis, and inflammation.

Lucas Derbocio dos Santos and Henning Ulrich

Lucas Derbocio dos Santos and Henning Ulrich

Graciele Almeida de Oliveira, Micheli Mainardi Pillat,

The imbalance between reactive oxygen species (ROS) production and antioxidant defenses determines the condition called oxidative stress. When there is an increase in ROS production or a decrease in the antioxidant defenses, this systemic antioxidant/pro-oxidant imbalance may lead to the accumulation of oxidative damage, which, in turn, may lead to a modification of biomolecules. These consist of reactions resulting in protein adducts, DNA oxidation, and formation of lipid peroxides, which, in turn, reduce the cellular functional capacity and increase the risk of disease development. The body has natural scavenging systems against free radicals and other reactive species. However, sometimes the endogenous antioxidant capacity is exceeded by the production of ROS. When this occurs, exogenous antioxidants exert important function for the human health. These bioactive compounds act preventing and neutralizing the formation of new reactive species and free radicals. In some cases, an increase of ROS can help the host to resolve an infection or even to control the tumor growth. Finally, the levels of ROS can be perceived by signal transduction pathways involving known targets (i.e., p53, Ras, and NF-κB) and regulate physiopathological events such as the cellular

Keywords: reactive species, cellular oxidation, antioxidant system, health, disease

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and eproduction in any medium, provided the original work is properly cited.

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use,

distribution, and reproduction in any medium, provided the original work is properly cited.

Additional information is available at the end of the chapter

Additional information is available at the end of the chapter

### **Chapter 1**

Provisional chapter

## **Oxidative Stress: Noxious but Also Vital**

DOI: 10.5772/intechopen.73394

Oxidative Stress: Noxious but Also Vital

Margarete Dulce Bagatini, Jeandre Augusto dos Santos Jaques, Carla Santos de Oliveira, Graciele Almeida de Oliveira, Micheli Mainardi Pillat, Aline Mânica, Cintia dos Santos Moser, Lucas Derbocio dos Santos and Henning Ulrich Margarete Dulce Bagatini, Jeandre Augusto dos Santos Jaques, Carla Santos de Oliveira, Graciele Almeida de Oliveira, Micheli Mainardi Pillat, Aline Mânica, Cintia dos Santos Moser, Lucas Derbocio dos Santos and Henning Ulrich

Additional information is available at the end of the chapter Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.73394

#### Abstract

VIII Preface

The imbalance between reactive oxygen species (ROS) production and antioxidant defenses determines the condition called oxidative stress. When there is an increase in ROS production or a decrease in the antioxidant defenses, this systemic antioxidant/pro-oxidant imbalance may lead to the accumulation of oxidative damage, which, in turn, may lead to a modification of biomolecules. These consist of reactions resulting in protein adducts, DNA oxidation, and formation of lipid peroxides, which, in turn, reduce the cellular functional capacity and increase the risk of disease development. The body has natural scavenging systems against free radicals and other reactive species. However, sometimes the endogenous antioxidant capacity is exceeded by the production of ROS. When this occurs, exogenous antioxidants exert important function for the human health. These bioactive compounds act preventing and neutralizing the formation of new reactive species and free radicals. In some cases, an increase of ROS can help the host to resolve an infection or even to control the tumor growth. Finally, the levels of ROS can be perceived by signal transduction pathways involving known targets (i.e., p53, Ras, and NF-κB) and regulate physiopathological events such as the cellular cycle, apoptosis, and inflammation.

Keywords: reactive species, cellular oxidation, antioxidant system, health, disease

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons © 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and eproduction in any medium, provided the original work is properly cited.

### 1. Cellular respiration and generation of reactive species in the mitochondria: implications in cell viability and aging

Oxidative phosphorylation is the center of energy metabolism in plants, animals and several microbial life forms [1]. In eukaryotes, this process occurs in mitochondria. The mitochondria is a cytoplasmic organelle surrounded by two membranes, outer and inner membrane, which main function is the production of most of the phosphate compounds necessary for the energetic balance of the cell. In addition, other functions such as the regulation of the body's heat generation [2–4] programmed cell death [5–7], reactive oxygen species (ROS) generation and cell signaling [8] is also associated with mitochondria. Cellular vitality is directly related to mitochondria, and mitochondrial dysfunctions are frequent causes of accidental cell death [5, 9–11], cancer [12, 13], diabetes [14–16] and neurodegenerative diseases [17–19], among others.

The characterization of the respiratory electron chain could be performed in studies using the fractionation of its components by certain detergents that at low concentrations break the interactions between proteins and lipids in the membranes, leaving associations between proteins intact [20]. In electron transport chain, through this process, four protein complexes were found. They were named complex I (or NADH-Ubiquinone oxidoreductase), complex II (succinate dehydrogenase), complex III (Ubiquinol -cytochrome c oxidoreductase, or complex bc1) e complex IV (cytochrome c oxidase). The complex V is also known as ATP synthase. Despite glycerophosphate dehydrogenase (glycerol-3-phosphate dehydrogenase) and ETF–ubiquinone oxidoreductase have not complex nomenclature, they are connect to the electron transport chain, as complex I and complex II, i.e., delivering electron to ubiquinone [21].

The redox carriers within the respiratory chain consists of flavoprotein containing tightly bound FAD or FMN as prosthetic groups, protein-bound couper, ironsulphur (nonhaem iron) proteins and cytochromes, with haem prosthetic groups. The ubiquinone also participated in electron transport chain as a free and diffusible cofactor [20]. While electron transport occurs through the mitochondrial complexes, complexes I, II, and III pump protons from mitochondrial matrix to the intermembrane space. The energy associated to this process is used to the production of ATP by ATP synthase (Figure 1) [22].

> superoxide in both sides of inner mitochondrial membrane [29]. Complex II could theoretically generate superoxide, due presence of flavoprotein in its structure. However, the redox centers are arranged in a manner that aids the prevention of ROS by avoiding the access of O2 to the flavoprotein. This may explain the reason why this complex does not show a ROS formation by itself [30], but only due reverse electron transfer, i.e., when electrons flow from succinate to

•, superoxide radical anion; SOD2, superoxide dismutase 2 or

Oxidative Stress: Noxious but Also Vital http://dx.doi.org/10.5772/intechopen.73394 3

Figure 1. Electron transport chain, ROS, and antioxidant defense. The electron transport chain receives electrons from reduced compounds, as NADH in complex I (also called NADH coenzyme Q reductase) and succinate or FADH2 in complex II (succinate dehydrogenase) and transfers them successively to coenzyme Q or ubiquinone, complex III, complex IV and finally to molecular oxygen with the formation of water. Concomitant with electron transport, protons are transferred from the mitochondrial matrix to the intermembrane space by complexes I (in mammals, but not in yeasts), complex III and IV. The difference in electrochemical potential between intermembrane space and matrix is used by ATP synthase to produce ATP from ADP and inorganic phosphate. During the passage of electrons through the complexes, a small fraction is leaked to oxygen at intermediate points, producing the superoxide radical anion, which in the mitochondrial matrix can be removed by SOD2 forming H2O2. Hydrogen peroxide can be converted to water in the mitochondrial matrix by glutathione peroxidase (mammals). Abbreviations: ADP, adenosine diphosphate; ATP, adenosine triphosphate; UQ, ubiquinone; Cyt, cytochrome c; SDH, succinate dehydrogenase; I, complex I or NADH coenzyme Q reductase; II, complex II or succinate dehydrogenase; III,

In addition to the electron transport chain, recent studies in mammalian tissues have shown that proteins belonging to the α-ketoglutarate dehydrogenase complex located in the mitochondrial matrix are also a source of ROS in a mechanism stimulated by the low concentration of NAD<sup>+</sup> [32, 33]. In Saccharomyces cerevisiae, the deletion of the LPD1 gene, which leads to the inactivation of the enzyme dihydrolipoyl dehydrogenase, E3 subunit of the pyruvate dehydrogenase complex, also leads to a decrease in ROS production. This finding shows the importance of other mitochondrial proteins, other than those associated with the electron

ubiquinone and back to complex I [31].

complex III; IV, complex IV or cytochrome c oxidase; O2

mitochondrial MnSOD; GPx, glutathione peroxidase. Modified from Ref. [22].

transport chain, in the regulation of redox balance [34].

#### 1.1. Reactive species in mitochondria

The ROS comprise a variety of molecules derived from molecular oxygen, including oxygen radicals and non-radical oxygen derivate. The major intracellular site of ROS formation in most tissues is mitochondria [23, 24]. Within mitochondria, the electron transport chain continuously generates water from O2 through the electronic reduction at the cytochrome c oxidase level (Figure 1). These electrons reach cytochrome c oxidase by sequential transfer from the reduction of other components, and are initially removed from NADH and FADH2. During this transfer, a small amount of electrons are lost at intermediate stages in the electron transport chain, mainly in the complex I and complex III [25–27] in mammals, leading to a monoelectronic reduction of O2 [28].

This monoelectronic reduction of O2 results in the formation of anion superoxide radical. While complex I releases superoxide only in the mitochondrial matrix, complex III releases

1. Cellular respiration and generation of reactive species in the

Oxidative phosphorylation is the center of energy metabolism in plants, animals and several microbial life forms [1]. In eukaryotes, this process occurs in mitochondria. The mitochondria is a cytoplasmic organelle surrounded by two membranes, outer and inner membrane, which main function is the production of most of the phosphate compounds necessary for the energetic balance of the cell. In addition, other functions such as the regulation of the body's heat generation [2–4] programmed cell death [5–7], reactive oxygen species (ROS) generation and cell signaling [8] is also associated with mitochondria. Cellular vitality is directly related to mitochondria, and mitochondrial dysfunctions are frequent causes of accidental cell death [5, 9–11],

cancer [12, 13], diabetes [14–16] and neurodegenerative diseases [17–19], among others.

port chain, as complex I and complex II, i.e., delivering electron to ubiquinone [21].

production of ATP by ATP synthase (Figure 1) [22].

1.1. Reactive species in mitochondria

monoelectronic reduction of O2 [28].

The redox carriers within the respiratory chain consists of flavoprotein containing tightly bound FAD or FMN as prosthetic groups, protein-bound couper, ironsulphur (nonhaem iron) proteins and cytochromes, with haem prosthetic groups. The ubiquinone also participated in electron transport chain as a free and diffusible cofactor [20]. While electron transport occurs through the mitochondrial complexes, complexes I, II, and III pump protons from mitochondrial matrix to the intermembrane space. The energy associated to this process is used to the

The ROS comprise a variety of molecules derived from molecular oxygen, including oxygen radicals and non-radical oxygen derivate. The major intracellular site of ROS formation in most tissues is mitochondria [23, 24]. Within mitochondria, the electron transport chain continuously generates water from O2 through the electronic reduction at the cytochrome c oxidase level (Figure 1). These electrons reach cytochrome c oxidase by sequential transfer from the reduction of other components, and are initially removed from NADH and FADH2. During this transfer, a small amount of electrons are lost at intermediate stages in the electron transport chain, mainly in the complex I and complex III [25–27] in mammals, leading to a

This monoelectronic reduction of O2 results in the formation of anion superoxide radical. While complex I releases superoxide only in the mitochondrial matrix, complex III releases

The characterization of the respiratory electron chain could be performed in studies using the fractionation of its components by certain detergents that at low concentrations break the interactions between proteins and lipids in the membranes, leaving associations between proteins intact [20]. In electron transport chain, through this process, four protein complexes were found. They were named complex I (or NADH-Ubiquinone oxidoreductase), complex II (succinate dehydrogenase), complex III (Ubiquinol -cytochrome c oxidoreductase, or complex bc1) e complex IV (cytochrome c oxidase). The complex V is also known as ATP synthase. Despite glycerophosphate dehydrogenase (glycerol-3-phosphate dehydrogenase) and ETF–ubiquinone oxidoreductase have not complex nomenclature, they are connect to the electron trans-

mitochondria: implications in cell viability and aging

2 Novel Prospects in Oxidative and Nitrosative Stress

Figure 1. Electron transport chain, ROS, and antioxidant defense. The electron transport chain receives electrons from reduced compounds, as NADH in complex I (also called NADH coenzyme Q reductase) and succinate or FADH2 in complex II (succinate dehydrogenase) and transfers them successively to coenzyme Q or ubiquinone, complex III, complex IV and finally to molecular oxygen with the formation of water. Concomitant with electron transport, protons are transferred from the mitochondrial matrix to the intermembrane space by complexes I (in mammals, but not in yeasts), complex III and IV. The difference in electrochemical potential between intermembrane space and matrix is used by ATP synthase to produce ATP from ADP and inorganic phosphate. During the passage of electrons through the complexes, a small fraction is leaked to oxygen at intermediate points, producing the superoxide radical anion, which in the mitochondrial matrix can be removed by SOD2 forming H2O2. Hydrogen peroxide can be converted to water in the mitochondrial matrix by glutathione peroxidase (mammals). Abbreviations: ADP, adenosine diphosphate; ATP, adenosine triphosphate; UQ, ubiquinone; Cyt, cytochrome c; SDH, succinate dehydrogenase; I, complex I or NADH coenzyme Q reductase; II, complex II or succinate dehydrogenase; III, complex III; IV, complex IV or cytochrome c oxidase; O2 •, superoxide radical anion; SOD2, superoxide dismutase 2 or mitochondrial MnSOD; GPx, glutathione peroxidase. Modified from Ref. [22].

superoxide in both sides of inner mitochondrial membrane [29]. Complex II could theoretically generate superoxide, due presence of flavoprotein in its structure. However, the redox centers are arranged in a manner that aids the prevention of ROS by avoiding the access of O2 to the flavoprotein. This may explain the reason why this complex does not show a ROS formation by itself [30], but only due reverse electron transfer, i.e., when electrons flow from succinate to ubiquinone and back to complex I [31].

In addition to the electron transport chain, recent studies in mammalian tissues have shown that proteins belonging to the α-ketoglutarate dehydrogenase complex located in the mitochondrial matrix are also a source of ROS in a mechanism stimulated by the low concentration of NAD<sup>+</sup> [32, 33]. In Saccharomyces cerevisiae, the deletion of the LPD1 gene, which leads to the inactivation of the enzyme dihydrolipoyl dehydrogenase, E3 subunit of the pyruvate dehydrogenase complex, also leads to a decrease in ROS production. This finding shows the importance of other mitochondrial proteins, other than those associated with the electron transport chain, in the regulation of redox balance [34].

The term reactive specie is not restricted to oxygen, but is also include others, as reactive nitrogen (RNS). Nitric oxide is a membrane permeable free radical that participates in a multiple process in the cells as signaling molecule, but also can contribute in cell oxidative damage. Its effect depends on NO levels and localization in the cell microenvironment [35, 36]. When nitric oxide is present in environment, as in mitochondrial matrix, the reaction of this free radical with superoxide can form others RNS, as peroxynitrite.

the lipids, proteins and nucleic acids. In other words, when the generation of reactive species exceeds antioxidant capacity, the cellular macromolecules also become targets of oxidation by these species. The possible consequences originated from this extensive oxidation, including an increased risk for cardiovascular disease, cancer and neurodegenerative disease (as

Under oxidative stress conditions, proteins suffer extensive modification [62–65]. Basically, ROS can oxidize amino acids cysteine and methionine, resulting in the production of dithiol and methionine sulfoxide crosslinks, respectively [66]. Moreover, reactive species also can cause protein modification by nitration of tyrosine and by nitrosation of amino acids with thiol group. These changes often result in the alteration of function or inhibition of enzyme activities. The protein adducts have been observed in several pathologic conditions [67, 68], suggesting their deleterious effects. However, whether these endogenous modifications are produced in a controlled manner, they may also control physiological responses [69, 70].

It is important to stress that the presence of proteins containing nitrotyrosine residues, for example, has been a biomarker of damage by reactive species [67, 68]. The tyrosine nitration occurs by addition of NO2 to the ortho position of the phenolic ring of this amino acid. In fact, this NO2 group is obtained from peroxynitrite (ONOO), a very strong oxidant [71]. During

the tyrosine residues, can damage several classes of molecules. ONOO, its protonated form peroxynitrous acid (ONOOH), and its secondary radical product, react with electron-rich groups, such as sulfhydryls, ironsulphur centers, zinc-thiolates and active site sulfhydryl in

The thiol group (-SH) of cysteine, for example, it is another relevant protein targets of ROS. Disulfide bond is important in protein structure and function [74], and recently its role in redox signaling has also been evidenced [75]. The reaction of H2O2 with the deprotonated thiol group of cysteine produces a sulfenic acid (R-SOH). This last may be oxidized again producing a sulfinic acid (R-SO2H). With high levels of stress oxidative, cysteines can further be oxidized to a sulfonic acid (R-SO3H) [70, 76]. While sulfenic and sulfinic acids can be enzymatically reversible by the glutathione and thioredoxin enzyme systems [77] (Details about antioxidant mechanisms in next section), the sulfonic acid in cysteine residues seems to represent an

The reactive species react directly with nucleic acids producing oxidative damage. Since oxidative DNA damage is a major threat to genetic integrity, causing mutations and modifications in gene expression pattern, it has been implicated in a wide variety of diseases, including cancer, cardiovascular and neurodegeneration disease, as well as aging process [46, 73].

• reacts

• and, beyond

Oxidative Stress: Noxious but Also Vital http://dx.doi.org/10.5772/intechopen.73394 5

oxidative stress conditions, especially in inflammatory processes, a proportion of O2

with NO to form ONOO. This last is a much more powerful oxidant than O2

detailed in Section 4).

2.1. Protein adducts

tyrosine phosphatases [67, 68, 72, 73].

irreversible protein damage.

2.2. DNA oxidation

Besides mitochondria electron chain and enzyme linked to mitochondrial dehydrogenase complexes, other sources of ROS in cells include enzymes, as NADPH oxidases, cytochrome P450, cyclooxygenases, and the system xanthine/xanthine oxidase. Autoxidation is another example of source of ROS that in cells occurs when a biochemical compound is exposure to O2, as it occurs in FADH2, L-DOPA and in nitric oxide synthase with generation of superoxide. The auto oxidation can be catalyzed by metallic ions, finally, harm proteins, in which O2 bind Fe2+ could lead to superoxide, as in hemoglobin [37].

### 1.2. Mitochondria and reactive species: physiological level, oxidative stress, and its implications

ROS and RNS are normally produced in metabolism and have an important role as signaling molecules regulating diverse physiological cell events, as cell signaling, metabolism and regulation of transcription factors [35, 38–42].

The steady state of reactive species will depend on their generation, reactivity and removal by antioxidant defenses. When the level of reactive species generation is much larger than their removal it is said that there is a condition called oxidative stress, i.e., an imbalance between reactive species and antioxidants in favor of reactive species. The maintenance of cell redox state is important to cell viability [43]. The increased level of reactive species can lead to oxidative damage to a vast number of biological molecules, as DNA [44–46], proteins [47], lipids [48], including membranes [3] leading to a range of pathologies, as cancer [36], neurological disease [49], cardiac disease [50, 51], inflammation process [52] and aging.

There is a grand amount of theories about aging process, at least 300 theories according Medvedev [53]. In 1956, Harman proposed in his "free radical theory of aging" that the damage of biomolecules that occurs during aging is due oxidative stress, ROS increments [54]. Mitochondria, as the major site of ROS production, have been associated with aging process [55, 56]. Moreover, studies with caloric restriction in yeast and mammals have shown that the mitochondria, ROS, and RNS have an important role in the aging process [34, 56–61].

### 2. Protein adducts, DNA oxidation and epigenetic regulation, and effects on biological membranes

During oxidative stress, ROS can attack molecules at electron-dense sites or abstract protons, producing secondary radical species, which undergo conformational change generating more stable products. The molecules that are vulnerable to these deleterious modifications include the lipids, proteins and nucleic acids. In other words, when the generation of reactive species exceeds antioxidant capacity, the cellular macromolecules also become targets of oxidation by these species. The possible consequences originated from this extensive oxidation, including an increased risk for cardiovascular disease, cancer and neurodegenerative disease (as detailed in Section 4).

#### 2.1. Protein adducts

The term reactive specie is not restricted to oxygen, but is also include others, as reactive nitrogen (RNS). Nitric oxide is a membrane permeable free radical that participates in a multiple process in the cells as signaling molecule, but also can contribute in cell oxidative damage. Its effect depends on NO levels and localization in the cell microenvironment [35, 36]. When nitric oxide is present in environment, as in mitochondrial matrix, the reaction of this

Besides mitochondria electron chain and enzyme linked to mitochondrial dehydrogenase complexes, other sources of ROS in cells include enzymes, as NADPH oxidases, cytochrome P450, cyclooxygenases, and the system xanthine/xanthine oxidase. Autoxidation is another example of source of ROS that in cells occurs when a biochemical compound is exposure to O2, as it occurs in FADH2, L-DOPA and in nitric oxide synthase with generation of superoxide. The auto oxidation can be catalyzed by metallic ions, finally, harm proteins, in which O2 bind

1.2. Mitochondria and reactive species: physiological level, oxidative stress, and its

logical disease [49], cardiac disease [50, 51], inflammation process [52] and aging.

ROS and RNS are normally produced in metabolism and have an important role as signaling molecules regulating diverse physiological cell events, as cell signaling, metabolism and regu-

The steady state of reactive species will depend on their generation, reactivity and removal by antioxidant defenses. When the level of reactive species generation is much larger than their removal it is said that there is a condition called oxidative stress, i.e., an imbalance between reactive species and antioxidants in favor of reactive species. The maintenance of cell redox state is important to cell viability [43]. The increased level of reactive species can lead to oxidative damage to a vast number of biological molecules, as DNA [44–46], proteins [47], lipids [48], including membranes [3] leading to a range of pathologies, as cancer [36], neuro-

There is a grand amount of theories about aging process, at least 300 theories according Medvedev [53]. In 1956, Harman proposed in his "free radical theory of aging" that the damage of biomolecules that occurs during aging is due oxidative stress, ROS increments [54]. Mitochondria, as the major site of ROS production, have been associated with aging process [55, 56]. Moreover, studies with caloric restriction in yeast and mammals have shown that the mitochondria, ROS, and RNS have an important role in the aging process [34, 56–61].

2. Protein adducts, DNA oxidation and epigenetic regulation, and effects

During oxidative stress, ROS can attack molecules at electron-dense sites or abstract protons, producing secondary radical species, which undergo conformational change generating more stable products. The molecules that are vulnerable to these deleterious modifications include

free radical with superoxide can form others RNS, as peroxynitrite.

Fe2+ could lead to superoxide, as in hemoglobin [37].

lation of transcription factors [35, 38–42].

4 Novel Prospects in Oxidative and Nitrosative Stress

on biological membranes

implications

Under oxidative stress conditions, proteins suffer extensive modification [62–65]. Basically, ROS can oxidize amino acids cysteine and methionine, resulting in the production of dithiol and methionine sulfoxide crosslinks, respectively [66]. Moreover, reactive species also can cause protein modification by nitration of tyrosine and by nitrosation of amino acids with thiol group. These changes often result in the alteration of function or inhibition of enzyme activities. The protein adducts have been observed in several pathologic conditions [67, 68], suggesting their deleterious effects. However, whether these endogenous modifications are produced in a controlled manner, they may also control physiological responses [69, 70].

It is important to stress that the presence of proteins containing nitrotyrosine residues, for example, has been a biomarker of damage by reactive species [67, 68]. The tyrosine nitration occurs by addition of NO2 to the ortho position of the phenolic ring of this amino acid. In fact, this NO2 group is obtained from peroxynitrite (ONOO), a very strong oxidant [71]. During oxidative stress conditions, especially in inflammatory processes, a proportion of O2 • reacts with NO to form ONOO. This last is a much more powerful oxidant than O2 • and, beyond the tyrosine residues, can damage several classes of molecules. ONOO, its protonated form peroxynitrous acid (ONOOH), and its secondary radical product, react with electron-rich groups, such as sulfhydryls, ironsulphur centers, zinc-thiolates and active site sulfhydryl in tyrosine phosphatases [67, 68, 72, 73].

The thiol group (-SH) of cysteine, for example, it is another relevant protein targets of ROS. Disulfide bond is important in protein structure and function [74], and recently its role in redox signaling has also been evidenced [75]. The reaction of H2O2 with the deprotonated thiol group of cysteine produces a sulfenic acid (R-SOH). This last may be oxidized again producing a sulfinic acid (R-SO2H). With high levels of stress oxidative, cysteines can further be oxidized to a sulfonic acid (R-SO3H) [70, 76]. While sulfenic and sulfinic acids can be enzymatically reversible by the glutathione and thioredoxin enzyme systems [77] (Details about antioxidant mechanisms in next section), the sulfonic acid in cysteine residues seems to represent an irreversible protein damage.

#### 2.2. DNA oxidation

The reactive species react directly with nucleic acids producing oxidative damage. Since oxidative DNA damage is a major threat to genetic integrity, causing mutations and modifications in gene expression pattern, it has been implicated in a wide variety of diseases, including cancer, cardiovascular and neurodegeneration disease, as well as aging process [46, 73].

The nitrogenous bases as well as the sugar suffer radical attacks, causing several base alterations and strand breaks [78]. In fact, around 80 different bases have been observed in DNA exposed to oxidants [79]. In this context, •OH is the most important reactive species that attacks DNA, since it reacts with the four bases and sugar moiety of the DNA backbone [78, 80] with a reaction rate limited by diffusion (4.5 <sup>10</sup><sup>9</sup> to 9 109 <sup>M</sup><sup>1</sup> s 1 ) [79]. •OH attacks carbo-carbon double bonds of bases due to the high electron density. These attacks produce the hydroxylation at C5 and C6 of pyrimidines and C4, C5 and C8 of purines [78, 80]. These secondary radicals are subjected to other oxidation and reduction reactions, producing a wide DNA lesions, including the well characterized derivatives, 7,8-dihydro-8-oxodeoxyguanine (8-oxoG) and 2,6-diamino-4-hydroxy-5-formamido-pyrimidine (FapyGua) [71]. 8-oxo-G is the most stable of these altered bases and can give rise to mutations due to insert Adenine (A) opposite 8-oxo-G during DNA replication, instead of the Cytosine (C) [46, 71].

of peroxidation can inhibit the activity of protein transporters and ion channels [89, 91]. The increase of the permeability also seems to occur in internal mitochondrial membrane, uncoupling respiratory-chain phosphorylation [93]. Finally, the lipid peroxidation leads the severe damages: modification of membrane permeability, enzymatic inhibitions, inac-

Oxidative Stress: Noxious but Also Vital http://dx.doi.org/10.5772/intechopen.73394 7

The exposure cells and tissues to the harmful effects of free radicals cause a cascade of reactions and induces activation of some strategies to damage prevent, repair mechanism to alleviate the oxidative damages, physical protection mechanism against damage, and the final

The antioxidant defenses are the first line of choice to take care of the stress. Endogenous antioxidant defenses include antioxidant enzymes and non-enzymatic molecules that are usually distributed within the cytoplasm and various cell organelles [94]. The exogenous antioxidants are present in consumed fruits, vegetables, juice, tea, coffee, nuts and cereal products [95].

The concept of biological antioxidant refers to any compound present at a lower concentration which is able to either delay or prevent the oxidation of the substrate. Antioxidant functions imply lowering oxidative stress, DNA mutations, malignant transformations, as well as other parameters of cell damage [96]. Antioxidants reactions can deplete molecular oxygen or decreasing its local concentration, removing pro oxidative metal ions, trapping aggressive ROS such as superoxide anion radical or hydrogen peroxide, scavenging chain initiating radicals like hydroxyl OH, alkoxyl RO or peroxyl ROO, breaking the chain of a radical

O2) [97].

The antioxidants include some high molecular weight (SOD, GPx, catalase, albumin, transferrin, and metallothionein) and some low molecular weight substances (uric acid, ascorbic acid, lipoic acid, glutathione, ubiquinol, tocopherol/vitamin E, flavonoids). Natural food-derived components have received great attention in the last 2 decades, and several biological activities showing promising anti-inflammatory, antioxidant, and anti-apoptotic-modulatory potential have been identified. These enzymatic and nonenzymatic antioxidant systems are necessary for sustaining life by maintaining a delicate intracellular redox balance and minimizing unde-

Antioxidant enzymes catalyze ROS conversion directly via an active-site metal ion or through pathways involving the donation of an electron from the moiety-conserved redox couples thioredoxin and glutathione, which require continuous regeneration of the reduced species [99]. Superoxide and H2O2 metabolizing enzymes are generally considered to be the primary

tivation of transporters [37, 92].

sequence or quenching singlet oxygen (<sup>1</sup>

sirable cellular damage caused by ROS [94, 97, 98].

antioxidant enzyme defense system in the body [98].

3.1. Enzymatic antioxidant system

3. Endogenous/exogenous defense mechanisms

most important is the antioxidant defense mechanisms [94, 95].

Another mutation produced by oxidative damage is C to thymine (T) transition, mainly due to the cytosine-derived products uracil glycol and 5-hydroxyuracil mispairing with A, instead of the G [71]. Although other pathways also induce this mutation, it is important to stress that C to T transition is the most frequent mutations found in cancers and in the tumor suppressor gene p53 [81, 82].

#### 2.3. Effects on biological membranes

Under conditions of oxidative stress occur an oxidative process termed lipid peroxidation that affects lipids containing multiple double bonds, such as fatty acids, phospholipids, glycolipids and cholesterol, modifying properties of cellular membranes [73, 83]. This degenerative process is believed to contribute to aging and several diseases, such as atherosclerosis, Alzheimer's disease, peptic ulcer disease, and cancer [84, 85].

Cellular membranes are especially vulnerable to lipid peroxidation not only because of their high levels of unsaturated fatty acids, but also because of their connection with molecules capable of producing reactive species. They attack mainly the unsaturated fatty acids which contain carbon-carbon double bonds and CH2 groups with particularly reactive hydrogen, and start radical peroxidation chain reactions [86]. These chain reactions are going to terminate when primary or secondary radicals directly react. Lipid peroxidation is accelerated by the presence of Fe2+ and Cu2+ ions [87, 88]. It is important to stress that lipid peroxides are unstable derivatives from the oxidation of unsaturated fatty acids and decompose to form reactive carbonyl molecules, such as malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE) [85, 89]. These two products are abundant biomarkers of lipid peroxidation [85, 90].

Membrane-bound proteins are also involved in the process of lipid peroxidation. Aldehyde products, such as MDA and 4-HNE, react with amine and thiol groups of membrane protein, causing several damages, including inactivation of enzymes. Conformational changes of membrane molecules also include lipid–lipid cross-links and lipid–protein cross-links [91, 92].

Moreover, lipid peroxidation modifies the global biophysical properties of the membranes. This process affects the packing of lipids and the permeability to solutes, which in turn, changes its function, including the membrane potential. Furthermore, the process of peroxidation can inhibit the activity of protein transporters and ion channels [89, 91]. The increase of the permeability also seems to occur in internal mitochondrial membrane, uncoupling respiratory-chain phosphorylation [93]. Finally, the lipid peroxidation leads the severe damages: modification of membrane permeability, enzymatic inhibitions, inactivation of transporters [37, 92].

### 3. Endogenous/exogenous defense mechanisms

The nitrogenous bases as well as the sugar suffer radical attacks, causing several base alterations and strand breaks [78]. In fact, around 80 different bases have been observed in DNA exposed to oxidants [79]. In this context, •OH is the most important reactive species that attacks DNA, since it reacts with the four bases and sugar moiety of the DNA backbone [78,

carbo-carbon double bonds of bases due to the high electron density. These attacks produce the hydroxylation at C5 and C6 of pyrimidines and C4, C5 and C8 of purines [78, 80]. These secondary radicals are subjected to other oxidation and reduction reactions, producing a wide DNA lesions, including the well characterized derivatives, 7,8-dihydro-8-oxodeoxyguanine (8-oxoG) and 2,6-diamino-4-hydroxy-5-formamido-pyrimidine (FapyGua) [71]. 8-oxo-G is the most stable of these altered bases and can give rise to mutations due to insert Adenine (A)

Another mutation produced by oxidative damage is C to thymine (T) transition, mainly due to the cytosine-derived products uracil glycol and 5-hydroxyuracil mispairing with A, instead of the G [71]. Although other pathways also induce this mutation, it is important to stress that C to T transition is the most frequent mutations found in cancers and in the tumor suppressor

Under conditions of oxidative stress occur an oxidative process termed lipid peroxidation that affects lipids containing multiple double bonds, such as fatty acids, phospholipids, glycolipids and cholesterol, modifying properties of cellular membranes [73, 83]. This degenerative process is believed to contribute to aging and several diseases, such as atherosclerosis,

Cellular membranes are especially vulnerable to lipid peroxidation not only because of their high levels of unsaturated fatty acids, but also because of their connection with molecules capable of producing reactive species. They attack mainly the unsaturated fatty acids which contain carbon-carbon double bonds and CH2 groups with particularly reactive hydrogen, and start radical peroxidation chain reactions [86]. These chain reactions are going to terminate when primary or secondary radicals directly react. Lipid peroxidation is accelerated by the presence of Fe2+ and Cu2+ ions [87, 88]. It is important to stress that lipid peroxides are unstable derivatives from the oxidation of unsaturated fatty acids and decompose to form reactive carbonyl molecules, such as malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE)

[85, 89]. These two products are abundant biomarkers of lipid peroxidation [85, 90].

Membrane-bound proteins are also involved in the process of lipid peroxidation. Aldehyde products, such as MDA and 4-HNE, react with amine and thiol groups of membrane protein, causing several damages, including inactivation of enzymes. Conformational changes of membrane molecules also include lipid–lipid cross-links and lipid–protein cross-links [91, 92].

Moreover, lipid peroxidation modifies the global biophysical properties of the membranes. This process affects the packing of lipids and the permeability to solutes, which in turn, changes its function, including the membrane potential. Furthermore, the process

s 1

) [79]. •OH attacks

80] with a reaction rate limited by diffusion (4.5 <sup>10</sup><sup>9</sup> to 9 109 <sup>M</sup><sup>1</sup>

opposite 8-oxo-G during DNA replication, instead of the Cytosine (C) [46, 71].

gene p53 [81, 82].

2.3. Effects on biological membranes

6 Novel Prospects in Oxidative and Nitrosative Stress

Alzheimer's disease, peptic ulcer disease, and cancer [84, 85].

The exposure cells and tissues to the harmful effects of free radicals cause a cascade of reactions and induces activation of some strategies to damage prevent, repair mechanism to alleviate the oxidative damages, physical protection mechanism against damage, and the final most important is the antioxidant defense mechanisms [94, 95].

The antioxidant defenses are the first line of choice to take care of the stress. Endogenous antioxidant defenses include antioxidant enzymes and non-enzymatic molecules that are usually distributed within the cytoplasm and various cell organelles [94]. The exogenous antioxidants are present in consumed fruits, vegetables, juice, tea, coffee, nuts and cereal products [95].

The concept of biological antioxidant refers to any compound present at a lower concentration which is able to either delay or prevent the oxidation of the substrate. Antioxidant functions imply lowering oxidative stress, DNA mutations, malignant transformations, as well as other parameters of cell damage [96]. Antioxidants reactions can deplete molecular oxygen or decreasing its local concentration, removing pro oxidative metal ions, trapping aggressive ROS such as superoxide anion radical or hydrogen peroxide, scavenging chain initiating radicals like hydroxyl OH, alkoxyl RO or peroxyl ROO, breaking the chain of a radical sequence or quenching singlet oxygen (<sup>1</sup> O2) [97].

The antioxidants include some high molecular weight (SOD, GPx, catalase, albumin, transferrin, and metallothionein) and some low molecular weight substances (uric acid, ascorbic acid, lipoic acid, glutathione, ubiquinol, tocopherol/vitamin E, flavonoids). Natural food-derived components have received great attention in the last 2 decades, and several biological activities showing promising anti-inflammatory, antioxidant, and anti-apoptotic-modulatory potential have been identified. These enzymatic and nonenzymatic antioxidant systems are necessary for sustaining life by maintaining a delicate intracellular redox balance and minimizing undesirable cellular damage caused by ROS [94, 97, 98].

#### 3.1. Enzymatic antioxidant system

Antioxidant enzymes catalyze ROS conversion directly via an active-site metal ion or through pathways involving the donation of an electron from the moiety-conserved redox couples thioredoxin and glutathione, which require continuous regeneration of the reduced species [99]. Superoxide and H2O2 metabolizing enzymes are generally considered to be the primary antioxidant enzyme defense system in the body [98].

The SOD is a family of enzymes catalyzing dismutation of superoxide into oxygen and H2O2. Three types of superoxide dismutases can be encountered in mammalian tissues: copper-zinc containing superoxide dismutase (SOD1) present in the cytosol, manganese containing superoxide dismutase (SOD2) found in the mitochondrial matrix and extracellular superoxide dismutase (SOD3). All three are highly expressed, mainly in the renal tubules of healthy kidneys [15, 98, 100]. The final product of the SOD activity - H2O2, is then converted into water and oxygen by the catalase (CAT). This enzyme is a homotetrameric protein containing four iron heme and largely located in the peroxisomes [15, 100].

The flavonoid group is the most diverse within phenolic compounds, with two aromatic rings associated via C-C bonds by a 3C oxygenated heterocycle. Flavonoids have antioxidant and chelating properties, inactivate ROS, acting against the oxidation of low density lipoproteins (LDL) and improving inflammation of the blood vessels. They also reduce the activity of the xanthine oxidase enzymes and the nicotinamide adenine dinucleotide phosphate oxidase,

In cellular compartments, flavonoids function as antioxidants inactivating free radicals both in hydrophilic and lipophilic compartments. For example, the antioxidant activity of phenolic compounds present in spices (cinnamon, sweet weed and mustard) differs between aqueous

Vitamins C and E act together to inhibit lipid peroxidation and protect the cell against oxidative damage, as DNA damage. The antioxidant activity of vitamin C involves the transfer of an electron to the free radical and the consequent formation of the radical ascorbate [109]. In addition, vitamin C acts synergistically with vitamin E, which regenerate the vitamin C has better antioxidant activity in hydrophilic media, and in aqueous phase of extracellular fluids, it is able to neutralize ROS in the aqueous phase before they can attack lipids. Vitamin E is an important fat soluble antioxidant, acting as the chain breaking antioxidant within the cell membrane and playing an important role in the protection of

Vitamins C and E inhibit lipid peroxidation and protect against oxidative damage by their scavenging actions of ROS, as well as by modulateing numerous enzymatic complexes involved in the production of ROS, endothelial function and aggregation of platelets. These vitamins can

It has been reported that ascorbic acid and α-tocopherol, derivated from vitamin C and E

The most common carotenoids are xanthophylls and carotenes. Carotenoids can neutralize singlet oxygen by quenching it or can break the chain reaction of free radicals, or scavenging it, not so effective action (scavenging). The structure of the free radical is the main factor that determines if the carotenoid will have quenching or scavenging action. It also depends on the region where the radical is in heterogeneous biological tissue, aqueous or lipid region (plasma, blood, heart, liver, brain etc.), and the structure of the carotenoids (number of conjugated,

The physical quenching is the transfer of excitation energy from the singlet oxygen to the carotenoid. The oxygen returns to ground state and the carotenoid is in the excited triplet state, the energy is dissipated producing stable carotenoid and thermal energy and the carot-

The chemical quenching the carotenoid combines with oxygen or is oxidized, leading to its destruction and producing a variety of oxidized products. Carotenoids can also extinguish the triplet-excited state of chlorophyll or other excited sensitizers, thus preventing the formation of singlet oxygen [112]. The free radical scavenging can occur in three ways, by electron transfer,

respectively, may involved in the transcriptional modulation of NADPH oxidase [111].

cyclic or acyclic double bonds), polar or nonpolar groups, redox properties [112–114].

enoid can undergo other cycles of singlet oxygen quenching [112, 115].

by hydrogen abstraction, and by addition [112, 116].

• in the cardiovascular system.

Oxidative Stress: Noxious but Also Vital http://dx.doi.org/10.5772/intechopen.73394 9

enzymes that stimulate the production of ROS [107].

membrane fatty acids against lipid peroxidation [110].

also regulate NADPH oxidase, the most important source of O2

and lipid systems [108].

Other important enzymatic antioxidants in the first line of defense include glutathione peroxidase (GPX) and myeloperoxidase (MPO) enzymes. The GPX is a selenium-containing enzyme, catalyzes both the reduction of H2O2, and organic hydroperoxides to water or corresponding alcohols. Reduced glutathione functions as effective electron donor in the process, as free thiol groups are oxidized to disulfide bonds: H2O2 + 2GSH ! GS-SG + 2H2O [97]. The MPO, a heme peroxidase, abundant in granules of human inflammatory cells, catalyzes the conversion of H2O2 to HClO with the production of ROS. The ROS production is associated with cardiovascular disease, chronic obstructive pulmonary disease, and Alzheimer's disease. Oxidant species derived from MPO lead to the production of specific oxidation products, such as 3-Cl-Tyr. This can be used as biomarker in several diseases, as above described, and its levels correlate with MPO [100].

Other enzymes could be cited by our antioxidant activity, such as Peroxiredoxin Family (PRX). These enzymes are a family of abundantly present 20–30 kDa peroxidases that are excessively reactive with H2O2. So, they are likely to be critical for both oxidative stress protection as well as redox signaling [98]. The antioxidant enzymes may possibly offer novel treatment options for redox-related diseases, provided that the molecular mechanisms are known and can be specifically targeted. Besides that, inhibiting a given antioxidant enzyme or specifically silencing its gene expression may help treat disorders related to a gain of enzymatic function [98] and this fact can will help the researchers to explore future options in enzymatic antioxidant system and diseases.

#### 3.2. Nonenzymatic antioxidant systems

Among the nonenzymatic antioxidant compounds, the principals are obtained from dietary as the class of phenolic compounds, vitamins C and E, and carotenoids [101]. Phenolic compounds represent a large group of secondary metabolites [102], among them flavonoids, phenolic acids, tannins and tocopherols as the most common natural source phenolic antioxidants [103].

The phenolic compounds are composed of one or more aromatic rings with varying degrees of hydroxylation, methoxylation and glycosylation, and various studies have associated the structure of phenolic compounds with their antioxidant properties [102, 104]. The antioxidant activity generally increases with the degree of hydroxylation in aromatic rings and decreases with C-3 methoxylation [105, 106]. The antioxidant activity is based on the availability of electrons to neutralize the free radicals; in addition, it is related to the number and nature of the hydroxylation pattern in the aromatic ring and the ability to act as a hydrogen donor [106]. The flavonoid group is the most diverse within phenolic compounds, with two aromatic rings associated via C-C bonds by a 3C oxygenated heterocycle. Flavonoids have antioxidant and chelating properties, inactivate ROS, acting against the oxidation of low density lipoproteins (LDL) and improving inflammation of the blood vessels. They also reduce the activity of the xanthine oxidase enzymes and the nicotinamide adenine dinucleotide phosphate oxidase, enzymes that stimulate the production of ROS [107].

The SOD is a family of enzymes catalyzing dismutation of superoxide into oxygen and H2O2. Three types of superoxide dismutases can be encountered in mammalian tissues: copper-zinc containing superoxide dismutase (SOD1) present in the cytosol, manganese containing superoxide dismutase (SOD2) found in the mitochondrial matrix and extracellular superoxide dismutase (SOD3). All three are highly expressed, mainly in the renal tubules of healthy kidneys [15, 98, 100]. The final product of the SOD activity - H2O2, is then converted into water and oxygen by the catalase (CAT). This enzyme is a homotetrameric protein containing four

Other important enzymatic antioxidants in the first line of defense include glutathione peroxidase (GPX) and myeloperoxidase (MPO) enzymes. The GPX is a selenium-containing enzyme, catalyzes both the reduction of H2O2, and organic hydroperoxides to water or corresponding alcohols. Reduced glutathione functions as effective electron donor in the process, as free thiol groups are oxidized to disulfide bonds: H2O2 + 2GSH ! GS-SG + 2H2O [97]. The MPO, a heme peroxidase, abundant in granules of human inflammatory cells, catalyzes the conversion of H2O2 to HClO with the production of ROS. The ROS production is associated with cardiovascular disease, chronic obstructive pulmonary disease, and Alzheimer's disease. Oxidant species derived from MPO lead to the production of specific oxidation products, such as 3-Cl-Tyr. This can be used as biomarker in several diseases, as above described, and its levels correlate

Other enzymes could be cited by our antioxidant activity, such as Peroxiredoxin Family (PRX). These enzymes are a family of abundantly present 20–30 kDa peroxidases that are excessively reactive with H2O2. So, they are likely to be critical for both oxidative stress protection as well as redox signaling [98]. The antioxidant enzymes may possibly offer novel treatment options for redox-related diseases, provided that the molecular mechanisms are known and can be specifically targeted. Besides that, inhibiting a given antioxidant enzyme or specifically silencing its gene expression may help treat disorders related to a gain of enzymatic function [98] and this fact can will help the researchers to explore future options in enzymatic antioxidant

Among the nonenzymatic antioxidant compounds, the principals are obtained from dietary as the class of phenolic compounds, vitamins C and E, and carotenoids [101]. Phenolic compounds represent a large group of secondary metabolites [102], among them flavonoids, phenolic acids, tannins and tocopherols as the most common natural source phenolic antioxidants [103].

The phenolic compounds are composed of one or more aromatic rings with varying degrees of hydroxylation, methoxylation and glycosylation, and various studies have associated the structure of phenolic compounds with their antioxidant properties [102, 104]. The antioxidant activity generally increases with the degree of hydroxylation in aromatic rings and decreases with C-3 methoxylation [105, 106]. The antioxidant activity is based on the availability of electrons to neutralize the free radicals; in addition, it is related to the number and nature of the hydroxylation pattern in the aromatic ring and the ability to act as a hydrogen donor [106].

iron heme and largely located in the peroxisomes [15, 100].

8 Novel Prospects in Oxidative and Nitrosative Stress

with MPO [100].

system and diseases.

3.2. Nonenzymatic antioxidant systems

In cellular compartments, flavonoids function as antioxidants inactivating free radicals both in hydrophilic and lipophilic compartments. For example, the antioxidant activity of phenolic compounds present in spices (cinnamon, sweet weed and mustard) differs between aqueous and lipid systems [108].

Vitamins C and E act together to inhibit lipid peroxidation and protect the cell against oxidative damage, as DNA damage. The antioxidant activity of vitamin C involves the transfer of an electron to the free radical and the consequent formation of the radical ascorbate [109]. In addition, vitamin C acts synergistically with vitamin E, which regenerate the vitamin C has better antioxidant activity in hydrophilic media, and in aqueous phase of extracellular fluids, it is able to neutralize ROS in the aqueous phase before they can attack lipids. Vitamin E is an important fat soluble antioxidant, acting as the chain breaking antioxidant within the cell membrane and playing an important role in the protection of membrane fatty acids against lipid peroxidation [110].

Vitamins C and E inhibit lipid peroxidation and protect against oxidative damage by their scavenging actions of ROS, as well as by modulateing numerous enzymatic complexes involved in the production of ROS, endothelial function and aggregation of platelets. These vitamins can also regulate NADPH oxidase, the most important source of O2 • in the cardiovascular system. It has been reported that ascorbic acid and α-tocopherol, derivated from vitamin C and E respectively, may involved in the transcriptional modulation of NADPH oxidase [111].

The most common carotenoids are xanthophylls and carotenes. Carotenoids can neutralize singlet oxygen by quenching it or can break the chain reaction of free radicals, or scavenging it, not so effective action (scavenging). The structure of the free radical is the main factor that determines if the carotenoid will have quenching or scavenging action. It also depends on the region where the radical is in heterogeneous biological tissue, aqueous or lipid region (plasma, blood, heart, liver, brain etc.), and the structure of the carotenoids (number of conjugated, cyclic or acyclic double bonds), polar or nonpolar groups, redox properties [112–114].

The physical quenching is the transfer of excitation energy from the singlet oxygen to the carotenoid. The oxygen returns to ground state and the carotenoid is in the excited triplet state, the energy is dissipated producing stable carotenoid and thermal energy and the carotenoid can undergo other cycles of singlet oxygen quenching [112, 115].

The chemical quenching the carotenoid combines with oxygen or is oxidized, leading to its destruction and producing a variety of oxidized products. Carotenoids can also extinguish the triplet-excited state of chlorophyll or other excited sensitizers, thus preventing the formation of singlet oxygen [112]. The free radical scavenging can occur in three ways, by electron transfer, by hydrogen abstraction, and by addition [112, 116].

### 4. Interaction between reactive species, enzymes, and antioxidant molecules in health and disease

All living cells have molecular tools to perceive and respond properly to environmental cues. All the cascades of intracellular reactions involved in promoting a biochemical response are denoted as signal transduction. There are well known receptor types or systems of signal transduction such as the G protein-coupled receptors (GPCR), tyrosine kinase receptors (TKR), ion channels, cell adhesion receptors, nuclear receptors and guanylyl-cyclases. Since cells often need to deal with many signals at the same time, the final biochemical response is a result of the integrations of many simultaneous cascades produced by one or more systems.

several targets, such as GSK3, BAD, FOXO, p53, NF-kB, mTOR/p70S6K1 and HIF-1 [122, 123]. In this way, ROS increase the final cascade response in cell, i.e., cell cycle progression, proliferation, anti-apoptosis, invasion, autophagy and angiogenesis [124]. The PI3K/AKT pathway hyper

An important class of redox regulated proteins is the Src family of nonreceptor tyrosine kinases (SFKs), a group of structurally related kinases that catalyze the phosphorylation of tyrosine in downstream targets to regulate cellular functions coupling receptors such as the TKR, the cell adhesion molecules (CAMs), and the GPCR to the cellular signaling machinery [125]. For example, during focal adhesion while the extracellular matrix (ECM) contact triggers a slight or partial activation of SFKs, the ROS production is associated with a strong oxidative-dependent activation and recruitment of Src kinases to cell membranes. The redoxactivation of SFKs can induce sustained PI3K, protein kinase C (PKC), and extracellular regulated kinase (ERK) activation and, thereby, create conditions for tumor cell growth, invasion, angiogenesis, and resistance to apoptosis [126]. In a variety of human cancers an increased activity of Src kinases have been described, as well as activation of important Src downstream targets such as PI3K/Akt, focal adhesion kinase (FAK), paxillin, p130Cas, signal transducer and activator of transcription 3 (STAT3) and stress-activated protein kinase/c-Jun

Carcinogenesis is also related with activator protein-1 (AP-1) transcription factor activation. Among other systems, ROS are recognized as activators of AP-1; however, the signaling transduction events involved are not totally understood. Chromium, cobalt, cadmium and vanadium are metals involved in the activation of AP-1 through signaling cascades involving the production of ROS and comprised of proteins and enzymes such as thioredoxin (Trx), redox factor-1 (Ref-1), ERK/MAPK, NADPH oxidase, I kappa B kinase (IKK), p38, JNK/c-jun

ROS are important for the regulation of vascular tone, however an excess of reactive species might be associated with pathological dysfunction. Endothelial nitric oxide synthase (eNOS) regulates smooth muscle cells (SMC) relaxation through the production of the second messenger nitric oxide (NO) from L-arginine, which activates guanylyl cyclases to initiate the conversion of GTP to cyclic guanosine monophosphate (cGMP), which allosterically activates the cGMPdependent protein kinases (PKGs). The enzyme eNOS can be uncoupled and, consequently,

in eNOS uncoupling. First, an increase of ROS might generate the peroxynitrite (ONOO)

biopterin (THB/BH4), a cofactor of eNOS [133]. Second, an increased ratio of oxidized glutathione (GSSG)/reduced glutathione (GSH) cause reversible S-glutathionylation and uncoupling of eNOS [134]. Paradoxically, H2O2 produced by NADPH oxidase increases eNOS expression and NO production, but this effect is not believed to counteract the effects of oxidative stress [135]. Interestingly, in a scenario of reduced NO levels, in which it would be expected a lack of input signals to PKG activation (e.g., cGMP), the H2O2 can cause vasodilation through PKG oxidation [136]. Another target of ROS is the small GTPase RhoA, which when oxidized activates its downstream partner Rho kinase (ROCK), leading to inhibitory phosphorylation of myosin light chain (MLC) phosphatase and, ultimately, to SMC contraction [137, 138]. For a more

<sup>∙</sup> production instead. Two major events are involved

Oxidative Stress: Noxious but Also Vital http://dx.doi.org/10.5772/intechopen.73394 11

<sup>∙</sup>. The anion ONOO reacts with and oxidizes tetrahydro-

activated by ROS might favor carcinogenesis in the end of the process.

N-terminal kinase (SAPK/JNK) [127–130].

change its profile from NO synthesis to O2

through the reaction of NO and O2

[117, 131, 132].

Before we move on exploring the targets of ROS in health and disease, an important question is raised: "Which are the main sources of cellular ROS?" Enzymes such as NADPH oxidases (Nox), xanthine oxidase (XO), lipoxygenase, MPO and uncoupled nitric oxide synthase are involved in the production of the anion radical superoxide (O2 <sup>∙</sup>�). Furthermore, the mitochondrial aerobic respiration contributes with a huge amount of O2 <sup>∙</sup>�. Peroxynitrite (ONOO�) is formed by the reaction of nitric oxide and superoxide and is thought to contribute to eNOS uncoupling [69]. The majority of O2 <sup>∙</sup>� generated within the mitochondrial matrix or the cytosol is dismutated to H2O2 by the SOD antioxidant enzyme. Moreover, metal exposure can mediate the generation of H2O2, O2 <sup>∙</sup>�, and even the hydroxyl radical (OH<sup>∙</sup> ), mainly via the Fenton or the Haber-Weiss reactions [117].

Some ROS such as O2 <sup>∙</sup>� and HO<sup>∙</sup> are highly reactive and have a brief half life. For this reason they are not considered signaling molecules, but intermediates of nonselective nature. On the other hand, H2O2 is relatively stable and can both mediate intracellular signaling and also serve to paracrine signaling (i.e., cell-to-cell communication involving nearby cells), since it can cross biological membranes [118].

Up to date, several proteins have been recognized as downstream targets of ROS, such as kinases, phosphatases, mitogen-activated protein kinases (MAPK), small G proteins, transcription factors, microRNAs, and phospholipases. In this section, we do not intend to deeply review the literature, but to show an overview of important targets and exemplify their involvement in the signal transduction by ROS in health and disease.

ROS can induce alterations in the intracellular and extracellular processes, for example, in the PI3K/AKT signaling. The lipid phosphatidylinositol 3,4,5-triphosphate (PIP3) has a function as a second messenger and is not present in the quiescent cells, but it rises within seconds to minutes when there is a stimuli. PIP3 is produced by the phosphorylation of the phosphatidylinositol 4,5 bisphosphate (PIP2) catalyzed by the phosphatidylinositol 3-kinase (PI3K). This enzyme is activated by ROS through two different pathways, or directly, throught amplications of downstream PI3K pathway, or indirectly by inhibition of the phosphatase and tensing homolog deleted on chromosome 10 (PTEN). PTEN is responsible for the degradation of PIP3 signaling, since it catalyzes the hydrolysis of phosphate in the 3<sup>0</sup> position on PIP3 to produce PIP2 [119]. ROS, mainly, H2O2, can oxidize and inhibit PTEN, which culminates in an increase in the PIP3 production, that acts in cell signaling, through activation of proteins, as serine/threonine protein kinase, AKT/PKB, among others [120, 121]. The AKT activation provides the transcription of several targets, such as GSK3, BAD, FOXO, p53, NF-kB, mTOR/p70S6K1 and HIF-1 [122, 123]. In this way, ROS increase the final cascade response in cell, i.e., cell cycle progression, proliferation, anti-apoptosis, invasion, autophagy and angiogenesis [124]. The PI3K/AKT pathway hyper activated by ROS might favor carcinogenesis in the end of the process.

4. Interaction between reactive species, enzymes, and antioxidant

All living cells have molecular tools to perceive and respond properly to environmental cues. All the cascades of intracellular reactions involved in promoting a biochemical response are denoted as signal transduction. There are well known receptor types or systems of signal transduction such as the G protein-coupled receptors (GPCR), tyrosine kinase receptors (TKR), ion channels, cell adhesion receptors, nuclear receptors and guanylyl-cyclases. Since cells often need to deal with many signals at the same time, the final biochemical response is a result of the integrations of many simultaneous cascades produced by one or more systems.

Before we move on exploring the targets of ROS in health and disease, an important question is raised: "Which are the main sources of cellular ROS?" Enzymes such as NADPH oxidases (Nox), xanthine oxidase (XO), lipoxygenase, MPO and uncoupled nitric oxide synthase are

formed by the reaction of nitric oxide and superoxide and is thought to contribute to eNOS

is dismutated to H2O2 by the SOD antioxidant enzyme. Moreover, metal exposure can mediate

they are not considered signaling molecules, but intermediates of nonselective nature. On the other hand, H2O2 is relatively stable and can both mediate intracellular signaling and also serve to paracrine signaling (i.e., cell-to-cell communication involving nearby cells), since it can

Up to date, several proteins have been recognized as downstream targets of ROS, such as kinases, phosphatases, mitogen-activated protein kinases (MAPK), small G proteins, transcription factors, microRNAs, and phospholipases. In this section, we do not intend to deeply review the literature, but to show an overview of important targets and exemplify their

ROS can induce alterations in the intracellular and extracellular processes, for example, in the PI3K/AKT signaling. The lipid phosphatidylinositol 3,4,5-triphosphate (PIP3) has a function as a second messenger and is not present in the quiescent cells, but it rises within seconds to minutes when there is a stimuli. PIP3 is produced by the phosphorylation of the phosphatidylinositol 4,5 bisphosphate (PIP2) catalyzed by the phosphatidylinositol 3-kinase (PI3K). This enzyme is activated by ROS through two different pathways, or directly, throught amplications of downstream PI3K pathway, or indirectly by inhibition of the phosphatase and tensing homolog deleted on chromosome 10 (PTEN). PTEN is responsible for the degradation of PIP3 signaling, since it catalyzes the hydrolysis of phosphate in the 3<sup>0</sup> position on PIP3 to produce PIP2 [119]. ROS, mainly, H2O2, can oxidize and inhibit PTEN, which culminates in an increase in the PIP3 production, that acts in cell signaling, through activation of proteins, as serine/threonine protein kinase, AKT/PKB, among others [120, 121]. The AKT activation provides the transcription of

<sup>∙</sup>�, and even the hydroxyl radical (OH<sup>∙</sup>

<sup>∙</sup>�). Furthermore, the mitochon-

<sup>∙</sup>� generated within the mitochondrial matrix or the cytosol

are highly reactive and have a brief half life. For this reason

<sup>∙</sup>�. Peroxynitrite (ONOO�) is

), mainly via the Fenton or the

involved in the production of the anion radical superoxide (O2

<sup>∙</sup>� and HO<sup>∙</sup>

drial aerobic respiration contributes with a huge amount of O2

involvement in the signal transduction by ROS in health and disease.

molecules in health and disease

10 Novel Prospects in Oxidative and Nitrosative Stress

uncoupling [69]. The majority of O2

cross biological membranes [118].

the generation of H2O2, O2

Some ROS such as O2

Haber-Weiss reactions [117].

An important class of redox regulated proteins is the Src family of nonreceptor tyrosine kinases (SFKs), a group of structurally related kinases that catalyze the phosphorylation of tyrosine in downstream targets to regulate cellular functions coupling receptors such as the TKR, the cell adhesion molecules (CAMs), and the GPCR to the cellular signaling machinery [125]. For example, during focal adhesion while the extracellular matrix (ECM) contact triggers a slight or partial activation of SFKs, the ROS production is associated with a strong oxidative-dependent activation and recruitment of Src kinases to cell membranes. The redoxactivation of SFKs can induce sustained PI3K, protein kinase C (PKC), and extracellular regulated kinase (ERK) activation and, thereby, create conditions for tumor cell growth, invasion, angiogenesis, and resistance to apoptosis [126]. In a variety of human cancers an increased activity of Src kinases have been described, as well as activation of important Src downstream targets such as PI3K/Akt, focal adhesion kinase (FAK), paxillin, p130Cas, signal transducer and activator of transcription 3 (STAT3) and stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK) [127–130].

Carcinogenesis is also related with activator protein-1 (AP-1) transcription factor activation. Among other systems, ROS are recognized as activators of AP-1; however, the signaling transduction events involved are not totally understood. Chromium, cobalt, cadmium and vanadium are metals involved in the activation of AP-1 through signaling cascades involving the production of ROS and comprised of proteins and enzymes such as thioredoxin (Trx), redox factor-1 (Ref-1), ERK/MAPK, NADPH oxidase, I kappa B kinase (IKK), p38, JNK/c-jun [117, 131, 132].

ROS are important for the regulation of vascular tone, however an excess of reactive species might be associated with pathological dysfunction. Endothelial nitric oxide synthase (eNOS) regulates smooth muscle cells (SMC) relaxation through the production of the second messenger nitric oxide (NO) from L-arginine, which activates guanylyl cyclases to initiate the conversion of GTP to cyclic guanosine monophosphate (cGMP), which allosterically activates the cGMPdependent protein kinases (PKGs). The enzyme eNOS can be uncoupled and, consequently, change its profile from NO synthesis to O2 <sup>∙</sup> production instead. Two major events are involved in eNOS uncoupling. First, an increase of ROS might generate the peroxynitrite (ONOO) through the reaction of NO and O2 <sup>∙</sup>. The anion ONOO reacts with and oxidizes tetrahydrobiopterin (THB/BH4), a cofactor of eNOS [133]. Second, an increased ratio of oxidized glutathione (GSSG)/reduced glutathione (GSH) cause reversible S-glutathionylation and uncoupling of eNOS [134]. Paradoxically, H2O2 produced by NADPH oxidase increases eNOS expression and NO production, but this effect is not believed to counteract the effects of oxidative stress [135].

Interestingly, in a scenario of reduced NO levels, in which it would be expected a lack of input signals to PKG activation (e.g., cGMP), the H2O2 can cause vasodilation through PKG oxidation [136]. Another target of ROS is the small GTPase RhoA, which when oxidized activates its downstream partner Rho kinase (ROCK), leading to inhibitory phosphorylation of myosin light chain (MLC) phosphatase and, ultimately, to SMC contraction [137, 138]. For a more

explored involvement of ROS in the regulation of signal transduction in the cardiovascular system, check the review of Brown and Griendling [118].

ethanolamine [147]. A link between oxidative stress and PLD has been proposed by Kim et al. [148], in a study that suggests that H2O2 induces rat vascular smooth muscle cells tyrosine

Oxidative Stress: Noxious but Also Vital http://dx.doi.org/10.5772/intechopen.73394 13

In the innate immune system, mononuclear monocytes/macrophages eliminate pathogen, antigen and cellular components through generation of ROS/RNS [149]. When there is an imbalance in the equilibrium between oxidative/nitrosative stress and cellular requirements, the stress can generates pathological complications. Among others, rheumatoid arthritis is an autoimmune disease that has oxidative/nitrosative stress as one of the causes. The cellular immune system is vulnerable to reactions caused by ROS, which in turn can affect the regular physiological process and activates inflammatory signaling pathways that produce pro-inflammatory cytokines, chemokines and prostaglandins. The inflammatory mechanism involves synovial cellular infiltrate and peripheral blood inflammatory cells following by polymorphonuclear neutrophils and lymphocytes culminating in the joint damage [150, 151]. The signaling cascade occurs via activation of NFkB for synthesizing pro-inflammatory cytokines and chemokines [149]. The Th1 cytokines are one of the most important because can provide the development of autoimmune disorders. These cytokines can directly or indirectly promote oxidative stress in the cells, inten-

Prostaglandins have a pivotal role in the formation of the inflammatory response, since they mediate pathogenic mechanisms and provide the development of the cardinal signs of acute inflammation. Their biosynthesis involves the initial enzyme, phospholipase A2 (PLA2). PLA2 catalyzes the conversion of membrane phospholipids in AA. Then, cyclooxygenases convert AA into prostaglandins. Prostaglandin E2, in particular, rises vasoactive components (histamine, bradykinin, and nitric oxide), hence generating edema, pain and hyperalgesia at the local inflammatory sites, and so the inflammation [152]. ROS stimulate this process through the activation of cyclooxygenases. Prostaglandins, also, activate NADPH oxidase, which produces superoxide anion radical [153]. Therefore, this system becomes cyclic, ROS activate cyclooxygenases and so the prostaglandins biosynthesis, further prostaglandins trigger NAPH

The microRNA (miRNA) is a small noncoding endogenous RNA, that has an important role, since it regulates gene expression. Its function can be modified depending on epigenetic changes, chromosomal abnormalities and oxidative stress. It has been found that miRNA can respond to ROS, implying in its ability to activate certain genes transcription during stress, and this is prominent in cancer cells, which was correlated to the adaptation of these cells to unfavorable and/or hypoxic environment [130, 154, 155]. However, studies showed that some types of miRNAs can regulate gene expression of protective proteins and antioxidant enzymes [156, 157]. Some ROS dependent miRNAs play a role as oncogenic (miR21 and miR155), but interesting miR21 also targets SOD, which can be interpreted that this miRNA regulate the ROS levels in the cell. When miR21 is stimulated, it also affects the immune system through the chemokine CXCL10. CXCL10 adjusts innate and adaptive immune response by activating T lymphocytes, macrophages and inflammatory dendritic cells. The miR155 also has opposite actions, it can be oncogenic (the targets are BCL2, FOXO3a, RhoA) or tumor suppressor (the targets are TGF-beta/SMAD) [158]. The literature about miR155 is vast, and we suggest the articles by Higgs and Slack [158] and Mattiske et al. [159] for a deep reading. Besides these two

kinase activity, and PLD1-dependent PKC-α activation.

sifying the rheumatoid arthritis.

oxidases, increasing ROS.

The activating or deactivating switch, in which a group of kinases is active or a group of phosphatases is active, provokes different downstream cascades with consequences in the cellular response. As we described above, several kinases are susceptible to ROS reactions, but also phosphatases are vulnerable to ROS, since they react with a group of amino acids presents in different enzymes. The reaction between ROS and phosphatases causes the oxidation and inhibition of those enzymes, increasing the kinases signaling [139]. Another phosphatase inhibited by ROS is PTEN, which increases the PIP3 signaling, as described above.

A vascular injury promotes an increase in the expression of platelet derived growth factor (PDGF) and PDGF receptor, which in turn cause stimulation for the vascular smooth muscle cells to migrate [140]. The activation of the PDGF receptor is controlled by the action of low molecular weight protein tyrosine phosphatase (LMW-PTP). The Cys12 and Cys17 in LMW-PTP is susceptible to a reaction with ROS resulting in a disulfide bond, and so its inactivation [141]. Therefore, without the LMW-PTP deactivation upon PDGF receptor, its signal is amplified, which generates migration. Oxidized LMW-PTP also increases the Rho family signal, since PDGF receptor is stimulated, and it binds to phospholipase C, Src, and PI3K. As described before, PI3K catalyzes the reaction and formation of PIP3. The Rho-guanine nucleotide exchange factors are activated by PIP3, which triggers Rho-GTPase family members' activation (Rho, Rac, and cdc42). As Nox family is activated by Rac, it produces ROS. Therefore, this process is kept by a positive feedback: generated ROS oxide Rho in a redox sensitive motif and restrain the LMW-PTP action [118, 138].

Phospholipases are enzymes that hydrolyze phospholipids and generate second messengers involved in the regulation of many physiological functions. Phospholipase A2 (PLA2) cleaves the fatty acyl group at the sn-2 position of the glycerol backbone, releasing arachidonic acid (AA) and lysophospholipid. It was attributed a role for the Ca2+-independent PLA2 (iPLA2) isoform in the excessive production of O2 <sup>∙</sup> by primed neutrophils of patients with poorly controlled diabetes. This study suggested that hyperglycemia is related to the activation of iPLA2 and AA formation which, in part, regulate NADPH oxidase activity (i.e., generation of O2 <sup>∙</sup>) [142].

PLA2 activation has also been related to alterations implicated in the pathogenesis of neurodegenerative diseases, such as neuronal excitation, cognitive and behavioral function, oxidative and nitrosative stress [143]. Phospholipase C (PLC) is a well-known enzyme especially involved in the signaling transduction of GPCR coupled to Gq/11 protein and some G protein βγ subunits (PLC-β), but also in RTK (PLC-γ and PLC-ε), Ras and Rho small GTPases (PLC-ε) and Ca2+ (PLC-δ) signaling pathways, which involves the generation of the phosphatecontaining head group inositol 1,4,5-triphosphate (IP3) and diacylglycerol (DAG) through the hydrolysis of the membrane phospholipid PIP2 [144]. The activation of PLC-γ1 was shown to have an important protective function during mouse embryonic fibroblasts (MEF) response to oxidative stress (H2O2) treatment [145]. A further study suggested that this function of PLC-γ1 involved the PKC-dependent phosphorylation of Bcl-2 and inhibition of caspase-3 [146]. Phospholipase D (PLD) cleaves a phosphodiester bold in membrane-bound lipids, similarly to PLC. However, its activity generates phosphatidic acid (PA) and an alcohol, usually choline or ethanolamine [147]. A link between oxidative stress and PLD has been proposed by Kim et al. [148], in a study that suggests that H2O2 induces rat vascular smooth muscle cells tyrosine kinase activity, and PLD1-dependent PKC-α activation.

explored involvement of ROS in the regulation of signal transduction in the cardiovascular

The activating or deactivating switch, in which a group of kinases is active or a group of phosphatases is active, provokes different downstream cascades with consequences in the cellular response. As we described above, several kinases are susceptible to ROS reactions, but also phosphatases are vulnerable to ROS, since they react with a group of amino acids presents in different enzymes. The reaction between ROS and phosphatases causes the oxidation and inhibition of those enzymes, increasing the kinases signaling [139]. Another phosphatase inhibited by ROS is PTEN, which increases the PIP3 signaling, as described above.

A vascular injury promotes an increase in the expression of platelet derived growth factor (PDGF) and PDGF receptor, which in turn cause stimulation for the vascular smooth muscle cells to migrate [140]. The activation of the PDGF receptor is controlled by the action of low molecular weight protein tyrosine phosphatase (LMW-PTP). The Cys12 and Cys17 in LMW-PTP is susceptible to a reaction with ROS resulting in a disulfide bond, and so its inactivation [141]. Therefore, without the LMW-PTP deactivation upon PDGF receptor, its signal is amplified, which generates migration. Oxidized LMW-PTP also increases the Rho family signal, since PDGF receptor is stimulated, and it binds to phospholipase C, Src, and PI3K. As described before, PI3K catalyzes the reaction and formation of PIP3. The Rho-guanine nucleotide exchange factors are activated by PIP3, which triggers Rho-GTPase family members' activation (Rho, Rac, and cdc42). As Nox family is activated by Rac, it produces ROS. Therefore, this process is kept by a positive feedback: generated ROS oxide Rho in a redox sensitive

Phospholipases are enzymes that hydrolyze phospholipids and generate second messengers involved in the regulation of many physiological functions. Phospholipase A2 (PLA2) cleaves the fatty acyl group at the sn-2 position of the glycerol backbone, releasing arachidonic acid (AA) and lysophospholipid. It was attributed a role for the Ca2+-independent PLA2 (iPLA2)

controlled diabetes. This study suggested that hyperglycemia is related to the activation of iPLA2 and AA formation which, in part, regulate NADPH oxidase activity (i.e., generation of

PLA2 activation has also been related to alterations implicated in the pathogenesis of neurodegenerative diseases, such as neuronal excitation, cognitive and behavioral function, oxidative and nitrosative stress [143]. Phospholipase C (PLC) is a well-known enzyme especially involved in the signaling transduction of GPCR coupled to Gq/11 protein and some G protein βγ subunits (PLC-β), but also in RTK (PLC-γ and PLC-ε), Ras and Rho small GTPases (PLC-ε) and Ca2+ (PLC-δ) signaling pathways, which involves the generation of the phosphatecontaining head group inositol 1,4,5-triphosphate (IP3) and diacylglycerol (DAG) through the hydrolysis of the membrane phospholipid PIP2 [144]. The activation of PLC-γ1 was shown to have an important protective function during mouse embryonic fibroblasts (MEF) response to oxidative stress (H2O2) treatment [145]. A further study suggested that this function of PLC-γ1 involved the PKC-dependent phosphorylation of Bcl-2 and inhibition of caspase-3 [146]. Phospholipase D (PLD) cleaves a phosphodiester bold in membrane-bound lipids, similarly to PLC. However, its activity generates phosphatidic acid (PA) and an alcohol, usually choline or

<sup>∙</sup> by primed neutrophils of patients with poorly

system, check the review of Brown and Griendling [118].

12 Novel Prospects in Oxidative and Nitrosative Stress

motif and restrain the LMW-PTP action [118, 138].

isoform in the excessive production of O2

O2

<sup>∙</sup>) [142].

In the innate immune system, mononuclear monocytes/macrophages eliminate pathogen, antigen and cellular components through generation of ROS/RNS [149]. When there is an imbalance in the equilibrium between oxidative/nitrosative stress and cellular requirements, the stress can generates pathological complications. Among others, rheumatoid arthritis is an autoimmune disease that has oxidative/nitrosative stress as one of the causes. The cellular immune system is vulnerable to reactions caused by ROS, which in turn can affect the regular physiological process and activates inflammatory signaling pathways that produce pro-inflammatory cytokines, chemokines and prostaglandins. The inflammatory mechanism involves synovial cellular infiltrate and peripheral blood inflammatory cells following by polymorphonuclear neutrophils and lymphocytes culminating in the joint damage [150, 151]. The signaling cascade occurs via activation of NFkB for synthesizing pro-inflammatory cytokines and chemokines [149]. The Th1 cytokines are one of the most important because can provide the development of autoimmune disorders. These cytokines can directly or indirectly promote oxidative stress in the cells, intensifying the rheumatoid arthritis.

Prostaglandins have a pivotal role in the formation of the inflammatory response, since they mediate pathogenic mechanisms and provide the development of the cardinal signs of acute inflammation. Their biosynthesis involves the initial enzyme, phospholipase A2 (PLA2). PLA2 catalyzes the conversion of membrane phospholipids in AA. Then, cyclooxygenases convert AA into prostaglandins. Prostaglandin E2, in particular, rises vasoactive components (histamine, bradykinin, and nitric oxide), hence generating edema, pain and hyperalgesia at the local inflammatory sites, and so the inflammation [152]. ROS stimulate this process through the activation of cyclooxygenases. Prostaglandins, also, activate NADPH oxidase, which produces superoxide anion radical [153]. Therefore, this system becomes cyclic, ROS activate cyclooxygenases and so the prostaglandins biosynthesis, further prostaglandins trigger NAPH oxidases, increasing ROS.

The microRNA (miRNA) is a small noncoding endogenous RNA, that has an important role, since it regulates gene expression. Its function can be modified depending on epigenetic changes, chromosomal abnormalities and oxidative stress. It has been found that miRNA can respond to ROS, implying in its ability to activate certain genes transcription during stress, and this is prominent in cancer cells, which was correlated to the adaptation of these cells to unfavorable and/or hypoxic environment [130, 154, 155]. However, studies showed that some types of miRNAs can regulate gene expression of protective proteins and antioxidant enzymes [156, 157]. Some ROS dependent miRNAs play a role as oncogenic (miR21 and miR155), but interesting miR21 also targets SOD, which can be interpreted that this miRNA regulate the ROS levels in the cell. When miR21 is stimulated, it also affects the immune system through the chemokine CXCL10. CXCL10 adjusts innate and adaptive immune response by activating T lymphocytes, macrophages and inflammatory dendritic cells. The miR155 also has opposite actions, it can be oncogenic (the targets are BCL2, FOXO3a, RhoA) or tumor suppressor (the targets are TGF-beta/SMAD) [158]. The literature about miR155 is vast, and we suggest the articles by Higgs and Slack [158] and Mattiske et al. [159] for a deep reading. Besides these two miRNAs cited above, others miRNAs are upregulated by ROS, such as miR23, miR200, miR210, etc., affecting migration, invasion; tumor growth, angiogenesis; cell cycle, DNA damage (among others), respectively [126].

summary of some important miRNAs and their responses in carcinogenesis, for more informa-

As previously discussed in this chapter, cells have a repertoire of antioxidant molecules and enzymes as a defense mechanism to an increase in ROS production. However, oxidative stress takes place when the antioxidant capacity is overwhelmed by reactive species production. In this scenario, to maintain cell homeostasis and/or terminate the ROS signal transduction there are some stress sensors that regulate the translation of antioxidant proteins. The antioxidant responsive element (ARE) is a region of non-coding DNA (short consensus sequence) which is localized upstream and regulates the transcription of many antioxidant neighboring genes such as glutathione S-transferases (GST), NAD(P)H:quinone oxidoreductase (NQO1) [162], heme oxygenase 1 (HO-1), γ-glutamylcysteine synthetase (γ-GCS) [163], metallothionein-1

It was shown that ARE induction protected against oxidative stress mediated by 6- hydroxydopamine in vitro, a mitochondrial inhibitor used to model Parkinson's disease [166]. The nuclear-factor erythroid-2 related factor (Nrf2) is a central transcription factor involved in the upregulation of ARE-containing genes and, consequently, synthesis of proteins with antioxidant function. However, there are also nuclear factors that negatively regulate ARE-mediated gene

Finally, in this section, we showed an overview of processes regulated by fluctuating levels of ROS and their molecular sensors. Furthermore, we showed that in response to oxidative stress and to maintain homeostasis, cells can upregulate the synthesis of antioxidant defenses

H.U.'s research is supported by the São Paulo Research Foundation (FAPESP proj. No. 2012/

\*, Jeandre Augusto dos Santos Jaques<sup>2</sup>

, Lucas Derbocio dos Santos<sup>2</sup> and Henning Ulrich3

, Micheli Mainardi Pillat3

\*Address all correspondence to: margaretebagatini@yahoo.com.br

2 Universidade Federal de Mato Grosso do Sul, Campo Grande, MS, Brazil

1 Universidade Federal da Fronteira Sul, Chapecó, SC, Brazil

4 Universidade Federal de Santa Maria, Santa Maria, RS, Brazil

3 Universidade de São Paulo, São Paulo, SP, Brazil

, Carla Santos de Oliveira<sup>2</sup>

Oxidative Stress: Noxious but Also Vital http://dx.doi.org/10.5772/intechopen.73394 15

,

, Aline Mânica<sup>4</sup>

,

expression, such as Mafs (MafG and MAfK), large Maf (c-Maf), c-Fos, and Fra1 [163].

tion check the review Mu and Liu [126].

and -2 (MT-1 and MT-2) [164], and SOD [165].

(Figure 2).

50880-4).

Author details

Margarete Dulce Bagatini<sup>1</sup>

Cintia dos Santos Moser<sup>1</sup>

Graciele Almeida de Oliveira3

Acknowledgements

In addition to the miRNAs that are ROS upregulated as cited above, there are ROS downregulated miRNAs important in the carcinogenic process, such as miR34 family. Some miR34 members regulate p53 causing a cell cycle arrest in G1 and apoptosis when DNA is impaired. The miR34a, for example, induce tumor suppression and metastasis inhibition. Another miRNA, miR124, has been shown to be affected by H2O2 [160]. This miRNA is correlated to the regulation of tumor cell proliferation, migration and drug resistance through its action upon R-Ras, PI3- KCA, AKT2, ROCK1, Src, DNA methyltransferases and others. The miR199a is also downregulated by ROS, some of its targets are ERBB2, ERBB3, IKKB, HIF-1alfa, ApoE, CCR7, having an effect upon cell proliferation, invasion, metabolism and metastasis [126, 161]. This is just a

Figure 2. Examples of molecular targets involved in the signal transduction mediated by reactive oxygen species. Abbreviations: AA, arachidonic acid; AP1, activator protein 1; ARE, antioxidant-responsive element; BAD, Bcl-2-associated death promoter; Bcl-2, B-cell lymphoma 2 protein; cGMP, cyclic guanosine monophosphate; DAG, diacylglycerol; eNOS, endothelial nitric oxide synthase; ERK, extracellular-signal regulated kinase; FAK, focal adhesion kinase; FOXO, Forkhead box protein O; GC, guanylyl cyclase; GSK-3, glycogen synthase kinase 3; GST, glutathione S-transferases; HIF-1, hypoxia-inducible factor 1; HO-1, heme oxygenase 1; IP3, inositol 1,4,5-triphosphate; LMW-PTP, low molecular weight phosphotyrosine protein phosphatase; MT-1, metallothionein-1; MT-2, metallothionein-2; mTOR, mammalian target of rapamycin; NF-κB, nuclear factor-kappa B; NO, nitric oxide; NQO1, NAD(P)H:quinone oxidoreductase; Nrf2, nuclearfactor erythroid-2 related factor; O2 <sup>∙</sup>, superoxide anion radical; p130Cas, p130 Crk-associated substrate; p53, p53 tumor suppressor protein; p70S6K1, p70S6 kinase 1; PDK, phosphoinositide-dependent kinase; PI3K, phosphatidylinositol 3 kinase; PIP2, phosphatidylinositol 4,5-bisphosphate; PIP3, phosphatidylinositol 3,4,5-triphosphate; PKB/AKT, protein kinase B; PKC, protein kinase C; PKC, protein kinase C; PKG, cGMP-dependent protein kinases; PLA2, phospholipase A2; PLC, phospholipase C; PLD, phospholipase D; PTEN, phosphatase and tensin homolog deleted on chromosome 10; RhoA, Ras homolog family member A; ROCK, Rho-associated protein kinase; SAPK/JNK, stress-activated protein kinase or c-Jun N-terminal kinase; SOD, superoxide dismutase; STAT3, signal transducer and activator of transcription 3; γ-GCS, gamma-glutamylcysteine synthetase.

summary of some important miRNAs and their responses in carcinogenesis, for more information check the review Mu and Liu [126].

As previously discussed in this chapter, cells have a repertoire of antioxidant molecules and enzymes as a defense mechanism to an increase in ROS production. However, oxidative stress takes place when the antioxidant capacity is overwhelmed by reactive species production. In this scenario, to maintain cell homeostasis and/or terminate the ROS signal transduction there are some stress sensors that regulate the translation of antioxidant proteins. The antioxidant responsive element (ARE) is a region of non-coding DNA (short consensus sequence) which is localized upstream and regulates the transcription of many antioxidant neighboring genes such as glutathione S-transferases (GST), NAD(P)H:quinone oxidoreductase (NQO1) [162], heme oxygenase 1 (HO-1), γ-glutamylcysteine synthetase (γ-GCS) [163], metallothionein-1 and -2 (MT-1 and MT-2) [164], and SOD [165].

It was shown that ARE induction protected against oxidative stress mediated by 6- hydroxydopamine in vitro, a mitochondrial inhibitor used to model Parkinson's disease [166]. The nuclear-factor erythroid-2 related factor (Nrf2) is a central transcription factor involved in the upregulation of ARE-containing genes and, consequently, synthesis of proteins with antioxidant function. However, there are also nuclear factors that negatively regulate ARE-mediated gene expression, such as Mafs (MafG and MAfK), large Maf (c-Maf), c-Fos, and Fra1 [163].

Finally, in this section, we showed an overview of processes regulated by fluctuating levels of ROS and their molecular sensors. Furthermore, we showed that in response to oxidative stress and to maintain homeostasis, cells can upregulate the synthesis of antioxidant defenses (Figure 2).

### Acknowledgements

miRNAs cited above, others miRNAs are upregulated by ROS, such as miR23, miR200, miR210, etc., affecting migration, invasion; tumor growth, angiogenesis; cell cycle, DNA dam-

In addition to the miRNAs that are ROS upregulated as cited above, there are ROS downregulated miRNAs important in the carcinogenic process, such as miR34 family. Some miR34 members regulate p53 causing a cell cycle arrest in G1 and apoptosis when DNA is impaired. The miR34a, for example, induce tumor suppression and metastasis inhibition. Another miRNA, miR124, has been shown to be affected by H2O2 [160]. This miRNA is correlated to the regulation of tumor cell proliferation, migration and drug resistance through its action upon R-Ras, PI3- KCA, AKT2, ROCK1, Src, DNA methyltransferases and others. The miR199a is also downregulated by ROS, some of its targets are ERBB2, ERBB3, IKKB, HIF-1alfa, ApoE, CCR7, having an effect upon cell proliferation, invasion, metabolism and metastasis [126, 161]. This is just a

Figure 2. Examples of molecular targets involved in the signal transduction mediated by reactive oxygen species. Abbreviations: AA, arachidonic acid; AP1, activator protein 1; ARE, antioxidant-responsive element; BAD, Bcl-2-associated death promoter; Bcl-2, B-cell lymphoma 2 protein; cGMP, cyclic guanosine monophosphate; DAG, diacylglycerol; eNOS, endothelial nitric oxide synthase; ERK, extracellular-signal regulated kinase; FAK, focal adhesion kinase; FOXO, Forkhead box protein O; GC, guanylyl cyclase; GSK-3, glycogen synthase kinase 3; GST, glutathione S-transferases; HIF-1, hypoxia-inducible factor 1; HO-1, heme oxygenase 1; IP3, inositol 1,4,5-triphosphate; LMW-PTP, low molecular weight phosphotyrosine protein phosphatase; MT-1, metallothionein-1; MT-2, metallothionein-2; mTOR, mammalian target of rapamycin; NF-κB, nuclear factor-kappa B; NO, nitric oxide; NQO1, NAD(P)H:quinone oxidoreductase; Nrf2, nuclear-

suppressor protein; p70S6K1, p70S6 kinase 1; PDK, phosphoinositide-dependent kinase; PI3K, phosphatidylinositol 3 kinase; PIP2, phosphatidylinositol 4,5-bisphosphate; PIP3, phosphatidylinositol 3,4,5-triphosphate; PKB/AKT, protein kinase B; PKC, protein kinase C; PKC, protein kinase C; PKG, cGMP-dependent protein kinases; PLA2, phospholipase A2; PLC, phospholipase C; PLD, phospholipase D; PTEN, phosphatase and tensin homolog deleted on chromosome 10; RhoA, Ras homolog family member A; ROCK, Rho-associated protein kinase; SAPK/JNK, stress-activated protein kinase or c-Jun N-terminal kinase; SOD, superoxide dismutase; STAT3, signal transducer and activator of transcription 3; γ-GCS,

<sup>∙</sup>, superoxide anion radical; p130Cas, p130 Crk-associated substrate; p53, p53 tumor

age (among others), respectively [126].

14 Novel Prospects in Oxidative and Nitrosative Stress

factor erythroid-2 related factor; O2

gamma-glutamylcysteine synthetase.

H.U.'s research is supported by the São Paulo Research Foundation (FAPESP proj. No. 2012/ 50880-4).

### Author details

Margarete Dulce Bagatini<sup>1</sup> \*, Jeandre Augusto dos Santos Jaques<sup>2</sup> , Carla Santos de Oliveira<sup>2</sup> , Graciele Almeida de Oliveira3 , Micheli Mainardi Pillat3 , Aline Mânica<sup>4</sup> , Cintia dos Santos Moser<sup>1</sup> , Lucas Derbocio dos Santos<sup>2</sup> and Henning Ulrich3

\*Address all correspondence to: margaretebagatini@yahoo.com.br


### References

[1] Frey TG, Mannella CA. The internal structure of mitochondria. Trends in Biochemical Sciences [Internet]. 2000;25(7):319-324. Available from: http://www.ncbi.nlm.nih.gov/ entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list\_uids=10871882

[11] Friberg H, Ferrand-Drake M, Bengtsson F, Halestrap AP, Wieloch T. Cyclosporin A, but not FK 506, protects mitochondria and neurons against hypoglycemic damage and implicates the mitochondrial permeability transition in cell death. The Journal of Neuroscience [Internet]. 1998;18(14):5151-5159. Available from: http://www.ncbi.nlm.nih.gov/ entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list\_uids=9651198 [12] Charni S, de Bettignies G, Rathore MG, Aguilo JI, van den Elsen PJ, Haouzi D, et al. Oxidative phosphorylation induces de novo expression of the MHC class I in tumor cells through the ERK5 pathway. Journal of Immunology [Internet]. 2010/08/20. 2010 Sep 15;185(6):3498-3503. Available from: https://www.ncbi.nlm.nih.gov/pubmed/20729331 [13] D'Souza GGM, Wagle MA, Saxena V, Shah A. Approaches for targeting mitochondria in cancer therapy. Biochimica et Biophysica Acta (BBA) - Bioenergetics [Internet]. 2010/08/ 21. 2011 Jun;1807(6):689-696. Available from: https://www.ncbi.nlm.nih.gov/pubmed/

Oxidative Stress: Noxious but Also Vital http://dx.doi.org/10.5772/intechopen.73394 17

[14] Bek T. Mitochondrial dysfunction and diabetic retinopathy. Mitochondrion [Internet]. 2016/07/22. 2017 Sep;36:4-6. Available from: https://www.ncbi.nlm.nih.gov/pubmed/274

[15] Dey A, Swaminathan K. Hyperglycemia-induced mitochondrial alterations in liver. Life Sciences [Internet]. 2010/06/17. 2010 Aug;87(7–8):197-214. Available from: https://www.

[16] Tseng Y-H, Cypess AM, Kahn CR. Cellular bioenergetics as a target for obesity therapy. Nature Reviews. Drug Discovery [Internet]. 2010 Jun;9(6):465-482. Available from:

[17] Pickrell AM, Moraes CT. Protein Misfolding and Cellular Stress in Disease and Aging.

[18] Cassina P, Cassina A, Pehar M, Castellanos R, Gandelman M, de Leon A, et al. Mitochondrial dysfunction in SOD1G93A-bearing astrocytes promotes motor neuron degeneration: Prevention by mitochondrial-targeted antioxidants. The Journal of Neuroscience [Internet]. 2008 Apr 16;28(16):4115-4122. Available from: http://www.ncbi.nlm.nih.gov/ entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list\_uids=18417691 [19] Rose J, Brian C, Woods J, Pappa A, Panayiotidis MI, Powers R, et al. Mitochondrial dysfunction in glial cells: Implications for neuronal homeostasis and survival. Toxicology. 2017 Nov;391:109-115. Available from: https://www.ncbi.nlm.nih.gov/pubmed/28655545

[20] Nicholls DG, Ferguson SJ. Bioenergetics 4 [Internet]. 4th ed. Amsterdam: Elsevier Academic Press; 2013. xiv, 419 pp. Available from: http://www.sciencedirect.com/science/book/

[21] Mráček T, Drahota Z, Houštěk J. The function and the role of the mitochondrial glycerol-3-phosphate dehydrogenase in mammalian tissues. Biochimica et Biophysica Acta (BBA) - Bioenergetics [Internet]. 2012/12/07. 2013 Mar;1827(3):401-410. Available from:

20732297

56429

ncbi.nlm.nih.gov/pubmed/20600152

Totowa, NJ: Humana Press; 2010

9780123884251

http://www.nature.com/doifinder/10.1038/nrd3138

https://www.ncbi.nlm.nih.gov/pubmed/23220394


[11] Friberg H, Ferrand-Drake M, Bengtsson F, Halestrap AP, Wieloch T. Cyclosporin A, but not FK 506, protects mitochondria and neurons against hypoglycemic damage and implicates the mitochondrial permeability transition in cell death. The Journal of Neuroscience [Internet]. 1998;18(14):5151-5159. Available from: http://www.ncbi.nlm.nih.gov/ entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list\_uids=9651198

References

16 Novel Prospects in Oxidative and Nitrosative Stress

[1] Frey TG, Mannella CA. The internal structure of mitochondria. Trends in Biochemical Sciences [Internet]. 2000;25(7):319-324. Available from: http://www.ncbi.nlm.nih.gov/ entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list\_uids=10871882 [2] Nicholls DG, Locke RM. Thermogenic mechanisms in brown fat. Physiological Reviews [Internet]. 1984;64(1):1-64. Available from: http://www.ncbi.nlm.nih.gov/entrez/query.

[3] Kowaltowski AJ, Vercesi AE. Mitochondrial damage induced by conditions of oxidative stress. Free Radical Biology & Medicine [Internet]. 1999;26(3–4):463-471. Available from:

[4] Kowaltowski AJ. Alternative mitochondrial functions in cell physiopathology: Beyond ATP production. Brazilian Journal of Medical and Biological Research [Internet]. 2000;33(2):241- 250. Available from: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=-

[5] Lemasters JJ, Nieminen AL, Qian T, Trost LC, Elmore SP, Nishimura Y, et al. The mitochondrial permeability transition in cell death: A common mechanism in necrosis, apoptosis and autophagy. Biochimica et Biophysica Acta [Internet]. 1998;1366(1–2):177-196.

[6] Susin SA, Zamzami N, Kroemer G. Mitochondria as regulators of apoptosis: Doubt no more. Biochimica et Biophysica Acta [Internet]. 1998;1366(1–2):151-165. Available from:

[7] Green DR, Reed JC. Mitochondria and apoptosis. Science (80- ) [Internet]. 1998;281(5381): 1309-1312. Available from: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrie-

[8] Kalyanaraman B, Cheng G, Hardy M, Ouari O, Lopez M, Joseph J, et al. A review of the basics of mitochondrial bioenergetics, metabolism, and related signaling pathways in cancer cells: Therapeutic targeting of tumor mitochondria with lipophilic cationic compounds. Redox Biology [Internet]. 2017/09/29. 2018 Apr;14:316-327. Available from:

[9] Griffiths EJ, Halestrap AP. Mitochondrial non-specific pores remain closed during cardiac ischaemia, but open upon reperfusion. The Biochemical Journal [Internet]. 1995;307(Pt 1):93- 98. Available from: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=Pub

[10] Halestrap AP, Kerr PM, Javadov S, Woodfield KY. Elucidating the molecular mechanism of the permeability transition pore and its role in reperfusion injury of the heart. Biochimica et Biophysica Acta [Internet]. 1998;1366(1–2):79-94. Available from: http:// www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&-

fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list\_uids=6320232

Available from: https://www.ncbi.nlm.nih.gov/pubmed/9714796

https://www.ncbi.nlm.nih.gov/pubmed/9895239

PubMed&dopt=Citation&list\_uids=10657067

https://www.ncbi.nlm.nih.gov/pubmed/9714783

ve&db=PubMed&dopt=Citation&list\_uids=9721092

https://www.ncbi.nlm.nih.gov/pubmed/29017115

Med&dopt=Citation&list\_uids=7717999

list\_uids=9714750


[22] de Oliveira GA. Caloric Restriction and Mitochondria: Role in Saccharomyces cerevisiae aging [doctoral thesis]; 2010. DOI: 10.11606/T.46.2010.tde-01032011-114941. Available from: http://www.teses.usp.br/teses/disponiveis/46/46131/tde-01032011-114941/en.php

[33] Tretter L. Generation of reactive oxygen species in the reaction catalyzed by αketoglutarate dehydrogenase. The Journal of Neuroscience [Internet]. 2004 Sep 8;24(36): 7771-7778. Available from: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrie-

Oxidative Stress: Noxious but Also Vital http://dx.doi.org/10.5772/intechopen.73394 19

[34] Tahara EB, Barros MH, Oliveira GA, Netto LES, Kowaltowski AJ. Dihydrolipoyl dehydrogenase as a source of reactive oxygen species inhibited by caloric restriction and involved in Saccharomyces cerevisiae aging. The FASEB Journal [Internet]. 2006 Nov 29;21(1):274-283.

[35] Thomas DD, Heinecke JL, Ridnour LA, Cheng RY, Kesarwala AH, Switzer CH, et al. Signaling and stress: The redox landscape in NOS2 biology. Free Radical Biology & Medicine [Internet]. 2015 Oct;87:204-225. Available from: http://www.ncbi.nlm.nih.gov/

[36] de Oliveira GA, Cheng RYS, Ridnour LA, Basudhar D, Somasundaram V, McVicar DW, et al. Inducible nitric oxide synthase in the carcinogenesis of gastrointestinal cancers. Antioxidants & Redox Signaling [Internet]. 2016/10/31. 2017 Jun 20;26(18):1059-1077.

[37] Halliwell B, Gutteridge JMC. Free Radicals in Biology and Medicine. 4th ed. Oxford,

[38] Stamler JS. Redox signaling: Nitrosylation and related target interactions of nitric oxide. Cell [Internet]. 1994;78(6):931-936. Available from: https://www.ncbi.nlm.nih.

[39] Chakraborti S, Chakraborti T. Down-regulation of protein kinase C attenuates the oxidant hydrogen peroxide-mediated activation of phospholipase A2 in pulmonary vascular smooth muscle cells. Cellular Signalling [Internet]. 1995;7(1):75-83. Available from:

[40] Tournier C, Thomas G, Pierre J, Jacquemin C, Pierre M, Saunier B. Mediation by arachidonic acid metabolites of the H2O2-induced stimulation of mitogen-activated protein kinases (extracellular-signal-regulated kinase and c-Jun NH2-terminal kinase). European Journal of Biochemistry [Internet]. 1997;244(2):587-595. Available from: https://

[41] Kim JH, Kwack HJ, Choi SE, Kim BC, Kim YS, Kang IJ, et al. Essential role of Rac GTPase in hydrogen peroxide-induced activation of c-fos serum response element. FEBS Letters [Internet]. 1997;406(1–2):93-96. Available from: https://www.ncbi.nlm.nih.gov/pubmed/

[42] Sena LA, Chandel NS. Physiological roles of mitochondrial reactive oxygen species. Molecular Cell [Internet]. 2012 Oct;48(2):158-167. Available from: https://www.ncbi.

[43] Rharass T, Lantow M, Gbankoto A, Weiss DG, Panáková D, Lucas S. Ascorbic acid alters cell fate commitment of human neural progenitors in a WNT/β-catenin/ROS signaling

Available from: http://www.fasebj.org/cgi/doi/10.1096/fj.06-6686com

Available from: https://www.ncbi.nlm.nih.gov/pubmed/27494631

New York: Oxford University Press; 2007. xxxvi, 851 p

https://www.ncbi.nlm.nih.gov/pubmed/7756114

www.ncbi.nlm.nih.gov/pubmed/9119028

nlm.nih.gov/pubmed/23102266

ve&db=PubMed&dopt=Citation&list\_uids=15356188

pubmed/26117324

gov/pubmed/7923362

9109393


[33] Tretter L. Generation of reactive oxygen species in the reaction catalyzed by αketoglutarate dehydrogenase. The Journal of Neuroscience [Internet]. 2004 Sep 8;24(36): 7771-7778. Available from: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list\_uids=15356188

[22] de Oliveira GA. Caloric Restriction and Mitochondria: Role in Saccharomyces cerevisiae aging [doctoral thesis]; 2010. DOI: 10.11606/T.46.2010.tde-01032011-114941. Available from: http://www.teses.usp.br/teses/disponiveis/46/46131/tde-01032011-114941/en.php

[23] Chance B, Sies H, Boveris A. Hydroperoxide metabolism in mammalian organs. Physiological Reviews [Internet]. 1979;59(3):527-605. Available from: https://www.ncbi.nlm.

[24] Turrens JF, Boveris A. Generation of superoxide anion by the NADH dehydrogenase of bovine heart mitochondria. The Biochemical Journal [Internet]. 1980;191(2):421-427. Available from: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=Pub-

[25] Boveris A, Cadenas E, Stoppani AO. Role of ubiquinone in the mitochondrial generation of hydrogen peroxide. The Biochemical Journal [Internet]. 1976;156(2):435-444. Available

[26] Quinlan CL, Gerencser AA, Treberg JR, Brand MD. The mechanism of superoxide production by the antimycin-inhibited mitochondrial Q-cycle. The Journal of Biological Chemistry [Internet]. 2011/06/27. 2011 Sep 9;286(36):31361-31372. Available from: https://www.

[27] Bleier L, Dröse S. Superoxide generation by complex III: From mechanistic rationales to functional consequences. Biochimica et Biophysica Acta (BBA) - Bioenergetics [Internet]. 2012/12/23. 2013 Nov;1827(11–12):1320-1331. Available from: https://www.ncbi.nlm.nih.

[28] Oliveira GA, Kowaltowski AJ. Phosphate increases mitochondrial reactive oxygen species release. Free Radical Research [Internet]. 2004 Oct 7;38(10):1113-1118. Available

[29] Muller FL, Liu Y, Van Remmen H. Complex III releases superoxide to both sides of the inner mitochondrial membrane. The Journal of Biological Chemistry [Internet]. 2004/08/17. 2004 Nov 19;279(47):49064-49073. Available from: https://www.ncbi.nlm.nih.gov/pubmed/

[30] Yankovskaya V. Architecture of succinate dehydrogenase and reactive oxygen species generation. Science (80-) [Internet]. 2003 Jan 31;299(5607):700-704. Available from: https://

[31] Liu Y, Fiskum G, Schubert D. Generation of reactive oxygen species by the mitochondrial electron transport chain. Journal of Neurochemistry [Internet]. 2002;80(5):780-787.

[32] Starkov AA. Mitochondrial alpha-ketoglutarate dehydrogenase complex generates reactive oxygen species. The Journal of Neuroscience [Internet]. 2004 Sep 8;24(36):7779-7788. Available from: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=Pub-

Available from: https://www.ncbi.nlm.nih.gov/pubmed/11948241

from: http://www.tandfonline.com/doi/full/10.1080/10715760400009258

nih.gov/pubmed/37532

18 Novel Prospects in Oxidative and Nitrosative Stress

Med&dopt=Citation&list\_uids=6263247

ncbi.nlm.nih.gov/pubmed/21708945

www.ncbi.nlm.nih.gov/pubmed/12560550

Med&dopt=Citation&list\_uids=15356189

gov/pubmed/23269318

15317809

from: https://www.ncbi.nlm.nih.gov/pubmed/182149


dependent manner. Journal of Biomedical Science [Internet]. 2017/10/16. 2017 Dec 16;24(1):78. Available from: https://www.ncbi.nlm.nih.gov/pubmed/29037191

[55] Harman D. Free radical theory of aging: An update: Increasing the functional life span. Annals of the New York Academy of Sciences [Internet]. 2006 May 1;1067(1):10-21.

Oxidative Stress: Noxious but Also Vital http://dx.doi.org/10.5772/intechopen.73394 21

[56] Barros MH, da Cunha FM, Oliveira GA, Tahara EB, Kowaltowski AJ. Yeast as a model to study mitochondrial mechanisms in ageing. Mechanisms of Ageing and Development [Internet]. 2010 Jul;131(7–8):494-502. Available from: http://www.ncbi.nlm.nih.gov/entrez/

[57] Barros MH, Bandy B, Tahara EB, Kowaltowski AJ. Higher respiratory activity decreases mitochondrial reactive oxygen release and increases life span in Saccharomyces cerevisiae. The Journal of Biological Chemistry [Internet]. 2004 Nov 26;279(48):49883-49888. Available from: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=-

[58] Nisoli E. Calorie restriction promotes mitochondrial biogenesis by inducing the expression of eNOS. Science (80-) [Internet]. 2005 Oct 14;310(5746):314-317. Available from:

[59] Oliveira GA, Tahara EB, Gombert AK, Barros MH, Kowaltowski AJ. Increased aerobic metabolism is essential for the beneficial effects of caloric restriction on yeast life span. Journal of Bioenergetics and Biomembranes [Internet]. 2008 Aug 15;40(4):381-388. Avail-

[60] Caldeira da Silva CC, Cerqueira FM, Barbosa LF, Medeiros MHG, Kowaltowski AJ. Mild mitochondrial uncoupling in mice affects energy metabolism, redox balance and longevity. Aging Cell [Internet]. 2008 Aug;7(4):552-560. Available from: http://www.ncbi.nlm.nih. gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list\_uids=18505478

[61] López-Otín C, Galluzzi L, Freije JMP, Madeo F, Kroemer G. Metabolic control of longevity. Cell [Internet]. 2016 Aug;166(4):802-821. Available from: https://www.ncbi.nlm.nih.

[62] Castilho RF, Kowaltowski AJ, Meinicke A, Bechara EJH, Vercesi AE. Permeabilization of the inner mitochondrial membrane by Ca2+ ions is stimulated by t-butyl hydroperoxide and mediated by reactive oxygen species generated by mitochondria. Free Radical Biology & Medicine [Internet]. 1995 Mar;18(3):479-486. Available from: http://linkinghub.elsevier.

[63] Castilho RF, Kowaltowski AJ, Vercesi AE. The irreversibility of inner mitochondrial membrane permeabilization by Ca2+ plus prooxidants is determined by the extent of membrane protein thiol cross-linking. Journal of Bioenergetics and Biomembranes [Internet]. 1996;28(6):523-529. Available from: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?

[64] Kowaltowski AJ, Castilho RF, Grijalba MT, Bechara EJH, Vercesi AE. Effect of inorganic phosphate concentration on the nature of inner mitochondrial membrane alterations

query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list\_uids=20450928

Available from: http://doi.wiley.com/10.1196/annals.1354.003

Citation&list\_uids=15383542

gov/pubmed/27518560

com/retrieve/pii/089158499400166H

http://www.ncbi.nlm.nih.gov/pubmed/16224023

able from: http://link.springer.com/10.1007/s10863-008-9159-5

cmd=Retrieve&db=PubMed&dopt=Citation&list\_uids=8953384


[55] Harman D. Free radical theory of aging: An update: Increasing the functional life span. Annals of the New York Academy of Sciences [Internet]. 2006 May 1;1067(1):10-21. Available from: http://doi.wiley.com/10.1196/annals.1354.003

dependent manner. Journal of Biomedical Science [Internet]. 2017/10/16. 2017 Dec

16;24(1):78. Available from: https://www.ncbi.nlm.nih.gov/pubmed/29037191

ncbi.nlm.nih.gov/pubmed/6249344

20 Novel Prospects in Oxidative and Nitrosative Stress

www.ncbi.nlm.nih.gov/pubmed/28963982

www.ncbi.nlm.nih.gov/pubmed/25244505

able from: https://www.ncbi.nlm.nih.gov/pubmed/27867086

https://www.ncbi.nlm.nih.gov/pubmed/28212523

nlm.nih.gov/pubmed/28600903

nlm.nih.gov/pubmed/28167130

nih.gov/pubmed/29080524

Citation&list\_uids=2205304

gov/pubmed/13332224

[44] Lesko SA, Lorentzen RJ, Ts'o PO. Role of superoxide in deoxyribonucleic acid strand scission. Biochemistry [Internet]. 1980;19(13):3023-3028. Available from: https://www.

[45] Cadet J, Wagner JR. DNA base damage by reactive oxygen species, oxidizing agents, and UV radiation. Cold Spring Harbor Perspectives in Biology [Internet]. 2013/02/01. 2013 Feb 1;5(2):a012559-a012559. Available from: https://www.ncbi.nlm.nih.gov/pubmed/23378590

[46] Markkanen E. Not breathing is not an option: How to deal with oxidative DNA damage. DNA Repair (Amst) [Internet]. 2017/09/22. 2017 Nov;59:82-105. Available from: https://

[47] Samardzic K, Rodgers KJ. Oxidised protein metabolism: Recent insights. Biological Chemistry [Internet]. 2017 Jan 26;398(11):1165-1175. Available from: https://www.ncbi.

[48] Eckl PM, Bresgen N. Genotoxicity of lipid oxidation compounds. Free Radical Biology & Medicine [Internet]. 2017/02/05. 2017 Oct;111:244-252. Available from: https://www.ncbi.

[49] Cheignon C, Tomas M, Bonnefont-Rousselot D, Faller P, Hureau C, Collin F. Oxidative stress and the amyloid beta peptide in Alzheimer's disease. Redox Biology [Internet]. 2017/10/18. 2017 Oct;14:450-464. Available from: https://www.ncbi.nlm.

[50] Li H, Horke S, Förstermann U. Vascular oxidative stress, nitric oxide and atherosclerosis. Atherosclerosis [Internet]. 2014/09/09. 2014 Nov;237(1):208-219. Available from: https://

[51] Lucas AM, Caldas FR, da Silva AP, Ventura MM, Leite IM, Filgueiras AB, et al. Diazoxide prevents reactive oxygen species and mitochondrial damage, leading to anti-hypertrophic effects. Chemico-Biological Interactions [Internet]. 2016/11/17. 2017 Jan;261:50-55. Avail-

[52] Griffiths HR, Gao D, Pararasa C. Redox regulation in metabolic programming and inflammation. Redox Biology [Internet]. 2017/02/12. 2017 Aug;12:50-57. Available from:

[53] Medvedev ZA. An attempt at a rational classification of theories of ageing. Biological Reviews of the Cambridge Philosophical Society [Internet]. 1990;65(3):375-398. Available from: http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=-

[54] Harman D. Aging: A theory based on free radical and radiation chemistry. Journal of Gerontology [Internet]. 1956;11(3):298-300. Available from: https://www.ncbi.nlm.nih.


mediated by Ca ions. The Journal of Biological Chemistry [Internet]. 1996 Feb 9;271(6):2929- 2934. Available from: http://www.jbc.org/lookup/doi/10.1074/jbc.271.6.2929

[76] Poole LB, Karplus PA, Claiborne A. Protein sulfenic acids in redox signaling. Annual Review of Pharmacology and Toxicology [Internet]. 2004 Feb 10;44(1):325-347. Available from: http://www.annualreviews.org/doi/10.1146/annurev.pharmtox.44.101802.

Oxidative Stress: Noxious but Also Vital http://dx.doi.org/10.5772/intechopen.73394 23

[77] Berndt C, Lillig CH, Holmgren A. Thiol-based mechanisms of the thioredoxin and glutaredoxin systems: Implications for diseases in the cardiovascular system. American Journal of Physiology. Heart and Circulatory Physiology [Internet]. 2006 Oct 20;292(3): H1227-H1236. Available from: http://ajpheart.physiology.org/cgi/doi/10.1152/ajpheart.

[78] Cooke MS. Oxidative DNA damage: Mechanisms, mutation, and disease. The FASEB Journal [Internet]. 2003 Jul 1;17(10):1195-1214. Available from: http://www.fasebj.org/

[79] Evans MD, Dizdaroglu M, Cooke MS. Oxidative DNA damage and disease: Induction, repair and significance. Mutation Research. 2004 Sep;567(1):1-61. Available from: http://

[80] Dizdaroglu M. Oxidatively induced DNA damage: Mechanisms, repair and disease. Cancer Letters [Internet]. 2012 Dec;327(1–2):26-47. Available from: http://dx.doi.org/

[81] Greenman C, Stephens P, Smith R, Dalgliesh GL, Hunter C, Bignell G, et al. Patterns of somatic mutation in human cancer genomes. Nature [Internet]. 2007 Mar 8;446(7132):153- 158. Available from: http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=2712719

[82] Olivier M, Hollstein M, Hainaut P. TP53 mutations in human cancers: Origins, consequences, and clinical use. Cold Spring Harbor Perspectives in Biology [Internet]. 2010 Jan 1;2(1):a001008-a001008. Available from: http://cshperspectives.cshlp.org/lookup/doi/

[83] Girotti AW. Action in biological systems. Journal of Lipid Research. 1998;39:1529-1542 [84] Shigenaga MK, Hagen TM, Ames BN. Oxidative damage and mitochondrial decay in aging. Proceedings of the National Academy of Sciences [Internet]. 1994 Nov 8;91(23):10771- 10778. Available from: http://www.ncbi.nlm.nih.gov/pubmed/7971961%5Cnhttp://www.

[85] Dianzani MU. 4-Hydroxynonenal from pathology to physiology. Molecular Aspects of Medicine [Internet]. 2003 Aug;24(4–5):263-272. Available from: http://linkinghub.elsevier.

[86] Porter NA, Caldwell SE, Mills KA. Mechanisms of free radical oxidation of unsaturated lipids. Lipids [Internet]. 1995 Apr;30(4):277-290. Available from: http://link.springer.

[87] Lorrain B, Dangles O, Loonis M, Armand M, Dufour C. Dietary iron-initiated lipid oxidation and its inhibition by polyphenols in gastric conditions. Journal of Agricultural

linkinghub.elsevier.com/retrieve/pii/S138357420300139X

pubmedcentral.nih.gov/articlerender.fcgi?artid=PMC45108

121735

01162.2006

cgi/doi/10.1096/fj.02-0752rev

10.1016/j.canlet.2012.01.016

10.1101/cshperspect.a001008

com/retrieve/pii/S0098299703000219

com/10.1007/BF02536034

&tool=pmcentrez&rendertype=abstract


[76] Poole LB, Karplus PA, Claiborne A. Protein sulfenic acids in redox signaling. Annual Review of Pharmacology and Toxicology [Internet]. 2004 Feb 10;44(1):325-347. Available from: http://www.annualreviews.org/doi/10.1146/annurev.pharmtox.44.101802. 121735

mediated by Ca ions. The Journal of Biological Chemistry [Internet]. 1996 Feb 9;271(6):2929-

[65] Kowaltowski AJ, Castilho RF, Vercesi AE. Opening of the mitochondrial permeability transition pore by uncoupling or inorganic phosphate in the presence of Ca2+ is dependent on mitochondrial-generated reactive oxygen species. FEBS Letters. 1996;378:150-152.

[66] Costa RAP, Romagna CD, Pereira JL, Souza-Pinto NC. The role of mitochondrial DNA damage in the citotoxicity of reactive oxygen species. Journal of Bioenergetics and Biomembranes [Internet]. 2011 Feb 1;43(1):25-29. Available from: http://link.springer.

[67] Mohiuddin I, Chai H, Lin PH, Lumsden AB, Yao Q, Chen C. Nitrotyrosine and chlorotyrosine: Clinical significance and biological functions in the vascular system. The Journal of Surgical Research [Internet]. 2006 Jun;133(2):143-149. Available from:

[68] Viappiani S. Detection of specific nitrotyrosine-modified proteins as a marker of oxidative stress in cardiovascular disease. American Journal of Physiology. Heart and Circulatory Physiology [Internet]. 2006 Jun 1;290(6):H2167-H2168. Available from: http://

[69] Pacher P, Beckman JS, Liaudet L. Nitric oxide and peroxynitrite in health and disease. Physiological Reviews [Internet]. 2007 Jan 1;87(1):315-424. Available from: http://physrev.

[70] Wall SB, Oh J-Y, Diers AR, Landar A. Oxidative modification of proteins: An emerging mechanism of cell signaling. Frontiers in Physiology [Internet]. 2012;3(September):1-9. Available from: http://journal.frontiersin.org/article/10.3389/fphys.2012.00369/abstract

[71] Bridge G, Rashid S, Martin S. DNA mismatch repair and oxidative DNA damage: Implications for cancer biology and treatment. Cancers (Basel) [Internet]. 2014 Aug

[72] Ohkawa H, Ohishi N, Yagi K. Assay for lipid peroxides in animal tissues by thiobarbituric acid reaction. Analytical Biochemistry [Internet]. 1979 Jun;95(2):351-358. Available from:

[73] Lü J-M, Lin PH, Yao Q, Chen C. Chemical and molecular mechanisms of antioxidants: Experimental approaches and model systems. Journal of Cellular and Molecular Medicine [Internet]. 2010 Apr;14(4):840-860. Available from: http://doi.wiley.com/10.1111/

[74] Tu BP, Weissman JS. Oxidative protein folding in eukaryotes: Figure 1. The Journal of Cell Biology [Internet]. 2004 Feb 2;164(3):341-346. Available from: http://www.jcb.org/

[75] Jones DP. Cysteine/cystine couple is a newly recognized node in the circuitry for biologic redox signaling and control. The FASEB Journal [Internet]. 2004 Jun 18;18:1246-

1248. Available from: http://www.fasebj.org/cgi/doi/10.1096/fj.03-0971fje

5;6(3):1597-1614. Available from: http://www.mdpi.com/2072-6694/6/3/1597/

http://linkinghub.elsevier.com/retrieve/pii/S0022480405005445

ajpheart.physiology.org/cgi/doi/10.1152/ajpheart.00128.2006

http://linkinghub.elsevier.com/retrieve/pii/0003269779907383

j.1582-4934.2009.00897.x

lookup/doi/10.1083/jcb.200311055

physiology.org/cgi/doi/10.1152/physrev.00029.2006

2934. Available from: http://www.jbc.org/lookup/doi/10.1074/jbc.271.6.2929

com/10.1007/s10863-011-9329-8

22 Novel Prospects in Oxidative and Nitrosative Stress


and Food Chemistry [Internet]. 2012 Sep 12;60(36):9074-9081. Available from: http:// pubs.acs.org/doi/abs/10.1021/jf302348s

[98] Lei XG, Zhu J-H, Cheng W-H, Bao Y, Ho Y-S, Reddi AR, et al. Paradoxical roles of antioxidant enzymes: Basic mechanisms and health implications. Physiological Reviews [Internet]. 2016 Jan 17;96(1):307-364. Available from: http://physrev.physiology.org/

Oxidative Stress: Noxious but Also Vital http://dx.doi.org/10.5772/intechopen.73394 25

[99] Dey S, Sidor A, O'Rourke B. Compartment-specific control of reactive oxygen species scavenging by antioxidant pathway enzymes. The Journal of Biological Chemistry [Internet]. 2016 May 20;291(21):11185-11197. Available from: http://www.jbc.org/lookup/doi/

[100] Marrocco I, Altieri F, Peluso I. Measurement and clinical significance of biomarkers of oxidative stress in humans. Oxidative Medicine and Cellular Longevity [Internet]. 2017;2017:1-32. Available from: https://www.hindawi.com/journals/omcl/2017/6501046/

[101] Barbosa KBF, Costa NMB, Alfenas R de CG, De Paula SO, Minim VPR, Bressan J. Estresse oxidativo: conceito, implicações e fatores modulatórios. Revista de Nutrição [Internet]. 2010 Aug;23(4):629-643. Available from: http://www.scielo.br/scielo.php?script=sci\_art-

[102] Manganaris GA, Goulas V, Vicente AR, Terry LA. Berry antioxidants: Small fruits providing large benefits. Journal of the Science of Food and Agriculture [Internet]. 2014 Mar

[103] King A, Young G. Characteristics and occurrence of phenolic phytochemicals. Journal of the American Dietetic Association [Internet]. 1999 Feb;99(2):213-218. Available from:

[104] Angelo PM, Jorge N. Compostos fenólicos em alimentos – Uma breve revisão Phenolic compounds in foods – A brief review. Revista do Instituto Adolfo Lutz. 2007;66(1):1-9

[105] Fan G-J, Jin X-L, Qian Y-P, Wang Q, Yang R-T, Dai F, et al. Hydroxycinnamic acids as DNA-cleaving agents in the presence of Cu II ions: Mechanism, structure-activity relationship, and biological implications. Chemistry - A European Journal [Internet]. 2009 Nov 23;15(46):12889-12899. Available from: http://doi.wiley.com/10.1002/chem.200901627 [106] Gülçin İ. Antioxidant activity of food constituents: An overview. Archives of Toxicology [Internet]. 2012 Mar 20;86(3):345-391. Available from: http://link.springer.com/10.1007/

[107] Majewska-Wierzbicka M, Czeczot H. Flavonoids in the prevention and treatment of

[108] Moreira AVB, Mancini Filho J. Atividade antioxidante das especiarias mostarda, canela

[109] da Rosa JS, Godoy RL de O, Oiano Neto J, et al. Desenvolvimento de um método de análise de vitamina C em alimentos por cromatografa líquida de alta eficiência e exclusão iônica. Ciência e Tecnologia de Alimentos [Internet]. 2007 Dec;27(4):837-846. Available from: http:// www.scielo.br/scielo.php?script=sci\_arttext&pid=S0101-20612007000400025&lng=en&nrm=

cardiovascular diseases. Polski Merkuriusz Lekarski. 2012;32(188):50-54

e erva-doce em sistemas aquoso e lipídico. Nutrire. 2003;25:31-46

30;94(5):825-833. Available from: http://doi.wiley.com/10.1002/jsfa.6432

http://linkinghub.elsevier.com/retrieve/pii/S0002822399000516

lookup/doi/10.1152/physrev.00010.2014

text&pid=S1415-52732010000400013&lng=pt&tlng=pt

10.1074/jbc.M116.726968

s00204-011-0774-2

iso&tlng=pt


[98] Lei XG, Zhu J-H, Cheng W-H, Bao Y, Ho Y-S, Reddi AR, et al. Paradoxical roles of antioxidant enzymes: Basic mechanisms and health implications. Physiological Reviews [Internet]. 2016 Jan 17;96(1):307-364. Available from: http://physrev.physiology.org/ lookup/doi/10.1152/physrev.00010.2014

and Food Chemistry [Internet]. 2012 Sep 12;60(36):9074-9081. Available from: http://

[88] Getzoff ED, Tainer JA, Weiner PK, Kollman PA, Richardson JS, Richardson DC. Electrostatic recognition between superoxide and copper, zinc superoxide dismutase. Nature [Internet]. 1983 Nov 17;306(5940):287-290. Available from: http://www.nature.com/

[89] Esterbauer H, Schaur RJ, Zollner H. Chemistry and biochemistry of 4-hydroxynonenal, malonaldehyde and related aldehydes. Free Radical Biology & Medicine [Internet]. 1991 Jan;11(1):81-128. Available from: http://linkinghub.elsevier.com/retrieve/pii/0891584991

[90] Kadiiska MB, Basu S, Brot N, Cooper C, Saari Csallany A, Davies MJ, et al. Biomarkers of oxidative stress study V: Ozone exposure of rats and its effect on lipids, proteins, and DNA in plasma and urine. Free Radical Biology & Medicine [Internet]. 2013 Aug;61:408-415. Available from: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3968235/%5Cnhttp://www. ncbi.nlm.nih.gov/pmc/articles/PMC3968235/pdf/nihms525937.pdf%5Cnhttp://www.ncbi.nl

[91] Kourie JI. Interaction of reactive oxygen species with ion transport mechanisms. The American Journal of Physiology. 1998 Nov;275(1):1-11. Available from: http://ajpcell.

[92] Catalá A. Lipid peroxidation of membrane phospholipids generates hydroxy-alkenals and oxidized phospholipids active in physiological and/or pathological conditions. Chemistry and Physics of Lipids [Internet]. 2009 Jan;157(1):1-11. Available from: http://

[93] Siegel MP, Kruse SE, Knowels G, Salmon A, Beyer R, Xie H, et al. Reduced coupling of oxidative phosphorylation in vivo precedes electron transport chain defects due to mild oxidative stress in mice. Vina J, editor. PLoS One [Internet]. 2011 Nov 22;6(11):e26963.

[94] Rahal A, Kumar A, Singh V, Yadav B, Tiwari R, Chakraborty S, et al. Oxidative stress, prooxidants, and antioxidants: The interplay. BioMed Research International [Internet]. 2014;2014:1-19. Available from: http://www.hindawi.com/journals/bmri/2014/761264/

[95] Mirończuk-Chodakowska I, Witkowska AM, Zujko ME. Endogenous non-enzymatic antioxidants in the human body. Advances in Medical Sciences [Internet]. 2018 Mar;63 (1):68-78. Available from: http://linkinghub.elsevier.com/retrieve/pii/S1896112617300445

[96] Godic A, Poljšak B, Adamic M, Dahmane R. The role of antioxidants in skin cancer prevention and treatment. Oxidative Medicine and Cellular Longevity [Internet]. 2014;2014:1-6. Available from: http://www.hindawi.com/journals/omcl/2014/860479/ [97] Pisoschi AM, Pop A. The role of antioxidants in the chemistry of oxidative stress: A review. European Journal of Medicinal Chemistry [Internet]. 2015 Jun;97:55-74. Avail-

pubs.acs.org/doi/abs/10.1021/jf302348s

doifinder/10.1038/306287a0

24 Novel Prospects in Oxidative and Nitrosative Stress

m.nih.gov/pubmed/23608465

physiology.org/cgi/doi/10.1152/ajpcell.00167.2002

linkinghub.elsevier.com/retrieve/pii/S0009308408003708

able from: http://dx.doi.org/10.1016/j.ejmech.2015.04.040

Available from: http://dx.plos.org/10.1371/journal.pone.0026963

901926


[110] Prior RL. Oxygen radical absorbance capacity (ORAC): New horizons in relating dietary antioxidants/bioactives and health benefits. Journal of Functional Foods [Internet]. 2015 Oct;18:797-810. Available from: http://dx.doi.org/10.1016/j.jff.2014.12.018

[122] Park K-R, Nam D, Yun H-M, Lee S-G, Jang H-J, Sethi G, et al. β-Caryophyllene oxide inhibits growth and induces apoptosis through the suppression of PI3K/AKT/mTOR/ S6K1 pathways and ROS-mediated MAPKs activation. Cancer Letters [Internet]. 2011

Oxidative Stress: Noxious but Also Vital http://dx.doi.org/10.5772/intechopen.73394 27

Dec;312(2):178-188. Available from: http://dx.doi.org/10.1016/j.canlet.2011.08.001 [123] Zhang J, Wang X, Vikash V, Ye Q, Wu D, Liu Y, et al. ROS and ROS-mediated cellular signaling. Oxidative Medicine and Cellular Longevity [Internet]. 2016;2016(Figure 1):1-

[124] Franke TF, Kaplan DR, Cantley LC. PI3K: Downstream AKTion blocks apoptosis. Cell [Internet]. 1997 Feb;88(4):435-437. Available from: http://linkinghub.elsevier.com/

[125] Parsons SJ, Parsons JT. Src family kinases, key regulators of signal transduction. Oncogene [Internet]. 2004 Oct 18;23(48):7906-7909. Available from: http://www.nature.com/

[126] Mu W, Liu L-Z. Reactive oxygen species signaling in cancer development. Reactive Oxygen Species [Internet]. 2017;2(1):219-230. Available from: https://www.aimsci.com/

[127] Jones RJ, Brunton VG, Frame MC. Adhesion-linked kinases in cancer; emphasis on Src, focal adhesion kinase and PI 3-kinase. European Journal of Cancer [Internet]. 2000 Aug;36(13):1595-1606. Available from: http://linkinghub.elsevier.com/retrieve/pii/S09598

[128] Lai YH, Chen MH, Lin SY, Lin SY, Wong YH, Yu SL, et al. Rhodomycin A, a novel Srctargeted compound, can suppress lung cancer cell progression via modulating Src-related pathways. Oncotarget [Internet]. 2015;6(28):26252-26265. Available from: http://ovidsp. ovid.com/ovidweb.cgi?T=JS&CSC=Y&NEWS=N&PAGE=fulltext&D=medl&AN=2631276 6%5Cnhttp://nt2yt7px7u.search.serialssolutions.com/?sid=OVID:Ovid+MEDLINE%28R% 29+%3C2013+to+April+Week+3+2017%3E&genre=article&id=pmid:26312766&id=doi:10.1

[129] Patel A, Sabbineni H, Clarke A, Somanath PR. Novel roles of Src in cancer cell epithelialto-mesenchymal transition, vascular permeability, microinvasion and metastasis. Life Sciences [Internet]. 2016 Jul;157:52-61. Available from: http://linkinghub.elsevier.com/

[130] Lin S-Y, Chang H-H, Lai Y-H, Lin C-H, Chen M-H, Chang G-C, et al. Digoxin suppresses tumor malignancy through inhibiting multiple Src-related signaling pathways in nonsmall cell lung cancer. Chellappan SP, editor. PLoS One [Internet]. 2015 May 8;10(5):

[131] Zuo Z, Cai T, Li J, Zhang D, Yu Y, Huang C. Hexavalent chromium Cr(VI) up-regulates COX-2 expression through an NFκB/c-Jun/AP-1–dependent pathway. Environmental Health Perspectives [Internet]. 2012 Jan 6;120(4):547-553. Available from: http://www.

e0123305. Available from: http://dx.doi.org/10.1371/journal.pone.0123305

18. Available from: http://www.hindawi.com/journals/omcl/2016/4350965/

retrieve/pii/S0092867400818838

doifinder/10.1038/sj.onc.1208160

ros/index.php/ros/article/view/95

04900001532

8632%2Foncotarge

retrieve/pii/S0024320516303344

ncbi.nlm.nih.gov/pubmed/22472290


[122] Park K-R, Nam D, Yun H-M, Lee S-G, Jang H-J, Sethi G, et al. β-Caryophyllene oxide inhibits growth and induces apoptosis through the suppression of PI3K/AKT/mTOR/ S6K1 pathways and ROS-mediated MAPKs activation. Cancer Letters [Internet]. 2011 Dec;312(2):178-188. Available from: http://dx.doi.org/10.1016/j.canlet.2011.08.001

[110] Prior RL. Oxygen radical absorbance capacity (ORAC): New horizons in relating dietary antioxidants/bioactives and health benefits. Journal of Functional Foods [Internet]. 2015

[111] Vannucchi H, Melo SS. Hiper-homocisteinemia e risco cardiometabólico. Arquivos

[112] Rodriguez-Amaya DB. Food Carotenoids: Chemistry, Biology, and Technology. IFT Press/

[113] Morita M, Naito Y, Yoshikawa T, Niki E. Rapid assessment of singlet oxygen-induced plasma lipid oxidation and its inhibition by antioxidants with diphenyl-1-pyrenylphosphine (DPPP). Analytical and Bioanalytical Chemistry [Internet]. 2016 Jan 14;408(1):265-

[114] Takahashi S, Iwasaki-Kino Y, Aizawa K, Terao J, Mukai K. Development of a Singlet Oxygen Absorption Capacity (SOAC) assay method. Measurements of the SOAC values for carotenoids and α-tocopherol in an aqueous Triton X-100 micellar solution. Journal of Agricultural and Food Chemistry. 2017 Feb;65(4):784-792. Available from: http://pubs.

[115] Thomas B, Murray BG, Murphy DJ. Encyclopedia of Applied Plant Sciences. Elsevier, 2;

[116] El-Agamey A, Lowe GM, McGarvey DJ, Mortensen A, Phillip DM, Truscott TG, et al. Carotenoid radical chemistry and antioxidant/pro-oxidant properties. Archives of Biochemistry and Biophysics [Internet]. 2004 Oct;430(1):37-48. Available from: http://

[117] Leonard SS, Harris GK, Shi X. Metal-induced oxidative stress and signal transduction. Free Radical Biology & Medicine [Internet]. 2004 Dec;37(12):1921-1942. Available from:

[118] Brown DI, Griendling KK. Regulation of signal transduction by reactive oxygen species in the cardiovascular system. Circulation Research [Internet]. 2015 Jan 30;116(3):531-549. Available from: http://circres.ahajournals.org/cgi/doi/10.1161/CIRCRESAHA.116.303584

[119] Nakanishi A. Link between PI3K/AKT/PTEN pathway and NOX protein in diseases. Aging and Disease [Internet]. 2014 Jun 1;5(3):203. Available from: http://www.ncbi.nlm. nih.gov/pubmed/24900943%5Cnhttp://www.pubmedcentral.nih.gov/articlerender.fcgi?

[120] Alessi DR, Andjelkovic M, Caudwell B, Cron P, Morrice N, Cohen P, et al. Mechanism of activation of protein kinase B by insulin and IGF-1. The EMBO Journal. 1996;15(23):6541-

[121] Gào X, Schöttker B. Reduction–oxidation pathways involved in cancer development: A systematic review of literature reviews. Oncotarget [Internet]. 2017 Jul 31. Available

6551. Available from: http://www.ncbi.nlm.nih.gov/pubmed/8978681

from: http://www.oncotarget.com/fulltext/17128

Oct;18:797-810. Available from: http://dx.doi.org/10.1016/j.jff.2014.12.018

270. Available from: http://link.springer.com/10.1007/s00216-015-9102-7

Brasileiros de Endocrinologia e Metabologia. 2009;53(5):540-549

Wiley Blackwell; 2016

26 Novel Prospects in Oxidative and Nitrosative Stress

2017

artid=PMC4037312

acs.org/doi/abs/10.1021/acs.jafc.6b04329

linkinghub.elsevier.com/retrieve/pii/S0003986104001468

http://linkinghub.elsevier.com/retrieve/pii/S0891584904007191


[132] Lian S, Xia Y, Khoi PN, Ung TT, Yoon HJ, Kim NH, et al. Cadmium induces matrix metalloproteinase-9 expression via ROS-dependent EGFR, NF-кB, and AP-1 pathways in human endothelial cells. Toxicology [Internet]. 2015 Dec;338:104-116. Available from: http://dx.doi.org/10.1016/j.tox.2015.10.008

Journal of Immunology [Internet]. 2010 Feb 1;184(3):1507-1515. Available from: http://

Oxidative Stress: Noxious but Also Vital http://dx.doi.org/10.5772/intechopen.73394 29

[143] Sun GY, Chuang DY, Zong Y, Jiang J, Lee JCM, Gu Z, et al. Role of cytosolic phospholipase A2 in oxidative and inflammatory signaling pathways in different cell types in the central nervous system. Nixon AE, editor. Molecular Neurobiology [Internet]. 2014 Aug

27;50(1):6-14. Available from: http://link.springer.com/10.1007/978-1-62703-673-3 [144] Suh P-G, Park J-I, Manzoli L, Cocco L, Peak JC, Katan M, et al. Multiple roles of phosphoinositide-specific phospholipase C isozymes. BMB Reports [Internet]. 2008 Jun 30;41(6):415-434. Available from: http://koreascience.or.kr/journal/view.jsp?kj=E1MBB7&p

[145] Wang X-T, McCullough KD, Wang X-J, Carpenter G, Holbrook NJ. Oxidative stressinduced phospholipase C-γ1 activation enhances cell survival. The Journal of Biological Chemistry [Internet]. 2001 Jul 27;276(30):28364-28371. Available from: http://www.jbc.

[146] Bai X-C, Deng F, Liu A-L, Zou Z-P, Wang Y, Ke Z-Y, et al. Phospholipase C-γ1 is required for cell survival in oxidative stress by protein kinase C. The Biochemical Journal [Internet]. 2002 Apr 15;363(2):395. Available from: http://www.biochemj.org/bj/363/bj3630395.htm

[147] Kolesnikov YS, Nokhrina KP, Kretynin SV, Volotovski ID, Martinec J, Romanov GA, et al. Molecular structure of phospholipase D and regulatory mechanisms of its activity in plant and animal cells. The Biochemist [Internet]. 2012 Jan 28;77(1):1-14. Available

[148] Kim J, Min G, Bae Y-S, Min DS. Phospholipase D is involved in oxidative stress-induced migration of vascular smooth muscle cells via tyrosine phosphorylation and protein kinase C. Experimental & Molecular Medicine. 2004;36(2):103-109. Available from:

[149] Ryan KA, Smith MF, Sanders MK, Ernst PB. Reactive oxygen and nitrogen species differentially regulate Toll-like receptor 4-mediated activation of NF-kappa B and interleukin-8 expression. Infection and Immunity [Internet]. 2004 Apr 1;72(4):2123-2130.

[150] Bala A, Mondal C, Haldar PK, Khandelwal B. Oxidative stress in inflammatory cells of patient with rheumatoid arthritis: Clinical efficacy of dietary antioxidants. Inflammopharmacology [Internet]. 2017 Dec;25(6):595-607. Available from: http://link.springer.

[151] Datta S, Kundu S, Ghosh P, De S, Ghosh A, Chatterjee M. Correlation of oxidant status with oxidative tissue damage in patients with rheumatoid arthritis. Clinical Rheumatology [Internet]. 2014 Nov 10;33(11):1557-1564. Available from: http://www.ncbi.

[152] Ricciotti E, FitzGerald GA. Prostaglandins and inflammation. Arteriosclerosis, Thrombosis, and Vascular Biology [Internet]. 2011 May 1;31(5):986-1000. Available from: http://

atvb.ahajournals.org/cgi/doi/10.1161/ATVBAHA.110.207449

Available from: http://iai.asm.org/cgi/doi/10.1128/IAI.72.4.2123-2130.2004

www.jimmunol.org/cgi/doi/10.4049/jimmunol.0901219

y=2008&vnc=v41n6&sp=415

org/lookup/doi/10.1074/jbc.M102693200

from: http://www.ncbi.nlm.nih.gov/pubmed/22339628

http://www.ncbi.nlm.nih.gov/pubmed/15150437

com/10.1007/s10787-017-0397-1

nlm.nih.gov/pubmed/12377764


Journal of Immunology [Internet]. 2010 Feb 1;184(3):1507-1515. Available from: http:// www.jimmunol.org/cgi/doi/10.4049/jimmunol.0901219

[143] Sun GY, Chuang DY, Zong Y, Jiang J, Lee JCM, Gu Z, et al. Role of cytosolic phospholipase A2 in oxidative and inflammatory signaling pathways in different cell types in the central nervous system. Nixon AE, editor. Molecular Neurobiology [Internet]. 2014 Aug 27;50(1):6-14. Available from: http://link.springer.com/10.1007/978-1-62703-673-3

[132] Lian S, Xia Y, Khoi PN, Ung TT, Yoon HJ, Kim NH, et al. Cadmium induces matrix metalloproteinase-9 expression via ROS-dependent EGFR, NF-кB, and AP-1 pathways in human endothelial cells. Toxicology [Internet]. 2015 Dec;338:104-116. Available from:

[133] Landmesser U, Dikalov S, Price SR, McCann L, Fukai T, Holland SM, et al. Oxidation of tetrahydrobiopterin leads to uncoupling of endothelial cell nitric oxide synthase in hypertension. The Journal of Clinical Investigation [Internet]. 2003 Apr 15;111(8):1201-

[134] Chen C-A, De Pascali F, Basye A, Hemann C, Zweier JL. Redox modulation of endothelial nitric oxide synthase by glutaredoxin-1 through reversible oxidative posttranslational modification. Biochemistry [Internet]. 2013 Sep 24;52(38):6712-6723. Avail-

[135] Cai H, Li Z, Dikalov S, Holland SM, Hwang J, Jo H, et al. NAD(P)H oxidase-derived hydrogen peroxide mediates endothelial nitric oxide production in response to angiotensin II. The Journal of Biological Chemistry [Internet]. 2002 Dec 13;277(50):48311-

[136] Burgoyne JR, Prysyazhna O, Rudyk O, Eaton P. CGMP-dependent activation of protein kinase g precludes disulfide activation: Implications for blood pressure control. Hyper-

[137] Jin L, Ying Z, Webb R. Activation of Rho/Rho kinase signaling pathway by reactive oxygen species in rat aorta. American Journal of Physiology. Heart and Circulatory

[138] Aghajanian A, Wittchen ES, Campbell SL, Burridge K. Direct activation of RhoA by reactive oxygen species requires a redox-sensitive motif. Bezanilla M, editor. PLoS One [Internet]. 2009 Nov 26;4(11):e8045. Available from: http://dx.plos.org/10.1371/journal.

[139] Sun H, Tonks NK. The coordinated action of protein tyrosine phosphatases and kinases in cell signaling. Trends in Biochemical Sciences [Internet]. 1994 Nov;19(11):480-485.

[140] Gerthoffer WT. Mechanisms of vascular smooth muscle cell migration. Circulation Research [Internet]. 2007 Mar 16;100(5):607-621. Available from: http://circres.ahajournals.

[141] Chiarugi P, Fiaschi T, Taddei ML, Talini D, Giannoni E, Raugei G, et al. Two vicinal cysteines confer a peculiar redox regulation to low molecular weight protein tyrosine phosphatase in response to platelet-derived growth factor receptor stimulation. The Journal of Biological Chemistry [Internet]. 2001 Sep 7;276(36):33478-33487. Available

[142] Ayilavarapu S, Kantarci A, Fredman G, Turkoglu O, Omori K, Liu H, et al. Diabetesinduced oxidative stress is mediated by Ca2+-independent phospholipase A2 in neutrophils.

Available from: http://linkinghub.elsevier.com/retrieve/pii/0968000494901341

48317. Available from: http://www.ncbi.nlm.nih.gov/pubmed/12377764

http://dx.doi.org/10.1016/j.tox.2015.10.008

28 Novel Prospects in Oxidative and Nitrosative Stress

tension. 2012;60(5):1301-1308

pone.0008045

Physiology. 2004;287(4):H1495-H1500

org/cgi/doi/10.1161/01.RES.0000258492.96097.47

from: http://www.jbc.org/lookup/doi/10.1074/jbc.M102302200

1209. Available from: http://www.jci.org/articles/view/14172

able from: http://pubs.acs.org/doi/abs/10.1021/bi400404s


[153] Sarkar D, Saha P, Gamre S, Bhattacharjee S, Hariharan C, Ganguly S, et al. Antiinflammatory effect of allylpyrocatechol in LPS-induced macrophages is mediated by suppression of iNOS and COX-2 via the NF-κB pathway. International Immunopharmacology [Internet]. 2008 Sep;8(9):1264-1271. Available from: http://linkinghub.elsevier. com/retrieve/pii/S1567576908001562

rapid induction of metallothionein-1 and -2 in response to cerebral ischemia and reperfusion. The Journal of Neuroscience [Internet]. 2000;20(14):5200-5207. Available from:

Oxidative Stress: Noxious but Also Vital http://dx.doi.org/10.5772/intechopen.73394 31

[165] Yao J-W, Liu J, Kong X-Z, Zhang S-G, Wang X-H, Yu M, et al. Induction of activation of the antioxidant response element and stabilization of Nrf2 by 3-(3-pyridylmethylidene)- 2-indolinone (PMID) confers protection against oxidative stress-induced cell death. Toxicology and Applied Pharmacology [Internet]. 2012 Mar;259(2):227-235. Available from:

[166] Hara H. Increase of antioxidative potential by tert-butylhydroquinone protects against cell death associated with 6-hydroxydopamine-induced oxidative stress in neuroblastoma SH-SY5Y cells. Molecular Brain Research [Internet]. 2003 Nov 26;119(2):125-131.

Available from: http://linkinghub.elsevier.com/retrieve/pii/S0169328X03003462

http://www.ncbi.nlm.nih.gov/pubmed/10884303

http://dx.doi.org/10.1016/j.taap.2011.12.027


rapid induction of metallothionein-1 and -2 in response to cerebral ischemia and reperfusion. The Journal of Neuroscience [Internet]. 2000;20(14):5200-5207. Available from: http://www.ncbi.nlm.nih.gov/pubmed/10884303

[165] Yao J-W, Liu J, Kong X-Z, Zhang S-G, Wang X-H, Yu M, et al. Induction of activation of the antioxidant response element and stabilization of Nrf2 by 3-(3-pyridylmethylidene)- 2-indolinone (PMID) confers protection against oxidative stress-induced cell death. Toxicology and Applied Pharmacology [Internet]. 2012 Mar;259(2):227-235. Available from: http://dx.doi.org/10.1016/j.taap.2011.12.027

[153] Sarkar D, Saha P, Gamre S, Bhattacharjee S, Hariharan C, Ganguly S, et al. Antiinflammatory effect of allylpyrocatechol in LPS-induced macrophages is mediated by suppression of iNOS and COX-2 via the NF-κB pathway. International Immunopharmacology [Internet]. 2008 Sep;8(9):1264-1271. Available from: http://linkinghub.elsevier.

[154] Cairns RA, Harris IS, Mak TW. Regulation of cancer cell metabolism. Nature Reviews. Cancer [Internet]. 2011 Feb;11(2):85-95. Available from: http://www.nature.com/doifinder/

[155] Wang W, Zhang E, Lin C. MicroRNAs in tumor angiogenesis. Life Sciences [Internet]. 2015 Sep;136:28-35. Available from: http://dx.doi.org/10.1016/j.lfs.2015.06.025

[156] Hu Y, Deng H, Xu S, Zhang J. MicroRNAs regulate mitochondrial function in cerebral ischemia-reperfusion injury. International Journal of Molecular Sciences [Internet]. 2015 Oct 20;16(10):24895-24917. Available from: http://www.mdpi.com/1422-0067/16/10/24895/

[157] Zhang X, Ng W-L, Wang P, Tian L, Werner E, Wang H, et al. MicroRNA-21 modulates the levels of reactive oxygen species by targeting SOD3 and TNF. Cancer Research [Internet]. 2012 Sep 15;72(18):4707-4713. Available from: http://cancerres.aacrjournals.

[158] Higgs G, Slack F. The multiple roles of microRNA-155 in oncogenesis. Journal of Clinical Bioinformatics [Internet]. 2013;3(1):17. Available from: http://jclinbioinformatics.biomed

[159] Mattiske S, Suetani RJ, Neilsen PM, Callen DF. The oncogenic role of miR-155 in breast cancer. Cancer Epidemiology, Biomarkers & Prevention [Internet]. 2012 Aug 1;21(8):1236- 1243. Available from: http://cebp.aacrjournals.org/cgi/doi/10.1158/1055-9965.EPI-12-0173

[160] Feng C-Z, Yin J-B, Yang J-J, Cao L. Regulatory factor X1 depresses ApoE-dependent Aβ uptake by miRNA-124 in microglial response to oxidative stress. Neuroscience [Internet]. 2017 Mar;344:217-228. Available from: http://linkinghub.elsevier.com/retrieve/pii/

[161] He J, Xu Q, Jing Y, Agani F, Qian X, Carpenter R, et al. Reactive oxygen species regulate ERBB2 and ERBB3 expression via miR-199a/125b and DNA methylation. EMBO Reports [Internet]. 2012 Nov 13;13(12):1116-1122. Available from: http://embor.embopress.org/

[162] Rushmore TH, Morton MR, Pickett CB. The antioxidant responsive element: Activation by oxidative stress and identification of the DNA consensus sequence required for functional activity. The Journal of Biological Chemistry. 1991;266(18):11632-11639 [163] Jaiswal AK. Nrf2 signaling in coordinated activation of antioxidant gene expression. Free Radical Biology & Medicine [Internet]. 2004 May;36(10):1199-1207. Available from:

[164] Campagne MV, Thibodeaux H, van Bruggen N, Cairns B, Lowe DG. Increased binding activity at an antioxidant-responsive element in the metallothionein-1 promoter and

http://linkinghub.elsevier.com/retrieve/pii/S0891584904001923

com/retrieve/pii/S1567576908001562

30 Novel Prospects in Oxidative and Nitrosative Stress

org/cgi/doi/10.1158/0008-5472.CAN-12-0639

central.com/articles/10.1186/2043-9113-3-17

10.1038/nrc2981

S0306452216307072

cgi/doi/10.1038/embor.2012.162

[166] Hara H. Increase of antioxidative potential by tert-butylhydroquinone protects against cell death associated with 6-hydroxydopamine-induced oxidative stress in neuroblastoma SH-SY5Y cells. Molecular Brain Research [Internet]. 2003 Nov 26;119(2):125-131. Available from: http://linkinghub.elsevier.com/retrieve/pii/S0169328X03003462

**Chapter 2**

**Provisional chapter**

**The Role of Oxidative Stress and Systemic**

**The Role of Oxidative Stress and Systemic** 

**Cardiovascular Risk**

**Cardiovascular Risk**

Aye San, Magid Fahim, Katrina Campbell, Carmel M. Hawley and David W. Johnson

Aye San, Magid Fahim, Katrina Campbell, Carmel M. Hawley and David W. Johnson

Additional information is available at the end of the chapter

Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.73239

**Abstract**

potential therapies.

**Inflammation in Kidney Disease and Its Associated**

**Inflammation in Kidney Disease and Its Associated** 

Chronic kidney disease (CKD) is a major global health burden, with a prevalence of 10–15% and high mortality rates. In particular, CKD portends a disproportionately high risk of cardiovascular disease beyond the traditional cardiovascular risk factors, with pathophysiological factors such as oxidative stress, inflammation and hyperuricaemia considered to exert an additional role in accelerated atherosclerosis. The presence of heightened oxidative stress and systemic inflammation in CKD is associated with increased mortality. The possible underlying mechanisms include gut dysbiosis, dialysis factors, infections, metabolic acidosis and hyperuricaemia. The state of oxidative stress and systemic inflammation are closely linked and perpetuate each other resulting in progression of CKD and cardiovascular disease. Potential interventions to alleviate the oxidative stress and inflammation in CKD include lifestyle modifications including dietary changes and exercise, optimization of dialysis procedure and pharmacotherapeutic agents including antioxidants. They present a potentially highly effective approach to add to the currently available traditional risk-modification strategies. To date, the majority of the published trials have had a small number of participants with a short duration of follow up. Therefore, no robust evidence has been established. Larger trials with meaningful clinical outcomes and longer follow up are required to evaluate such

**Keywords:** cardiovascular disease, chronic kidney disease, endotoxin, nitric oxide,

oxidative stress, reactive oxygen species, systemic inflammation

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use,

distribution, and reproduction in any medium, provided the original work is properly cited.

DOI: 10.5772/intechopen.73239

#### **The Role of Oxidative Stress and Systemic Inflammation in Kidney Disease and Its Associated Cardiovascular Risk The Role of Oxidative Stress and Systemic Inflammation in Kidney Disease and Its Associated Cardiovascular Risk**

DOI: 10.5772/intechopen.73239

Aye San, Magid Fahim, Katrina Campbell, Carmel M. Hawley and David W. Johnson Aye San, Magid Fahim, Katrina Campbell, Carmel M. Hawley and David W. Johnson

Additional information is available at the end of the chapter Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.73239

#### **Abstract**

Chronic kidney disease (CKD) is a major global health burden, with a prevalence of 10–15% and high mortality rates. In particular, CKD portends a disproportionately high risk of cardiovascular disease beyond the traditional cardiovascular risk factors, with pathophysiological factors such as oxidative stress, inflammation and hyperuricaemia considered to exert an additional role in accelerated atherosclerosis. The presence of heightened oxidative stress and systemic inflammation in CKD is associated with increased mortality. The possible underlying mechanisms include gut dysbiosis, dialysis factors, infections, metabolic acidosis and hyperuricaemia. The state of oxidative stress and systemic inflammation are closely linked and perpetuate each other resulting in progression of CKD and cardiovascular disease. Potential interventions to alleviate the oxidative stress and inflammation in CKD include lifestyle modifications including dietary changes and exercise, optimization of dialysis procedure and pharmacotherapeutic agents including antioxidants. They present a potentially highly effective approach to add to the currently available traditional risk-modification strategies. To date, the majority of the published trials have had a small number of participants with a short duration of follow up. Therefore, no robust evidence has been established. Larger trials with meaningful clinical outcomes and longer follow up are required to evaluate such potential therapies.

**Keywords:** cardiovascular disease, chronic kidney disease, endotoxin, nitric oxide, oxidative stress, reactive oxygen species, systemic inflammation

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

### **1. Introduction**

Chronic kidney disease (CKD), defined as an estimated or measured glomerular filtration rate (GFR) <60 mL/min/1.73 m2 and/or evidence of kidney damage (usually manifested as proteinuria/albuminuria) for at least 3 months [1], is a major and growing public health burden [2]. It is currently estimated that approximately 10–15% of the world's population suffer from CKD and accounts for 4% of the deaths worldwide [3] and has progressively risen from the 21st to the 17th commonest cause of global years of life lost between 2005 and 2015 [4].

cardiovascular end-points by approximately 20%. There is therefore a pressing need to develop novel, and more effective therapeutic strategies. Therapies targeting oxidative stress and inflammation are promising and may be adjuncts to current therapies targeting traditional risk factors. This chapter reviews the pathophysiologic mechanisms underlying the heightened oxidative stress and systemic inflammation in CKD, and their association with mortality. Current evi-

The Role of Oxidative Stress and Systemic Inflammation in Kidney Disease and Its Associated…

http://dx.doi.org/10.5772/intechopen.73239

35

Oxidative stress is a state of excessive pathological pro-oxidant activities relative to antioxidant defense mechanisms. The majority of oxidizing agents are reactive oxygen species (ROS),

In CKD, there is accumulating evidence that excess oxidative activity exists with deficient antioxidant protection. Studies in dialysis populations show higher levels of end products of oxidation, such as protein carbonyl, oxidized lipoproteins, F2-isoproteins, advanced oxidation protein products (AOPPs), thiobarbituric acid reactive substances (TBARS), 8-hydroxyl-2′-deoxyguanosine (8OHdG). The free radical superoxide anion production by the pro-oxidant enzyme NADPH oxidase (NOX) is found to be elevated in haemodialysis patients [7]. At the same time, it has been shown that dialysis patients have reduced antioxidant activities of superoxide dismutase (SOD), glutathione, and reduced levels of antioxidants such as vitamin A, C, E, zinc (Zn) and selenium (Se) [7, 8]. In a cross-sectional study of 159 patients (28 to 36 patients in each of the CKD stages 1 to 5) compared with 30 healthy controls, Yilmaz et al. [9] also found that a progressive increase in the levels of oxidative stress marker, malondialdehyde (MDA), while concentrations of antioxidant elements, including SOD, glutathione peroxidase (GSH-Px), Zn, copper (Cu) and Se, fell with increasing levels of kidney dysfunction. The most profound redox imbalances were seen in haemodialysis patients. Therefore, the level of oxidative stress in CKD patients appears to

These perturbations in oxidant-antioxidant balance begin early in the course of CKD. Fortuño et al. [13] showed increased levels of NADPH-generated ROS in patients with early stage (stages 1 and 2) CKD. Similarly, Yilmaz et al. [9] reported lower levels of activity of the antioxidant enzymes, SOD and GSH-Px, in patients with CKD, including those in stages 1–2.

In kidney transplant recipients, there was an increase in oxidative stress, as evidenced by a rise in MDA immediately after allograft reperfusion [14]. Over the following 2 weeks after transplant, there was a continued rise in MDA, although this was somewhat counterbalanced by a concomitant rise in antioxidant levels, such as GSH-Px. Other oxidative and inflammatory markers, such as IL-6, CRP, tumour necrosis factor-α (TNF-α) and protein carbonyls, significantly declined by 2 months after transplantation [15]. The level of improvement in oxidative stress depended on the level of graft function, such that complete resolution was only possible if renal function returned to normal. At 1 year after transplant, the patients with higher serum creatinine concentrations also displayed higher levels of oxidative stress and

dence on various antioxidant and anti-inflammatory therapies are also reviewed.

but others include reactive nitrogen species (RNS), chlorine and carbonyl species.

**2. Oxidative stress in patients with CKD**

escalate with declining renal function.

The presence of CKD carries a disproportionately increased risk of cardiovascular disease, which increases with progressive declines in GFR and/or increases in albuminuria/proteinuria [1]. Indeed, CKD is considered the most potent known risk factor for cardiovascular disease [5]. Part of this risk is explained by the frequent clustering of traditional cardiovascular risk factors, such as diabetes mellitus, hypertension, obesity, smoking, dyslipidaemia and depression in patients with CKD. However, traditional cardiovascular risk factors account for less than half of the observed excess cardiovascular risk [6]. Consequently, investigators have evaluated the roles of a range of non-traditional cardiovascular risk factors in patients with CKD, including oxidative stress, inflammation and hyperuricaemia (**Table 1**).

Oxidative stress is a particularly promising avenue of investigation. It is becoming well established that CKD is a state of elevated oxidative stress. This is evidenced by the presence of elevated reactive oxygen and nitrogen species, oxidized end products and reduced levels of antioxidants in patients with CKD [7–10]. Systemic inflammation is also up-regulated in CKD evidenced by higher concentrations of inflammatory markers including C-reactive protein (CRP) and interleukin-6 (IL-6) which have been associated with higher mortality in CKD patients and patients with end stage kidney disease (ESKD) [11, 12]. Oxidative stress and chronically elevated inflammatory state may contribute to accelerated atherosclerosis via direct endothelial injury and alteration in nitrogen handling. Reactive oxygen species (ROS) can also cause direct glomerular and tubulointerstitial injury in the kidneys, resulting in further progression of CKD.

The therapeutic options currently available and shown to be beneficial for CKD are limited to anti-hypertensive agents, particularly renin-angiotensin-aldosterone system (RAAS) blockers. However, these agents are only partially effective, typically lowering the risk of renal and


**Table 1.** The traditional and non-traditional risk factors for development of cardiovascular disease.

cardiovascular end-points by approximately 20%. There is therefore a pressing need to develop novel, and more effective therapeutic strategies. Therapies targeting oxidative stress and inflammation are promising and may be adjuncts to current therapies targeting traditional risk factors.

This chapter reviews the pathophysiologic mechanisms underlying the heightened oxidative stress and systemic inflammation in CKD, and their association with mortality. Current evidence on various antioxidant and anti-inflammatory therapies are also reviewed.

### **2. Oxidative stress in patients with CKD**

**1. Introduction**

Age

Hypertension Hyperlipidaemia Tobacco use Diabetes Obesity

(GFR) <60 mL/min/1.73 m2

34 Novel Prospects in Oxidative and Nitrosative Stress

Chronic kidney disease (CKD), defined as an estimated or measured glomerular filtration rate

uria/albuminuria) for at least 3 months [1], is a major and growing public health burden [2]. It is currently estimated that approximately 10–15% of the world's population suffer from CKD and accounts for 4% of the deaths worldwide [3] and has progressively risen from the 21st to

The presence of CKD carries a disproportionately increased risk of cardiovascular disease, which increases with progressive declines in GFR and/or increases in albuminuria/proteinuria [1]. Indeed, CKD is considered the most potent known risk factor for cardiovascular disease [5]. Part of this risk is explained by the frequent clustering of traditional cardiovascular risk factors, such as diabetes mellitus, hypertension, obesity, smoking, dyslipidaemia and depression in patients with CKD. However, traditional cardiovascular risk factors account for less than half of the observed excess cardiovascular risk [6]. Consequently, investigators have evaluated the roles of a range of non-traditional cardiovascular risk factors in patients with

Oxidative stress is a particularly promising avenue of investigation. It is becoming well established that CKD is a state of elevated oxidative stress. This is evidenced by the presence of elevated reactive oxygen and nitrogen species, oxidized end products and reduced levels of antioxidants in patients with CKD [7–10]. Systemic inflammation is also up-regulated in CKD evidenced by higher concentrations of inflammatory markers including C-reactive protein (CRP) and interleukin-6 (IL-6) which have been associated with higher mortality in CKD patients and patients with end stage kidney disease (ESKD) [11, 12]. Oxidative stress and chronically elevated inflammatory state may contribute to accelerated atherosclerosis via direct endothelial injury and alteration in nitrogen handling. Reactive oxygen species (ROS) can also cause direct glomerular and tubulointerstitial injury in the kidneys, resulting in further progression of CKD. The therapeutic options currently available and shown to be beneficial for CKD are limited to anti-hypertensive agents, particularly renin-angiotensin-aldosterone system (RAAS) blockers. However, these agents are only partially effective, typically lowering the risk of renal and

Oxidative stress

Hyperuricaemia Thrombosis

**Table 1.** The traditional and non-traditional risk factors for development of cardiovascular disease.

Adipocyte dysfunction

Mitochondrial dysfunction Systemic inflammation Hyperhomocysteinaemia

the 17th commonest cause of global years of life lost between 2005 and 2015 [4].

CKD, including oxidative stress, inflammation and hyperuricaemia (**Table 1**).

**Traditional risk factors Non-traditional risk factors**

and/or evidence of kidney damage (usually manifested as protein-

Oxidative stress is a state of excessive pathological pro-oxidant activities relative to antioxidant defense mechanisms. The majority of oxidizing agents are reactive oxygen species (ROS), but others include reactive nitrogen species (RNS), chlorine and carbonyl species.

In CKD, there is accumulating evidence that excess oxidative activity exists with deficient antioxidant protection. Studies in dialysis populations show higher levels of end products of oxidation, such as protein carbonyl, oxidized lipoproteins, F2-isoproteins, advanced oxidation protein products (AOPPs), thiobarbituric acid reactive substances (TBARS), 8-hydroxyl-2′-deoxyguanosine (8OHdG). The free radical superoxide anion production by the pro-oxidant enzyme NADPH oxidase (NOX) is found to be elevated in haemodialysis patients [7]. At the same time, it has been shown that dialysis patients have reduced antioxidant activities of superoxide dismutase (SOD), glutathione, and reduced levels of antioxidants such as vitamin A, C, E, zinc (Zn) and selenium (Se) [7, 8]. In a cross-sectional study of 159 patients (28 to 36 patients in each of the CKD stages 1 to 5) compared with 30 healthy controls, Yilmaz et al. [9] also found that a progressive increase in the levels of oxidative stress marker, malondialdehyde (MDA), while concentrations of antioxidant elements, including SOD, glutathione peroxidase (GSH-Px), Zn, copper (Cu) and Se, fell with increasing levels of kidney dysfunction. The most profound redox imbalances were seen in haemodialysis patients. Therefore, the level of oxidative stress in CKD patients appears to escalate with declining renal function.

These perturbations in oxidant-antioxidant balance begin early in the course of CKD. Fortuño et al. [13] showed increased levels of NADPH-generated ROS in patients with early stage (stages 1 and 2) CKD. Similarly, Yilmaz et al. [9] reported lower levels of activity of the antioxidant enzymes, SOD and GSH-Px, in patients with CKD, including those in stages 1–2.

In kidney transplant recipients, there was an increase in oxidative stress, as evidenced by a rise in MDA immediately after allograft reperfusion [14]. Over the following 2 weeks after transplant, there was a continued rise in MDA, although this was somewhat counterbalanced by a concomitant rise in antioxidant levels, such as GSH-Px. Other oxidative and inflammatory markers, such as IL-6, CRP, tumour necrosis factor-α (TNF-α) and protein carbonyls, significantly declined by 2 months after transplantation [15]. The level of improvement in oxidative stress depended on the level of graft function, such that complete resolution was only possible if renal function returned to normal. At 1 year after transplant, the patients with higher serum creatinine concentrations also displayed higher levels of oxidative stress and inflammation, such as IL-6, MDA, and transforming growth factor beta (TGF-β), compared to those with normal serum creatinine levels [15].

(NF-kB). NF-kB is the master initiator of proinflammatory processes and induces the production of cytokines IL-1, IL-6 and TNF-α [21]. The influx of proinflammatory cytokines causes

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Hyperglycaemia in diabetic kidney disease has direct and indirect roles in increasing oxidative stress. Advanced glycation end products (AGEs) are formed when the carbonyl component is added non-enzymatically to the free amino acid group of proteins or lipids in the presence of chronic hyperglycaemia. The effects of AGEs are executed via various receptors for AGE (RAGE), which in turn activate transcription factors, such as NF-ΚB, AP1 and SP1, to activate various oxidative pathways. The pathways activated by AGEs cause production of reactive oxygen species with the end results of mesangial expansion, glomerular basement membrane (GBM) thickening and endothelial cell dysfunction. First, AGEs activate the protein kinase C (PKC) pathway in the glomeruli producing ROS and downstream pathologic changes. The second enzymatic pathway is the activation of NOX which directly generates free radicals. In addition to the enzymatic pathways, hyperglycaemia can directly stimulate the formation of TGF-β, again resulting in mesangial expansion, renal hypertrophy and glomerulosclerosis. The presence of increased renin-angiotensin-aldosterone system (RAAS) activity increases the level of angiotensin II which is a potent initiator of inflammatory pro-

**Figure 1.** The role of microbiota in the progression of chronic kidney disease and accelerated atherosclerosis. IS, Indoxyl sulphate; PCS, p-cresyl sulphate; TMAO, trimethylamine N-oxide; TLR, toll-like receptor, NF-kB, nuclear factor kB; ROS, reactive oxygen species; NO, nitric oxide; ADMA, asymmetric dimethylarginine; CVS, cardiovascular system.

generation of reactive species (**Figure 1**).

cesses and subsequent oxidative stress [10, 22].

*2.1.2. Hyperglycaemia*

#### **2.1. Mechanisms of increased oxidative stress**

A number of factors have been identified which may contribute to the state of heightened oxidative stress in people with CKD.

### *2.1.1. Gut dysbiosis*

A symbiotic relationship exists between the host and its bacterial flora (also known as microbiota), which are predominantly found in the gastrointestinal tract. In addition to regulating the nutrient absorption and protecting the host from pathological bacteria, the gut microbiota has been increasingly linked to progression of CKD through several mechanisms, including generation of uraemic toxins, enhanced intestinal permeability to endotoxins and alteration of nitrogen handling, all of which contribute to elevated oxidative state.

The breakdown of tyrosine and phenylalanine by the intestinal bacteria produces p-cresyl sulphate (PCS), and the breakdown of tryptophan produces the end product of indoxyl sulphate (IS) in the liver. These two nitrogenous end products have been extensively studied for their role in CKD. They are highly protein-bound in plasma, but as their levels become elevated in patients with CKD, the toxic free fraction level also rises [16]. The serum levels of these toxins have been found to correlate with the extent of damage observed in renal glomerular and tubular cells, tubulointerstitial damage and increased production of reactive oxygen species [16, 17].

Factors influencing the amount of toxins produced include the balance between carbohydrate (in the form of dietary fibre) and amino acids in the large intestine, the intestinal transit time, and the permeability of the gastrointestinal tract. Production of uremic toxins is influenced by substrate availability for fermentation in the colon, notably the balance between carbohydrate (fermentable fibre), and amino acids. A low concentration of fermentable fibre alters the composition of intestinal bacteria to favour more proteolytic bacterial species resulting in the higher production of nitrogenous waste and uraemic toxins. The slower intestinal transit time predisposes to bacteria overgrowth with subsequent production of pro-inflammatory toxins [18].

In addition to the increased production of uraemic toxins in CKD patients, there is evidence that changes in the composition of intestinal bacteria compromise the integrity of intestinal barrier resulting in increased intestinal permeability. This is supported by findings of depletion of the tight junction proteins in the gastrointestinal tract of uraemic patients and drops in transepithelial electrical resistance in colonocytes exposed to the plasma of uraemic patients [19]. Suppression of anti-inflammatory nuclear factor erythroid 2-related factor 2 (NRF2) has also been noted. This may contribute to gut inflammation and reduced expression of epithelial tight junctions, resulting in increased gut permeability [20]. Translocation of bacterial fragments and endotoxins due to increased intestinal permeability may activate the pro-inflammatory pathways in the systemic circulation. For example, increased plasma concentration of lipopolysaccharides due to increased intestinal permeability activates the tolllike receptors (TLR) 2 and TLR4, which in turn activates the nuclear transcription factor kB (NF-kB). NF-kB is the master initiator of proinflammatory processes and induces the production of cytokines IL-1, IL-6 and TNF-α [21]. The influx of proinflammatory cytokines causes generation of reactive species (**Figure 1**).

#### *2.1.2. Hyperglycaemia*

inflammation, such as IL-6, MDA, and transforming growth factor beta (TGF-β), compared to

A number of factors have been identified which may contribute to the state of heightened

A symbiotic relationship exists between the host and its bacterial flora (also known as microbiota), which are predominantly found in the gastrointestinal tract. In addition to regulating the nutrient absorption and protecting the host from pathological bacteria, the gut microbiota has been increasingly linked to progression of CKD through several mechanisms, including generation of uraemic toxins, enhanced intestinal permeability to endotoxins and alteration

The breakdown of tyrosine and phenylalanine by the intestinal bacteria produces p-cresyl sulphate (PCS), and the breakdown of tryptophan produces the end product of indoxyl sulphate (IS) in the liver. These two nitrogenous end products have been extensively studied for their role in CKD. They are highly protein-bound in plasma, but as their levels become elevated in patients with CKD, the toxic free fraction level also rises [16]. The serum levels of these toxins have been found to correlate with the extent of damage observed in renal glomerular and tubular cells, tubulointerstitial damage and increased production of reactive oxygen species [16, 17]. Factors influencing the amount of toxins produced include the balance between carbohydrate (in the form of dietary fibre) and amino acids in the large intestine, the intestinal transit time, and the permeability of the gastrointestinal tract. Production of uremic toxins is influenced by substrate availability for fermentation in the colon, notably the balance between carbohydrate (fermentable fibre), and amino acids. A low concentration of fermentable fibre alters the composition of intestinal bacteria to favour more proteolytic bacterial species resulting in the higher production of nitrogenous waste and uraemic toxins. The slower intestinal transit time predisposes to bacteria overgrowth with subsequent production of pro-inflammatory toxins [18]. In addition to the increased production of uraemic toxins in CKD patients, there is evidence that changes in the composition of intestinal bacteria compromise the integrity of intestinal barrier resulting in increased intestinal permeability. This is supported by findings of depletion of the tight junction proteins in the gastrointestinal tract of uraemic patients and drops in transepithelial electrical resistance in colonocytes exposed to the plasma of uraemic patients [19]. Suppression of anti-inflammatory nuclear factor erythroid 2-related factor 2 (NRF2) has also been noted. This may contribute to gut inflammation and reduced expression of epithelial tight junctions, resulting in increased gut permeability [20]. Translocation of bacterial fragments and endotoxins due to increased intestinal permeability may activate the pro-inflammatory pathways in the systemic circulation. For example, increased plasma concentration of lipopolysaccharides due to increased intestinal permeability activates the tolllike receptors (TLR) 2 and TLR4, which in turn activates the nuclear transcription factor kB

of nitrogen handling, all of which contribute to elevated oxidative state.

those with normal serum creatinine levels [15].

**2.1. Mechanisms of increased oxidative stress**

oxidative stress in people with CKD.

36 Novel Prospects in Oxidative and Nitrosative Stress

*2.1.1. Gut dysbiosis*

Hyperglycaemia in diabetic kidney disease has direct and indirect roles in increasing oxidative stress. Advanced glycation end products (AGEs) are formed when the carbonyl component is added non-enzymatically to the free amino acid group of proteins or lipids in the presence of chronic hyperglycaemia. The effects of AGEs are executed via various receptors for AGE (RAGE), which in turn activate transcription factors, such as NF-ΚB, AP1 and SP1, to activate various oxidative pathways. The pathways activated by AGEs cause production of reactive oxygen species with the end results of mesangial expansion, glomerular basement membrane (GBM) thickening and endothelial cell dysfunction. First, AGEs activate the protein kinase C (PKC) pathway in the glomeruli producing ROS and downstream pathologic changes. The second enzymatic pathway is the activation of NOX which directly generates free radicals. In addition to the enzymatic pathways, hyperglycaemia can directly stimulate the formation of TGF-β, again resulting in mesangial expansion, renal hypertrophy and glomerulosclerosis. The presence of increased renin-angiotensin-aldosterone system (RAAS) activity increases the level of angiotensin II which is a potent initiator of inflammatory processes and subsequent oxidative stress [10, 22].

**Figure 1.** The role of microbiota in the progression of chronic kidney disease and accelerated atherosclerosis. IS, Indoxyl sulphate; PCS, p-cresyl sulphate; TMAO, trimethylamine N-oxide; TLR, toll-like receptor, NF-kB, nuclear factor kB; ROS, reactive oxygen species; NO, nitric oxide; ADMA, asymmetric dimethylarginine; CVS, cardiovascular system.

#### *2.1.3. Dialysis*

The dialysis procedure itself accentuates the heightened state of oxidative stress observed in CKD patients. In haemodialysis patients, the mechanisms of oxidative stress include the use of bioincompatible membranes, contamination of dialysate with bacterial endotoxins, occult infection of clotted vascular access and potential loss of antioxidants during the dialysis procedure. Wu et al. [23] demonstrated that levels of myeloperoxidase (MPO), AOPP and 8-OHdG were significantly higher in patients dialysed with regenerated cellulose membranes compared to those dialysed with synthetic polysulphone membranes. However, even with the use of biocompatible membranes, the haemodialysis procedure can still increase systemic levels of reactive oxygen species by 14-fold during one session [24].

It is known that the endotoxins in the dialysate affect the level of cytokines produced by the peripheral leucocytes. Studies have shown that lower levels of endotoxin contamination correlate with lower levels of inflammatory cytokines and oxidative stress markers. In a meta-analysis of 31 studies involving 1580 dialysis patients, the use of ultrapure dialysate has been found to reduce the oxidative stress markers, pentosidine, MPO and oxidized LDL cholesterol [25].

The loss of antioxidants, especially water-soluble vitamins such as vitamin C, has been demonstrated during the dialysis procedure [24, 26].

#### *2.1.4. Inflammation*

CKD has also been noted to be a systemic inflammatory state, which is intertwined with oxidative stress. Inflammatory cells stimulate the release of reactive species at the site of inflammation. Conversely, oxidized end products and ROS stimulate phagocytic cells, such as macrophages and neutrophils, to release inflammatory cytokines as well as more ROS, thereby creating a positive feedback loop of inflammation and oxidative stress state. When the phagocytic cells release ROS, they also induce nearby non-phagocytic cells to release inflammatory cytokines. Studies of oxidative states in people with CKD commonly include the investigation of inflammatory cytokines as the two pathways are intimately inter-related.

Therefore an event inciting increased oxidative stress can result in an inflammatory response

**Figure 2.** Interrelation between ROS (reactive oxygen species) and oxidized end products signifying the oxidative stress, and cytokines as part of the inflammatory process. ROS, reactive oxygen species; MPO, myeloperoxidase; NOX, NADPH-oxidases; NF-kB, nuclear factor kB; NLRP3, nod-like receptor protein 3; TLR4, toll-like receptor 4; JNK, c-Jun

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A number of mechanisms have been reported to underpin the association between increased

Increased oxidized low density lipoprotein (LDL) levels have been recognized as a risk factor for cardiovascular disease in uraemic patients [27, 28]. Oxidized LDLs stimulate the release of inflammatory cytokines from macrophages with subsequent vascular phenotypic changes and platelet aggregation [29], which result in endothelial dysfunction. Oxidized LDL also stimulates further oxidative activity by enhancing the activity of NOX in endothelial cells, juxtaglomerular cells and mesangial cells. Increased levels of ox-LDL in CKD patients correlate

Similarly, in patients with CKD, increased carotid intima-media thickness, a marker of atherosclerosis, has been strongly positively correlated with increased levels and activities of NOX, MDA, AOPP, TBARS, 8-OHdG and MPO [24, 31–34] and negatively correlated with

**2.2. Association of oxidative stress with CKD progression and cardiovascular** 

oxidative stress and progression of CKD and CVD in patients with CKD.

which further propagates the oxidative state, and vice versa.

*2.2.1. Oxidative stress markers and vascular disease*

SOD and GSH-Px levels [34].

positively with carotid atherosclerosis and mortality [30].

**disease (CVD)**

N-terminal; IL, interleukins.

Multiple pathways have been identified that highlight the mechanisms of interplay between inflammation and oxidative stress (**Figure 2**). The master regulator of the inflammatory process is NF-kB transcription factor. Reactive oxygen species, such as H2 O2, activate NF-kB which induces production of an array of inflammatory cytokines as well as activates NOX. These in turn stimulate further release of reactive species. Free radical-induced DNA base modifications can also act via NF-kB to activate the inflammatory processes.

NOX are responsible for free radical production by cells. They can be activated by inflammatory mediators, such as TGF, angiotensin II and TNF-α via other redox sensitive signal transduction pathways, such as c-Jun N-terminal kinase (JNK).

Nucleotide-binding oligomerization domain, Leucine rich Repeat and Pyrin domain 3 (NLRP3) is an inflammasome complex involved in the production of cytokines, such as IL-1β and IL-18. Oxidative stress can activate NLRP3 via oxidized mitochondrial DNA and thioredoxin-interacting proteins. Damaged mitochondria can directly release ROS, which perpetuate oxidative stress and inflammation.

The Role of Oxidative Stress and Systemic Inflammation in Kidney Disease and Its Associated… http://dx.doi.org/10.5772/intechopen.73239 39

**Figure 2.** Interrelation between ROS (reactive oxygen species) and oxidized end products signifying the oxidative stress, and cytokines as part of the inflammatory process. ROS, reactive oxygen species; MPO, myeloperoxidase; NOX, NADPH-oxidases; NF-kB, nuclear factor kB; NLRP3, nod-like receptor protein 3; TLR4, toll-like receptor 4; JNK, c-Jun N-terminal; IL, interleukins.

Therefore an event inciting increased oxidative stress can result in an inflammatory response which further propagates the oxidative state, and vice versa.

#### **2.2. Association of oxidative stress with CKD progression and cardiovascular disease (CVD)**

A number of mechanisms have been reported to underpin the association between increased oxidative stress and progression of CKD and CVD in patients with CKD.

#### *2.2.1. Oxidative stress markers and vascular disease*

*2.1.3. Dialysis*

38 Novel Prospects in Oxidative and Nitrosative Stress

*2.1.4. Inflammation*

The dialysis procedure itself accentuates the heightened state of oxidative stress observed in CKD patients. In haemodialysis patients, the mechanisms of oxidative stress include the use of bioincompatible membranes, contamination of dialysate with bacterial endotoxins, occult infection of clotted vascular access and potential loss of antioxidants during the dialysis procedure. Wu et al. [23] demonstrated that levels of myeloperoxidase (MPO), AOPP and 8-OHdG were significantly higher in patients dialysed with regenerated cellulose membranes compared to those dialysed with synthetic polysulphone membranes. However, even with the use of biocompatible membranes, the haemodialysis procedure can still increase systemic

It is known that the endotoxins in the dialysate affect the level of cytokines produced by the peripheral leucocytes. Studies have shown that lower levels of endotoxin contamination correlate with lower levels of inflammatory cytokines and oxidative stress markers. In a meta-analysis of 31 studies involving 1580 dialysis patients, the use of ultrapure dialysate has been found to reduce the oxidative stress markers, pentosidine, MPO and oxidized LDL cholesterol [25]. The loss of antioxidants, especially water-soluble vitamins such as vitamin C, has been dem-

CKD has also been noted to be a systemic inflammatory state, which is intertwined with oxidative stress. Inflammatory cells stimulate the release of reactive species at the site of inflammation. Conversely, oxidized end products and ROS stimulate phagocytic cells, such as macrophages and neutrophils, to release inflammatory cytokines as well as more ROS, thereby creating a positive feedback loop of inflammation and oxidative stress state. When the phagocytic cells release ROS, they also induce nearby non-phagocytic cells to release inflammatory cytokines. Studies of oxidative states in people with CKD commonly include the investigation of inflammatory cytokines as the two pathways are intimately inter-related. Multiple pathways have been identified that highlight the mechanisms of interplay between inflammation and oxidative stress (**Figure 2**). The master regulator of the inflammatory pro-

induces production of an array of inflammatory cytokines as well as activates NOX. These in turn stimulate further release of reactive species. Free radical-induced DNA base modifica-

NOX are responsible for free radical production by cells. They can be activated by inflammatory mediators, such as TGF, angiotensin II and TNF-α via other redox sensitive signal

Nucleotide-binding oligomerization domain, Leucine rich Repeat and Pyrin domain 3 (NLRP3) is an inflammasome complex involved in the production of cytokines, such as IL-1β and IL-18. Oxidative stress can activate NLRP3 via oxidized mitochondrial DNA and thioredoxin-interacting proteins. Damaged mitochondria can directly release ROS, which perpetu-

O2, activate NF-kB which

levels of reactive oxygen species by 14-fold during one session [24].

cess is NF-kB transcription factor. Reactive oxygen species, such as H2

tions can also act via NF-kB to activate the inflammatory processes.

transduction pathways, such as c-Jun N-terminal kinase (JNK).

ate oxidative stress and inflammation.

onstrated during the dialysis procedure [24, 26].

Increased oxidized low density lipoprotein (LDL) levels have been recognized as a risk factor for cardiovascular disease in uraemic patients [27, 28]. Oxidized LDLs stimulate the release of inflammatory cytokines from macrophages with subsequent vascular phenotypic changes and platelet aggregation [29], which result in endothelial dysfunction. Oxidized LDL also stimulates further oxidative activity by enhancing the activity of NOX in endothelial cells, juxtaglomerular cells and mesangial cells. Increased levels of ox-LDL in CKD patients correlate positively with carotid atherosclerosis and mortality [30].

Similarly, in patients with CKD, increased carotid intima-media thickness, a marker of atherosclerosis, has been strongly positively correlated with increased levels and activities of NOX, MDA, AOPP, TBARS, 8-OHdG and MPO [24, 31–34] and negatively correlated with SOD and GSH-Px levels [34].

Reactive oxygen species may also promote direct injury of vascular endothelial cells by inducing significant increases in nucleic acid damage in CKD patients [35].

reducing availability of nitric oxide to the vascular smooth muscle. Peroxynitrite, due to its

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Systemic inflammation, most commonly evidenced by an elevation of CRP, is frequently observed in people with CKD, even early stage disease. In CKD, this systemic inflammation has been ascribed to a complex biological reaction in response to exposure to inciting exogenous stimuli such as bacterial pathogens or endogenous stimuli such as injured cells. It is mainly characterized by involvement of the cells of innate immune system and release of

The most studied inflammatory marker in CKD patients is CRP. Elevated levels of CRP and the presence of a chronic inflammatory process in haemodialysis patients was first observed in 1980s with evidence accumulating over time of the heightened inflammatory state in CKD. CRP is produced by the liver as part of anti-inflammatory response and the rate of metabolism is nearly constant. Earlier studies involved mainly in haemodialysis patients where the elevation was noted consistently. The interest then expanded to non-dialysis ESKD patients and peritoneal dialysis (PD) patients. It is found that CRP is elevated even in ESRD patients receiving conservative care and also in PD patients [41]. This inflammatory state has

More recently, other biomarkers of inflammation have been studied. In the large Chronic Renal Insufficiency Cohort (CRIC) study [42], inflammatory markers (IL-1β, IL-1receptor antagonist, TNF-α, and fibrinogen and in addition to CRP) were found to be negatively correlated with kidney function and positively correlated with albuminuria (a marker of kidney damage). Another study showed an inverse correlation between IL-6 and kidney function [12]. The pattern of elevated pro-inflammatory cytokines (IL-1, IL-6, TNF-α) with low anti-inflammatory cytokines (IL2, IL4, IL5, IL12, CH50 and T cell number) has also been described in haemodialysis patients [43]. CRP, IL-6 and IL-10 levels are significantly higher in ESKD patients compared to controls and the levels did not change significantly after initiation of maintenance haemodialysis [44]. In another study, it was noted that CRP levels actually increased after initiation

Other markers of inflammatory status involve adipokines. There are proinflammatory adipokines and anti-inflammatory adipokines secreted by adipose tissue. Finally, the National Health and Nutrition Examination Survey (NHANES) demonstrated that levels of pro-inflammatory adipokines were higher in CKD patients than in the general population and that and the ratio of pro- to anti- inflammatory adipokines predicted mortality in PD patients [45].

In addition to promoting oxidative stress, gut dysbiosis may also promote systemic inflammation in patients with CKD. Endotoxemia activates TLR4 on endothelial cells and macrophages

oxidative and nitrosative properties, perpetuates the vascular injury [40].

**3. Systemic inflammation in patients with CKD**

in turn been linked with progression of CKD and CVD.

**3.1. Mechanisms of increased systemic inflammation**

acute phase proteins and cytokines.

of haemodialysis [41].

*3.1.1. Gut dysbiosis*

#### *2.2.2. Direct toxicity of oxidized end products*

Oxidative stress (induced by H2 O2 ) in experimental models of human proximal tubular epithelial cells is demonstrated to cause apoptosis and reduce mitosis of the renal cells. The results are highest when combined with induction of cell senescence, which itself can be induced by oxidative stress [36].

Gut-derived uraemic toxins, in addition to promoting oxidative stress, may exert direct toxicity to renal and cardiovascular tissues. In animal models, IS is implicated in the causation of vascular smooth muscle proliferation, aortic calcification, and vascular wall thickening thereby contributing to increased cardiovascular risk [16]. At the same time, it is also associated with reduced renal function, increased glomerular hypertrophy and glomerular sclerosis, signifying nephrotoxicity [16]. Similar findings were noted in human observational studies, where strong associations were observed between IS, free IS, PCS with cardiovascular disease [16, 37]. In a cross-sectional study involving 149 patients with CKD stages 3 and 4, Rossi et al. [38] investigated the potential mechanisms of toxicity of these uraemic toxins. It was found that the free forms of IS and PCS correlated positively with the level of inflammatory markers (IFN-γ, IL-6 and TNF-α) and negatively with the antioxidant GSH-Px. The total PCS concentration correlated positively with carotid-femoral pulse wave velocity as a measure of arterial stiffness. These findings suggest roles for elevated inflammatory and oxidative stress states as the causes of vascular and renal dysfunction.

Trimethylamine-N-oxide (TMAO), a metabolic end product of choline that is metabolized by the gut microbiota, has also been found to be correlated with an increased risk of cardiovascular disease [39].

#### *2.2.3. Alterations in nitrogen handling*

Nitric oxide is a free radical that plays an important role in regulation of endothelial function and regional blood flow by acting as a smooth muscle relaxant. The major pathway for nitric oxide production is by conversion of L-arginine by endothelial nitric oxide synthase (NOS) enzyme. Asymmetric dimethyl arginine (ADMA) is an inhibitor of NOS by competing with L-arginine and therefore reducing the availability of nitric oxide. It plays a significant contributory role in the development of endothelial dysfunction, and also correlates with proteinuria and progression of renal disease [40].

Yilmaz et al. [9] studied 159 patients with CKD stages 1–5 compared to 30 healthy controls. In addition to findings of reduced antioxidant activity and increased oxidative end products, it was also found that the levels of ADMA were higher in CKD patients compared to controls and were negatively correlated with renal function. The levels of oxidized LDL and ADMA were also found to be inversely related to brachial artery endothelium vasodilation. These findings raise the possibility that the elevated oxidative stress present in patients with CKD results from impaired endothelial function due to the inhibition of NOS by ADMA.

Additionally, reactive oxygen species degrade the critical co-factor of NOS causing functional impairment. Super-oxide anion also reacts with nitric oxide forming peroxynitrite thereby reducing availability of nitric oxide to the vascular smooth muscle. Peroxynitrite, due to its oxidative and nitrosative properties, perpetuates the vascular injury [40].

### **3. Systemic inflammation in patients with CKD**

Reactive oxygen species may also promote direct injury of vascular endothelial cells by induc-

thelial cells is demonstrated to cause apoptosis and reduce mitosis of the renal cells. The results are highest when combined with induction of cell senescence, which itself can be

Gut-derived uraemic toxins, in addition to promoting oxidative stress, may exert direct toxicity to renal and cardiovascular tissues. In animal models, IS is implicated in the causation of vascular smooth muscle proliferation, aortic calcification, and vascular wall thickening thereby contributing to increased cardiovascular risk [16]. At the same time, it is also associated with reduced renal function, increased glomerular hypertrophy and glomerular sclerosis, signifying nephrotoxicity [16]. Similar findings were noted in human observational studies, where strong associations were observed between IS, free IS, PCS with cardiovascular disease [16, 37]. In a cross-sectional study involving 149 patients with CKD stages 3 and 4, Rossi et al. [38] investigated the potential mechanisms of toxicity of these uraemic toxins. It was found that the free forms of IS and PCS correlated positively with the level of inflammatory markers (IFN-γ, IL-6 and TNF-α) and negatively with the antioxidant GSH-Px. The total PCS concentration correlated positively with carotid-femoral pulse wave velocity as a measure of arterial stiffness. These findings suggest roles for elevated

inflammatory and oxidative stress states as the causes of vascular and renal dysfunction.

Trimethylamine-N-oxide (TMAO), a metabolic end product of choline that is metabolized by the gut microbiota, has also been found to be correlated with an increased risk of cardiovas-

Nitric oxide is a free radical that plays an important role in regulation of endothelial function and regional blood flow by acting as a smooth muscle relaxant. The major pathway for nitric oxide production is by conversion of L-arginine by endothelial nitric oxide synthase (NOS) enzyme. Asymmetric dimethyl arginine (ADMA) is an inhibitor of NOS by competing with L-arginine and therefore reducing the availability of nitric oxide. It plays a significant contributory role in the development of endothelial dysfunction, and also correlates with proteinuria

Yilmaz et al. [9] studied 159 patients with CKD stages 1–5 compared to 30 healthy controls. In addition to findings of reduced antioxidant activity and increased oxidative end products, it was also found that the levels of ADMA were higher in CKD patients compared to controls and were negatively correlated with renal function. The levels of oxidized LDL and ADMA were also found to be inversely related to brachial artery endothelium vasodilation. These findings raise the possibility that the elevated oxidative stress present in patients with CKD

Additionally, reactive oxygen species degrade the critical co-factor of NOS causing functional impairment. Super-oxide anion also reacts with nitric oxide forming peroxynitrite thereby

results from impaired endothelial function due to the inhibition of NOS by ADMA.

) in experimental models of human proximal tubular epi-

ing significant increases in nucleic acid damage in CKD patients [35].

O2

*2.2.2. Direct toxicity of oxidized end products*

Oxidative stress (induced by H2

40 Novel Prospects in Oxidative and Nitrosative Stress

induced by oxidative stress [36].

cular disease [39].

*2.2.3. Alterations in nitrogen handling*

and progression of renal disease [40].

Systemic inflammation, most commonly evidenced by an elevation of CRP, is frequently observed in people with CKD, even early stage disease. In CKD, this systemic inflammation has been ascribed to a complex biological reaction in response to exposure to inciting exogenous stimuli such as bacterial pathogens or endogenous stimuli such as injured cells. It is mainly characterized by involvement of the cells of innate immune system and release of acute phase proteins and cytokines.

The most studied inflammatory marker in CKD patients is CRP. Elevated levels of CRP and the presence of a chronic inflammatory process in haemodialysis patients was first observed in 1980s with evidence accumulating over time of the heightened inflammatory state in CKD. CRP is produced by the liver as part of anti-inflammatory response and the rate of metabolism is nearly constant. Earlier studies involved mainly in haemodialysis patients where the elevation was noted consistently. The interest then expanded to non-dialysis ESKD patients and peritoneal dialysis (PD) patients. It is found that CRP is elevated even in ESRD patients receiving conservative care and also in PD patients [41]. This inflammatory state has in turn been linked with progression of CKD and CVD.

More recently, other biomarkers of inflammation have been studied. In the large Chronic Renal Insufficiency Cohort (CRIC) study [42], inflammatory markers (IL-1β, IL-1receptor antagonist, TNF-α, and fibrinogen and in addition to CRP) were found to be negatively correlated with kidney function and positively correlated with albuminuria (a marker of kidney damage). Another study showed an inverse correlation between IL-6 and kidney function [12]. The pattern of elevated pro-inflammatory cytokines (IL-1, IL-6, TNF-α) with low anti-inflammatory cytokines (IL2, IL4, IL5, IL12, CH50 and T cell number) has also been described in haemodialysis patients [43]. CRP, IL-6 and IL-10 levels are significantly higher in ESKD patients compared to controls and the levels did not change significantly after initiation of maintenance haemodialysis [44]. In another study, it was noted that CRP levels actually increased after initiation of haemodialysis [41].

Other markers of inflammatory status involve adipokines. There are proinflammatory adipokines and anti-inflammatory adipokines secreted by adipose tissue. Finally, the National Health and Nutrition Examination Survey (NHANES) demonstrated that levels of pro-inflammatory adipokines were higher in CKD patients than in the general population and that and the ratio of pro- to anti- inflammatory adipokines predicted mortality in PD patients [45].

#### **3.1. Mechanisms of increased systemic inflammation**

#### *3.1.1. Gut dysbiosis*

In addition to promoting oxidative stress, gut dysbiosis may also promote systemic inflammation in patients with CKD. Endotoxemia activates TLR4 on endothelial cells and macrophages thereby leading to activation of NF-kB pathway and ultimately resulting in the production of inflammatory cytokines, chemokines, adhesion molecules, reactive oxygen species and systemic inflammation [39]. Gut-derived uraemic toxins, IS and PCS, are also associated with elevated levels of inflammatory markers such as IL-6, TNF-α, and interferon-γ (IFN- γ) [38]. Higher levels of IS and PCS have been documented in patients with CKD compared to the healthy population [46] and positively correlated with endothelial dysfunction [37].

*3.1.5. Vitamin D deficiency*

*3.1.6. Oxidative stress*

**inflammation**

by vitamin D agonist supplementation [55].

In CKD, vitamin D levels are invariably reduced to various extents. In addition to its role in bone mineral metabolism, vitamin D has been associated with immunologic regulatory and antioxidant functions. For example, vitamin D has been found to potentiate the antioxidant effect of alpha-Klotho protein by increasing its gene expression [55]. Consequently, vitamin D receptor knockout mice have been reported to have augmented DNA damage and increased production of NADPH-dependent superoxide anion production [55]. In mouse models of HIV nephropathy, the downregulation of vitamin D receptor expression was observed together with increased reactive oxygen species generation and DNA damage, which was improved

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A pathologic condition that increases oxidative stress, e.g. ischemia reperfusion injury, simultaneously and subsequently activates inflammatory pathways resulting in a state of both elevated inflammation and oxidative stress. The prominent feature is the two way cross-talk between NOX, NF-kB, inflammasomes and phagocytic cells such as macrophages, resulting in production of both ROS and inflammatory cytokines. The knowledge of underlying path-

In parallel with oxidative stress markers, increases in inflammatory markers have been associated with adverse cardiovascular and mortality outcomes. For example, plasma IL-6 levels have been recognized as an independent predictor of cardiovascular events, atherosclerosis progression and all-cause mortality [44]. Similarly, higher CRP levels correlate with increased carotid artery intima-media thickness in predialysis patients [56] and with increased mortality in both haemodialysis patients [57] and peritoneal patients [58]. Furthermore, Wanner et al. [11] found that a single CRP measurement can predict overall and cardiovascular mortality in the following 4 years. Haemodialysis patients with CRP levels in the highest quartile were associated with a 2.4- to 4.6-fold higher risk of all-cause mortality and a 1.7- to 5.5-fold higher cardiovascular mortality compared to those with CRP levels in the lowest quartile [11, 59]. Similarly, in a prospective observational study involving 62 haemodialysis patients, total mortality was 37.1% and cardiovascular mortality 16.1% at the 2 year follow up. All-cause and cardiovascular mortality were significantly increased at CRP levels above 5 mg/L [60].

ways and mechanisms are evolving due to ongoing research in this area.

**4. Potential therapeutic strategies targeting oxidative stress and** 

With the increasing understanding of the mechanisms underpinning oxidative stress and systemic inflammation in CKD, various interventions have been explored to address these issues (**Table 2**). The challenging aspect of this complex pathological state is that inflammation and

**3.2. Association of inflammation with CKD progression and CVD**

#### *3.1.2. Infection*

Localized or systemic infections are more common in patients with CKD and in turn activate inflammatory and oxidative stress pathways. Dialysis patients are also at higher risk of infections due to presence of foreign bodies such as venous catheters, PD catheters or arteriovenous grafts, predisposing to blood stream infections. Periodontitis is also much more common in patients with CKD compared to normal population and has been linked with heightened cardiovascular risk. In an analysis of 861 CKD patients from NHANES data with a median follow up of 14.3 years, periodontitis was found to be associated with higher mortality. The presence of periodontitis increased the 10-year all-cause mortality from 32% (95% CI 29–35%) to 41% (36–47%), comparable to addition of diabetes to CKD at 43% (95% CI 38–49%) [47].

#### *3.1.3. Dialysis*

CKD patients display increased levels of inflammatory markers even prior to initiation of dialysis. Following commencement of dialysis, studies have variably shown either no change or a worsening of inflammatory and oxidative state [41, 44]. In haemodialysis patients, traces of lipopolysaccharides from dialysate contamination may stimulate the inflammatory process, which may be ameliorated by the use of ultrapure dialysate [48]. The presence of a foreign body, such as an arteriovenous graft, has also been shown to be associated with higher CRP levels and lower albumin levels, indicating a chronic inflammatory process [49]. Dialysis membranes may also have an impact, with cuprophane membranes eliciting higher levels of inflammatory biomarkers than other biocompatible membranes (polyamide or polycarbonate) [50]. Similarly, in PD patients, inflammation may be triggered by PD catheters and dialysis fluids [51].

#### *3.1.4. Metabolic acidosis*

Metabolic acidosis is increasingly more common with more advanced stages of CKD due to impaired renal excretion of acid generated by the body's metabolic processes. In patients with stage 2–4 CKD in the CRIC study, each mmol/L reduction in plasma bicarbonate concentration was associated with a 3% increased risk of progression to ESKD, although there was no association with mortality [52]. In haemodialysis patients, metabolic acidosis has been associated with increased circulating levels of the pro-inflammatory cytokine, IL-6, which was counterbalanced to some extent by increased levels of the anti-inflammatory cytokine, IL-10, and likely reflected a counter-regulatory mechanism [53]. On the other hand, another study by Ori [54] reported increased levels of IL-6 and reduced levels of IL10 indicating greatly augmented inflammatory status.

#### *3.1.5. Vitamin D deficiency*

thereby leading to activation of NF-kB pathway and ultimately resulting in the production of inflammatory cytokines, chemokines, adhesion molecules, reactive oxygen species and systemic inflammation [39]. Gut-derived uraemic toxins, IS and PCS, are also associated with elevated levels of inflammatory markers such as IL-6, TNF-α, and interferon-γ (IFN- γ) [38]. Higher levels of IS and PCS have been documented in patients with CKD compared to the

Localized or systemic infections are more common in patients with CKD and in turn activate inflammatory and oxidative stress pathways. Dialysis patients are also at higher risk of infections due to presence of foreign bodies such as venous catheters, PD catheters or arteriovenous grafts, predisposing to blood stream infections. Periodontitis is also much more common in patients with CKD compared to normal population and has been linked with heightened cardiovascular risk. In an analysis of 861 CKD patients from NHANES data with a median follow up of 14.3 years, periodontitis was found to be associated with higher mortality. The presence of periodontitis increased the 10-year all-cause mortality from 32% (95% CI 29–35%) to 41% (36–47%), comparable to addition of diabetes to CKD at 43% (95% CI 38–49%) [47].

CKD patients display increased levels of inflammatory markers even prior to initiation of dialysis. Following commencement of dialysis, studies have variably shown either no change or a worsening of inflammatory and oxidative state [41, 44]. In haemodialysis patients, traces of lipopolysaccharides from dialysate contamination may stimulate the inflammatory process, which may be ameliorated by the use of ultrapure dialysate [48]. The presence of a foreign body, such as an arteriovenous graft, has also been shown to be associated with higher CRP levels and lower albumin levels, indicating a chronic inflammatory process [49]. Dialysis membranes may also have an impact, with cuprophane membranes eliciting higher levels of inflammatory biomarkers than other biocompatible membranes (polyamide or polycarbonate) [50]. Similarly,

in PD patients, inflammation may be triggered by PD catheters and dialysis fluids [51].

Metabolic acidosis is increasingly more common with more advanced stages of CKD due to impaired renal excretion of acid generated by the body's metabolic processes. In patients with stage 2–4 CKD in the CRIC study, each mmol/L reduction in plasma bicarbonate concentration was associated with a 3% increased risk of progression to ESKD, although there was no association with mortality [52]. In haemodialysis patients, metabolic acidosis has been associated with increased circulating levels of the pro-inflammatory cytokine, IL-6, which was counterbalanced to some extent by increased levels of the anti-inflammatory cytokine, IL-10, and likely reflected a counter-regulatory mechanism [53]. On the other hand, another study by Ori [54] reported increased levels of IL-6 and reduced levels of IL10 indicating greatly

healthy population [46] and positively correlated with endothelial dysfunction [37].

*3.1.2. Infection*

42 Novel Prospects in Oxidative and Nitrosative Stress

*3.1.3. Dialysis*

*3.1.4. Metabolic acidosis*

augmented inflammatory status.

In CKD, vitamin D levels are invariably reduced to various extents. In addition to its role in bone mineral metabolism, vitamin D has been associated with immunologic regulatory and antioxidant functions. For example, vitamin D has been found to potentiate the antioxidant effect of alpha-Klotho protein by increasing its gene expression [55]. Consequently, vitamin D receptor knockout mice have been reported to have augmented DNA damage and increased production of NADPH-dependent superoxide anion production [55]. In mouse models of HIV nephropathy, the downregulation of vitamin D receptor expression was observed together with increased reactive oxygen species generation and DNA damage, which was improved by vitamin D agonist supplementation [55].

#### *3.1.6. Oxidative stress*

A pathologic condition that increases oxidative stress, e.g. ischemia reperfusion injury, simultaneously and subsequently activates inflammatory pathways resulting in a state of both elevated inflammation and oxidative stress. The prominent feature is the two way cross-talk between NOX, NF-kB, inflammasomes and phagocytic cells such as macrophages, resulting in production of both ROS and inflammatory cytokines. The knowledge of underlying pathways and mechanisms are evolving due to ongoing research in this area.

#### **3.2. Association of inflammation with CKD progression and CVD**

In parallel with oxidative stress markers, increases in inflammatory markers have been associated with adverse cardiovascular and mortality outcomes. For example, plasma IL-6 levels have been recognized as an independent predictor of cardiovascular events, atherosclerosis progression and all-cause mortality [44]. Similarly, higher CRP levels correlate with increased carotid artery intima-media thickness in predialysis patients [56] and with increased mortality in both haemodialysis patients [57] and peritoneal patients [58]. Furthermore, Wanner et al. [11] found that a single CRP measurement can predict overall and cardiovascular mortality in the following 4 years. Haemodialysis patients with CRP levels in the highest quartile were associated with a 2.4- to 4.6-fold higher risk of all-cause mortality and a 1.7- to 5.5-fold higher cardiovascular mortality compared to those with CRP levels in the lowest quartile [11, 59]. Similarly, in a prospective observational study involving 62 haemodialysis patients, total mortality was 37.1% and cardiovascular mortality 16.1% at the 2 year follow up. All-cause and cardiovascular mortality were significantly increased at CRP levels above 5 mg/L [60].

### **4. Potential therapeutic strategies targeting oxidative stress and inflammation**

With the increasing understanding of the mechanisms underpinning oxidative stress and systemic inflammation in CKD, various interventions have been explored to address these issues (**Table 2**). The challenging aspect of this complex pathological state is that inflammation and oxidative stress are two very intertwined processes. A primary disorder with elevated oxidative stress would inevitably end up with increased inflammatory state and vice versa. Since multiple cellular components, pathways and end products are involved in the pathogenesis and perpetuation of oxidative stress and inflammation, it has been difficult to attain a clinically meaningful impact by therapy targeted at just one aspect. With the exception of RAAS inhibition that has been well established to improve renal and cardiovascular outcomes in patients with CKD, the evidence for the majority of other interventions has been limited by small studies with short follow up duration, high degrees of heterogeneity and limited data available on patient-level outcomes.

**Lifestyle modifications 1.**Dietary interventions

• Increased dietary fibre intake

• Increased fibre-to-protein ratio

• Protein restriction

• Sodium restriction • Fluid restriction • Pomegranates • Soy milk

**2.**Prebiotics and probiotics

**1.**Use of ultrapure dialysate

**Pharmacologic therapies**

**1.**Oral absorbents **2.**Allopurinol **3.**N-acetyl cysteine

**5.**Bardoxolone **6.**Statins

**Antioxidants 1.**Vitamin E **2.**Vitamin C **3.**L-carnitine **4.**Coenzyme Q-10

**7.**Cytokine therapies

**5.**Miscellaneous antioxidants

**Optimization of dialysis procedure**

**2.**Modification of dialysis membranes • Biocompatible membranes • High-flux membranes • Vitamin E-coated membranes

• Online haemodiafiltration

**4.**Omega-3-polyunsaturated fatty acids

**3.**Modification in dialysis technique and frequency

• Short daily, extended, frequent dialysis sessions

• Vitamin A, selenium, zinc, methionine, alpha-lipoic acid, curcumin

**Table 2.** Interventions in studies to improve oxidative stress and systemic inflammation in chronic kidney disease.

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**3.**Exercise

#### **4.1. Lifestyle modifications**

#### *4.1.1. Dietary interventions*

Dietary management is an integral part of the treatment of CKD, especially in its advanced stages. The current scope of dietary modification includes fluid restriction, sodium, potassium and phosphate restriction, and achievement and maintenance of a healthy body weight.

It has been shown that the uraemic toxins, IS and PCS, are produced by the breakdown of nitrogenous waste products by gut bacteria and are associated with negative renal and cardiovascular outcomes due to systemic inflammation and oxidative stress. Dietary modifications have been explored in order to reduce the production of uraemic toxins by the gut bacteria. It is found that the balance between protein and dietary fibre intake appears to have a more significant impact on the production of uraemic toxins than the absolute intake of either protein or fibre alone. Rossi et al. [61] performed a cross-sectional study of 40 CKD stage 4 and nondialysis CKD 5 patients and correlated the protein-fibre index with serum IS and PCS levels. It was found that the ratio of the protein to fibre intake had significant associations with the serum IS and PCS levels. The total dietary fibre intake had significant associations with PCS but not with IS and the total protein intake had no association with either toxin.

The benefit of dietary fibre supplementation has also been explored in order to increase the carbohydrate content in the colon and to reduce protein fermentation. In a randomized controlled study involving 56 haemodialysis patients, fibre supplementation was found to significantly reduce the level of IS, and to a lesser extent the level of PCS at 6-week follow up [62]. In another non-randomized prospective study, significant reductions in PCS but not IS were found in haemodialysis patients when given dietary fibre supplements [63]. In a large population based cohort study involving 1110 community-dwelling elderly men (around 45% of participants with GFR < 60 ml/min/1.73 m2 ), dietary fibre intake was found to be associated with higher GFR, lower CRP and lower mortality at 10 year follow-up [64]. In a meta-analysis of 14 studies on dietary fibre supplementation in CKD patients, reduction in serum creatinine and urea was noted [65]. However, the trials are very small (n = 3–22) and the dose of fibre supplementation was also highly variable. While the available data suggest dietary fibre supplementation may slow the progression of CKD, robust studies with meaningful clinical end points are lacking.

Nitrogenous waste products can be reduced by dietary protein restriction, which in turn would favour the growth of saccharolytic bacteria over proteolytic bacteria in the colon.

#### **Lifestyle modifications**

oxidative stress are two very intertwined processes. A primary disorder with elevated oxidative stress would inevitably end up with increased inflammatory state and vice versa. Since multiple cellular components, pathways and end products are involved in the pathogenesis and perpetuation of oxidative stress and inflammation, it has been difficult to attain a clinically meaningful impact by therapy targeted at just one aspect. With the exception of RAAS inhibition that has been well established to improve renal and cardiovascular outcomes in patients with CKD, the evidence for the majority of other interventions has been limited by small studies with short follow up duration, high degrees of heterogeneity and limited data

Dietary management is an integral part of the treatment of CKD, especially in its advanced stages. The current scope of dietary modification includes fluid restriction, sodium, potassium and phosphate restriction, and achievement and maintenance of a healthy body weight. It has been shown that the uraemic toxins, IS and PCS, are produced by the breakdown of nitrogenous waste products by gut bacteria and are associated with negative renal and cardiovascular outcomes due to systemic inflammation and oxidative stress. Dietary modifications have been explored in order to reduce the production of uraemic toxins by the gut bacteria. It is found that the balance between protein and dietary fibre intake appears to have a more significant impact on the production of uraemic toxins than the absolute intake of either protein or fibre alone. Rossi et al. [61] performed a cross-sectional study of 40 CKD stage 4 and nondialysis CKD 5 patients and correlated the protein-fibre index with serum IS and PCS levels. It was found that the ratio of the protein to fibre intake had significant associations with the serum IS and PCS levels. The total dietary fibre intake had significant associations with PCS

but not with IS and the total protein intake had no association with either toxin.

The benefit of dietary fibre supplementation has also been explored in order to increase the carbohydrate content in the colon and to reduce protein fermentation. In a randomized controlled study involving 56 haemodialysis patients, fibre supplementation was found to significantly reduce the level of IS, and to a lesser extent the level of PCS at 6-week follow up [62]. In another non-randomized prospective study, significant reductions in PCS but not IS were found in haemodialysis patients when given dietary fibre supplements [63]. In a large population based cohort study involving 1110 community-dwelling elderly men (around 45% of

with higher GFR, lower CRP and lower mortality at 10 year follow-up [64]. In a meta-analysis of 14 studies on dietary fibre supplementation in CKD patients, reduction in serum creatinine and urea was noted [65]. However, the trials are very small (n = 3–22) and the dose of fibre supplementation was also highly variable. While the available data suggest dietary fibre supplementation may slow the progression of CKD, robust studies with meaningful clinical

Nitrogenous waste products can be reduced by dietary protein restriction, which in turn would favour the growth of saccharolytic bacteria over proteolytic bacteria in the colon.

), dietary fibre intake was found to be associated

available on patient-level outcomes.

44 Novel Prospects in Oxidative and Nitrosative Stress

participants with GFR < 60 ml/min/1.73 m2

end points are lacking.

**4.1. Lifestyle modifications**

*4.1.1. Dietary interventions*

**1.**Dietary interventions


**2.**Prebiotics and probiotics

**3.**Exercise

#### **Optimization of dialysis procedure**

**1.**Use of ultrapure dialysate

**2.**Modification of dialysis membranes


**3.**Modification in dialysis technique and frequency


#### **Pharmacologic therapies**

**1.**Oral absorbents


**6.**Statins

**7.**Cytokine therapies

#### **Antioxidants**

	- Vitamin A, selenium, zinc, methionine, alpha-lipoic acid, curcumin

**Table 2.** Interventions in studies to improve oxidative stress and systemic inflammation in chronic kidney disease.

However, the benefit of protein restriction per se in CKD patients is overshadowed by the risk of malnutrition and its complications.

*4.1.3. Exercise*

uting factor for these benefits.

**4.2. Optimization of dialysis**

polysulphone and polypropylene.

Physical exercise has been reported to improve proteinuria and progression of renal function in non-dialysis CKD patients [75] and improve cardiac function and other cardiovascular risk factors in haemodialysis patients. Although multiple pathways can be involved, reduction in the systemic inflammatory state and modulation of immune function may be a major contrib-

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An improvement in inflammatory markers has been noted after bouts of acute exercise or regular long-term exercise with both resistance training or aerobic exercise programs. Viana et al. [76] studied 15 predialysis CKD patients and found that 30 min of walking promoted an anti-inflammatory milieu by increasing the anti-inflammatory cytokine IL-10. An increase in serum IL-6 concentration was also noted, although the authors concluded that the musclederived IL-6 in the study exerted anti-inflammatory effects rather than the pro-inflammatory IL-6β. The group also followed up the patients after 6 months of a regular walking exercise program (30 min per day for 5 days a week) and found that the increased IL-10 levels were sustained with a reduction in the ratio of IL-6 to IL-10. T cell and monocyte activation were downregulated without an effect on the numbers likely representing the modulation of a chronically elevated inflammatory state. In a randomized controlled trial involving 26 non-dialysis CKD patients, resistance exercises for 45 min 3 times a week were also found to reduce CRP and IL-6

after 12 weeks, together with improvements in muscle mass and endurance [77].

of ultrapure dialysate as a standard of care in dialysis therapy.

Given the significantly elevated oxidative stress and systemic inflammatory levels observed in dialysis patients, different modifications to conventional dialytic therapy have been investigated. These interventions include the use of ultrapure dialysate to reduce endotoxin load, changes in membrane types (such as biocompatible membranes, high-flux membranes, and vitamin E coated membranes), modifications in the dialytic techniques (such as on-line haemodiafiltration) and alteration of dialysis frequency (such as short daily dialysis or nocturnal dialysis).

To reduce the inflammation induced by bacterial endotoxins in the dialysate, the use of ultrapure dialysate was investigated. In the meta-analysis by Susantitaphong et al. [25], 31 studies were analysed which included 16 single-arm studies, 5 non-randomized studies and 10 randomized controlled trials. It was found that the use of ultrapure dialysate compared to conventional dialysate significantly reduced CRP and IL-6 levels in all studies. Single arm studies showed a decrease in TNF-α and IL-1 although the controlled trials failed to show a significant decrease in these markers. In terms of oxidative stress state, all studies showed significant decreases in pentosidine, MPO and ox-LDL levels. These findings support the use

Sequelae of bioincompatible membranes in activation of complement and inflammatory pathways have been long recognized [78]. The historic use of bio-incompatible cellulose-based membranes has now been replaced by use of biocompatible synthetic membranes such as

Due to its antioxidant properties, vitamin E has been used as a membrane surface modifier to improve biocompatibility and confer additional benefit with respect to oxidative stress.

Dietary sodium restriction has been explored as a potential strategy for reducing systemic inflammation either directly or indirectly via reducing fluid overload and extracellular volume expansion [66–68]. In a randomized controlled study involving 53 haemodialysis patients, significant reductions in CRP, IL-6 and TNF-α were found in the sodium-restricted group at 8 weeks, which persisted at 16 weeks [69]. However, in a randomized trial by Campbell et al. [70] in patients with stage 3 or 4 CKD, no differences in inflammatory markers were observed after 2 weeks. The apparent disparity in the results of these two studies may be explained by the different stages of renal failure and the duration of intervention.

Pomegranates contain polyphenols, which are known to have antioxidant and anti-inflammatory properties. The number of trials studying on the effect of pomegranate extract or juice on inflammatory markers has been increasing over the last 15 years [71]. There have been a few human trials with varying durations of follow up between 1 day and 12 months. Observed changes in inflammatory markers were variable but the randomized controlled study with longest duration of follow-up of 12 months demonstrated reductions in all inflammatory markers in haemodialysis patients [72].

Soy milk contains isoflavones that are a subgroup of polyphenols with antioxidant properties. Although there have only been small studies with short follow up durations, soy milk has been reported to reduce inflammatory markers and improve renal function deterioration, proteinuria and lipid profile [73].

#### *4.1.2. Prebiotics and probiotics*

A number of studies have been conducted to explore the effect of supplementation of synbiotics (a combination of prebiotics and probiotics) on clinical outcomes in both dialysis and non-dialysis CKD patients. These studies have generally noted reductions in uraemic toxins (dimethylamine, nitrosodimethylamine, blood urea nitrogen, plasma p-cresol) and improvement in gastrointestinal scores and quality of life, although reductions in uric acid and alterations in microbiota have not been consistently identified [16, 39]. In a recent randomized, placebo-controlled cross over trial (SYNERGY) [74], 37 patients with stage 4 or 5 CKD (non-dialysis) received synbiotic treatment or placebo for 6 weeks, followed by a washout period of 4 weeks and a cross over to the alternative treatment arm for a further 6 weeks. The prebiotic component consisted of high-molecular weight inulin, fructo-oligosaccharides and galacto-oligosaccharides. The probiotic component consisted of nine different bacterial strains including *lactobacillus*, *bifidobacteria* and *streptococcus* genera. The administration of synbiotics significantly reduced serum PCS levels and favourably modified the gut bacteria. In a pre-specified analysis in which patients who received antibiotics during the study were excluded, serum IS levels were also significantly reduced by the symbiotic intervention. No significant changes in anti-inflammatory markers were noted. Thus, the study provided proof of concept that synbiotics can modify the gut microbiota and serum uraemic toxin levels in people with CKD. A larger randomized, placebo-controlled trial with 12-month follow up is currently underway which may yield more definite answers regarding the clinical utility of the synbiotic therapy.

#### *4.1.3. Exercise*

However, the benefit of protein restriction per se in CKD patients is overshadowed by the

Dietary sodium restriction has been explored as a potential strategy for reducing systemic inflammation either directly or indirectly via reducing fluid overload and extracellular volume expansion [66–68]. In a randomized controlled study involving 53 haemodialysis patients, significant reductions in CRP, IL-6 and TNF-α were found in the sodium-restricted group at 8 weeks, which persisted at 16 weeks [69]. However, in a randomized trial by Campbell et al. [70] in patients with stage 3 or 4 CKD, no differences in inflammatory markers were observed after 2 weeks. The apparent disparity in the results of these two studies may be explained by

Pomegranates contain polyphenols, which are known to have antioxidant and anti-inflammatory properties. The number of trials studying on the effect of pomegranate extract or juice on inflammatory markers has been increasing over the last 15 years [71]. There have been a few human trials with varying durations of follow up between 1 day and 12 months. Observed changes in inflammatory markers were variable but the randomized controlled study with longest duration of follow-up of 12 months demonstrated reductions in all inflammatory

Soy milk contains isoflavones that are a subgroup of polyphenols with antioxidant properties. Although there have only been small studies with short follow up durations, soy milk has been reported to reduce inflammatory markers and improve renal function deterioration,

A number of studies have been conducted to explore the effect of supplementation of synbiotics (a combination of prebiotics and probiotics) on clinical outcomes in both dialysis and non-dialysis CKD patients. These studies have generally noted reductions in uraemic toxins (dimethylamine, nitrosodimethylamine, blood urea nitrogen, plasma p-cresol) and improvement in gastrointestinal scores and quality of life, although reductions in uric acid and alterations in microbiota have not been consistently identified [16, 39]. In a recent randomized, placebo-controlled cross over trial (SYNERGY) [74], 37 patients with stage 4 or 5 CKD (non-dialysis) received synbiotic treatment or placebo for 6 weeks, followed by a washout period of 4 weeks and a cross over to the alternative treatment arm for a further 6 weeks. The prebiotic component consisted of high-molecular weight inulin, fructo-oligosaccharides and galacto-oligosaccharides. The probiotic component consisted of nine different bacterial strains including *lactobacillus*, *bifidobacteria* and *streptococcus* genera. The administration of synbiotics significantly reduced serum PCS levels and favourably modified the gut bacteria. In a pre-specified analysis in which patients who received antibiotics during the study were excluded, serum IS levels were also significantly reduced by the symbiotic intervention. No significant changes in anti-inflammatory markers were noted. Thus, the study provided proof of concept that synbiotics can modify the gut microbiota and serum uraemic toxin levels in people with CKD. A larger randomized, placebo-controlled trial with 12-month follow up is currently underway which may yield more definite answers regarding the clinical utility of

the different stages of renal failure and the duration of intervention.

risk of malnutrition and its complications.

46 Novel Prospects in Oxidative and Nitrosative Stress

markers in haemodialysis patients [72].

proteinuria and lipid profile [73].

*4.1.2. Prebiotics and probiotics*

the synbiotic therapy.

Physical exercise has been reported to improve proteinuria and progression of renal function in non-dialysis CKD patients [75] and improve cardiac function and other cardiovascular risk factors in haemodialysis patients. Although multiple pathways can be involved, reduction in the systemic inflammatory state and modulation of immune function may be a major contributing factor for these benefits.

An improvement in inflammatory markers has been noted after bouts of acute exercise or regular long-term exercise with both resistance training or aerobic exercise programs. Viana et al. [76] studied 15 predialysis CKD patients and found that 30 min of walking promoted an anti-inflammatory milieu by increasing the anti-inflammatory cytokine IL-10. An increase in serum IL-6 concentration was also noted, although the authors concluded that the musclederived IL-6 in the study exerted anti-inflammatory effects rather than the pro-inflammatory IL-6β. The group also followed up the patients after 6 months of a regular walking exercise program (30 min per day for 5 days a week) and found that the increased IL-10 levels were sustained with a reduction in the ratio of IL-6 to IL-10. T cell and monocyte activation were downregulated without an effect on the numbers likely representing the modulation of a chronically elevated inflammatory state. In a randomized controlled trial involving 26 non-dialysis CKD patients, resistance exercises for 45 min 3 times a week were also found to reduce CRP and IL-6 after 12 weeks, together with improvements in muscle mass and endurance [77].

### **4.2. Optimization of dialysis**

Given the significantly elevated oxidative stress and systemic inflammatory levels observed in dialysis patients, different modifications to conventional dialytic therapy have been investigated. These interventions include the use of ultrapure dialysate to reduce endotoxin load, changes in membrane types (such as biocompatible membranes, high-flux membranes, and vitamin E coated membranes), modifications in the dialytic techniques (such as on-line haemodiafiltration) and alteration of dialysis frequency (such as short daily dialysis or nocturnal dialysis).

To reduce the inflammation induced by bacterial endotoxins in the dialysate, the use of ultrapure dialysate was investigated. In the meta-analysis by Susantitaphong et al. [25], 31 studies were analysed which included 16 single-arm studies, 5 non-randomized studies and 10 randomized controlled trials. It was found that the use of ultrapure dialysate compared to conventional dialysate significantly reduced CRP and IL-6 levels in all studies. Single arm studies showed a decrease in TNF-α and IL-1 although the controlled trials failed to show a significant decrease in these markers. In terms of oxidative stress state, all studies showed significant decreases in pentosidine, MPO and ox-LDL levels. These findings support the use of ultrapure dialysate as a standard of care in dialysis therapy.

Sequelae of bioincompatible membranes in activation of complement and inflammatory pathways have been long recognized [78]. The historic use of bio-incompatible cellulose-based membranes has now been replaced by use of biocompatible synthetic membranes such as polysulphone and polypropylene.

Due to its antioxidant properties, vitamin E has been used as a membrane surface modifier to improve biocompatibility and confer additional benefit with respect to oxidative stress. A recent meta-analysis conducted by D'Arrigo et al. [79] included 60 studies, 23 of which were randomized controlled studies and 37 were non-randomized studies. Improvement in oxidative stress was evidenced by a decrease in MDA and TBARS, without any change in other parameters, such as SOD or NOX. Reduction in ox-LDL levels became significant after improving heterogeneity by excluding three parallel studies. The use of vitamin E-coated membranes reduced the IL-6 levels without any change in CRP.

with placebo (HR 1.03, 95% CI 0.84–1.27, p = 0.78 in EPPIC-1 and HR-0.91, 95% CI 0.74–1.12, p = 0.37). Notwithstanding their apparent lack of efficacy in these large trials, the main limitations of orally administered absorbents is poor compliance due to pill burden with the need to take 30 pills a day, and gastrointestinal side effects, such as constipation, diarrhoea, nausea,

The Role of Oxidative Stress and Systemic Inflammation in Kidney Disease and Its Associated…

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49

Uric acid is the final product of purine metabolism. Production of uric acid is catalysed by the enzyme xanthine oxidase, which also generates reactive oxygen species in the process. Hyperuricaemia has been found to be associated with increased RAAS activity [85], hypertension [86], endothelial dysfunction [87] and cardiovascular disease [88]. It is also associated with higher mortality in patients with non-dialysis CKD patients [89] as well as haemodialysis patients [90]. A number of single centre studies have shown that inhibition of xanthine oxidase by allopurinol or febuxostat may slow the progression of renal disease and improve cardiovascular outcomes [91, 92]. In the meta-analysis by Bose et al. [93] involving eight randomized controlled studies of allopurinol treatment found that no significant changes in glomerular filtration rate (GFR) in five studies, but improvement in serum creatinine concentrations in three studies. There was appreciable heterogeneity in the studies in terms of baseline GFR (three studies with baseline GFR > 67 mL/

), follow up duration (4–24 months) and etiology of CKD. They were all single centre

in the preceding 12 months.

and

studies with small sample sizes (n = 36–113). Since the publication of the meta-analysis, a long-term follow up outcome of a previous trial by Goicoechea et al. [94], and four additional randomized trials have been published. Goicoechea et al. [94] found that there was reduction in the number of renal and cardiovascular events in the allopurinol arm compared to the control group (HR 0.32 and HR 0.43 respectively) at 5 years of follow up. However, definitive conclusions regarding the

Currently there are three multi-centre, randomized, double-blinded prospective trials being conducted. The first trial, *the CKD-FIX Trial: Controlled trial of slowing of Kidney Disease progression From the Inhibition of Xanthine oxidase*, will involve 620 patients with CKD stage 3 or 4

Hyperuricaemia is not mandatory for inclusion. The intervention will be allopurinol dose escalated from 100 mg daily to 300 mg daily in a stepwise manner according to patient tolerance. The follow up period will be 104 weeks and the primary outcome measure of changes in GFR will be assessed. The second trial, *FEATHER Trial: FEbuxostat versus placebo rAndomized controlled Trial regarding reduced renal function in patients with Hyperuricemia complicated by chRonic kidney disease stage 3,* will titrate febuxostat dose from 10 to 40 mg in the first 9 weeks in 400 participants with CKD stage 3. Hyperuricaemia with serum uric acid 7.1–10 mg/dL is required for inclusion in the study. The primary outcome measure of GFR slope will be assessed. The third trial, *The Preventing Early Renal Function Loss (PERL) Allopurinol Study,* will include subjects with type1 diabetes with albuminuria, GFR 45–100 ml/min/1.73 m2

serum uric acid ≥4.5 mg/dL. The primary outcome measure of GFR will be measured at the end of the three-year study period. At the conclusion of the current trials, it is hoped that more robust evidence will be obtained regarding the effect of uric acid lowering therapy on

safety and efficacy of urate lowering therapies in CKD cannot be drawn at this stage.

and albuminuria and decline in GFR of at least 3 ml/min/1.73 m2

progression of renal disease in patients with pre-existing CKD.

abdominal distension and flatulence.

*4.3.2. Allopurinol*

min/1.73 m2

The clearance of pro-oxidant middle molecules, such as uraemic toxins, in haemodialysis may be enhanced by online haemodiafiltration. Using data from a large randomized controlled trial CONvective TRAnsport Study (CONTRAST) investigating the mortality and cardiovascular outcomes of online haemodiafiltration versus conventional low flux haemodialysis, Den Hoedt et al. [80] analysed a sub-group of 405 patients for changes in inflammatory markers after 3 years. Significant differences in the CRP and IL-6 levels became apparent after 6 months of the study. However, mortality and cardiovascular benefits were not noted in the main study.

Increasing dialysate flow rate, utilizing super-flux membranes or adding a sorbent to dialysate could also potentially improve the clearance of gut-derived, protein-bound uraemic toxins. A small study found that increasing the dialyzer mass transfer area coefficient by using two dialyzers in series improved the clearance of protein-bound molecules compared to the conventional use of a single dialyzer. Another small study also showed that by increasing the dialyzer size and dialysate flow in nocturnal haemodialysis patients, the clearance of IS and PCS were enhanced [16].

In terms of dialysis duration and frequency, short daily dialysis (3 h sessions for 6 days a week) was associated with significantly reduced levels of CRP compared with conventional dialysis (4 h sessions for 3 days a week) with corresponding improvement in erythropoietin stimulating agent sensitivity [81]. In a meta-analysis by Susantitaphong [82], there was improvement in cardiac parameters, such as left ventricular mass index (LVMI), left ventricular ejection fraction (LVEF) and blood pressure, in patients using frequent (2–8 h, > thrice weekly) or extended (>4 h, thrice weekly) haemodialysis, compared to a conventional (≤4 h, thrice weekly) haemodialysis schedule. Beyond utilization of ultrapure dialysate and highflux online haemodiafiltration, the addition of further convective permeability by utilizing hyper high flux membranes did not confer any extra benefits in oxidative stress markers [83].

#### **4.3. Pharmacologic therapies**

#### *4.3.1. Oral absorbents*

AST-120 is the only oral charcoal absorbent available to reduce the absorption of indole in the gastrointestinal tract, thereby reducing the systemic concentrations of IS. It is possible that other uraemic toxins are also absorbed. It is widely used in CKD patients in a number of Asian countries to prolong the time to initiation of dialysis. Initial retrospective and prospective studies showed an enhanced clearance of the toxins as well as improved clinical outcomes with AST-120 [16]. However, in two randomized controlled trials (EPPIC-1 and EPPIC-2) [84] involving 2035 patients with moderate to severe CKD (sCr 2–5 mg/dL for men and 1.5–5 mg/dL for women at screening) in 13 countries, AST-120 did not significantly alter the primary composite end point of dialysis initiation, transplantation or doubling of serum creatinine compared with placebo (HR 1.03, 95% CI 0.84–1.27, p = 0.78 in EPPIC-1 and HR-0.91, 95% CI 0.74–1.12, p = 0.37). Notwithstanding their apparent lack of efficacy in these large trials, the main limitations of orally administered absorbents is poor compliance due to pill burden with the need to take 30 pills a day, and gastrointestinal side effects, such as constipation, diarrhoea, nausea, abdominal distension and flatulence.

#### *4.3.2. Allopurinol*

A recent meta-analysis conducted by D'Arrigo et al. [79] included 60 studies, 23 of which were randomized controlled studies and 37 were non-randomized studies. Improvement in oxidative stress was evidenced by a decrease in MDA and TBARS, without any change in other parameters, such as SOD or NOX. Reduction in ox-LDL levels became significant after improving heterogeneity by excluding three parallel studies. The use of vitamin E-coated

The clearance of pro-oxidant middle molecules, such as uraemic toxins, in haemodialysis may be enhanced by online haemodiafiltration. Using data from a large randomized controlled trial CONvective TRAnsport Study (CONTRAST) investigating the mortality and cardiovascular outcomes of online haemodiafiltration versus conventional low flux haemodialysis, Den Hoedt et al. [80] analysed a sub-group of 405 patients for changes in inflammatory markers after 3 years. Significant differences in the CRP and IL-6 levels became apparent after 6 months of the study. However, mortality and cardiovascular benefits were not noted in the main study. Increasing dialysate flow rate, utilizing super-flux membranes or adding a sorbent to dialysate could also potentially improve the clearance of gut-derived, protein-bound uraemic toxins. A small study found that increasing the dialyzer mass transfer area coefficient by using two dialyzers in series improved the clearance of protein-bound molecules compared to the conventional use of a single dialyzer. Another small study also showed that by increasing the dialyzer size and dialysate flow in nocturnal haemodialysis patients, the clearance of IS and PCS were enhanced [16].

In terms of dialysis duration and frequency, short daily dialysis (3 h sessions for 6 days a week) was associated with significantly reduced levels of CRP compared with conventional dialysis (4 h sessions for 3 days a week) with corresponding improvement in erythropoietin stimulating agent sensitivity [81]. In a meta-analysis by Susantitaphong [82], there was improvement in cardiac parameters, such as left ventricular mass index (LVMI), left ventricular ejection fraction (LVEF) and blood pressure, in patients using frequent (2–8 h, > thrice weekly) or extended (>4 h, thrice weekly) haemodialysis, compared to a conventional (≤4 h, thrice weekly) haemodialysis schedule. Beyond utilization of ultrapure dialysate and highflux online haemodiafiltration, the addition of further convective permeability by utilizing hyper high flux membranes did not confer any extra benefits in oxidative stress markers [83].

AST-120 is the only oral charcoal absorbent available to reduce the absorption of indole in the gastrointestinal tract, thereby reducing the systemic concentrations of IS. It is possible that other uraemic toxins are also absorbed. It is widely used in CKD patients in a number of Asian countries to prolong the time to initiation of dialysis. Initial retrospective and prospective studies showed an enhanced clearance of the toxins as well as improved clinical outcomes with AST-120 [16]. However, in two randomized controlled trials (EPPIC-1 and EPPIC-2) [84] involving 2035 patients with moderate to severe CKD (sCr 2–5 mg/dL for men and 1.5–5 mg/dL for women at screening) in 13 countries, AST-120 did not significantly alter the primary composite end point of dialysis initiation, transplantation or doubling of serum creatinine compared

membranes reduced the IL-6 levels without any change in CRP.

48 Novel Prospects in Oxidative and Nitrosative Stress

**4.3. Pharmacologic therapies**

*4.3.1. Oral absorbents*

Uric acid is the final product of purine metabolism. Production of uric acid is catalysed by the enzyme xanthine oxidase, which also generates reactive oxygen species in the process. Hyperuricaemia has been found to be associated with increased RAAS activity [85], hypertension [86], endothelial dysfunction [87] and cardiovascular disease [88]. It is also associated with higher mortality in patients with non-dialysis CKD patients [89] as well as haemodialysis patients [90]. A number of single centre studies have shown that inhibition of xanthine oxidase by allopurinol or febuxostat may slow the progression of renal disease and improve cardiovascular outcomes [91, 92]. In the meta-analysis by Bose et al. [93] involving eight randomized controlled studies of allopurinol treatment found that no significant changes in glomerular filtration rate (GFR) in five studies, but improvement in serum creatinine concentrations in three studies. There was appreciable heterogeneity in the studies in terms of baseline GFR (three studies with baseline GFR > 67 mL/ min/1.73 m2 ), follow up duration (4–24 months) and etiology of CKD. They were all single centre studies with small sample sizes (n = 36–113). Since the publication of the meta-analysis, a long-term follow up outcome of a previous trial by Goicoechea et al. [94], and four additional randomized trials have been published. Goicoechea et al. [94] found that there was reduction in the number of renal and cardiovascular events in the allopurinol arm compared to the control group (HR 0.32 and HR 0.43 respectively) at 5 years of follow up. However, definitive conclusions regarding the safety and efficacy of urate lowering therapies in CKD cannot be drawn at this stage.

Currently there are three multi-centre, randomized, double-blinded prospective trials being conducted. The first trial, *the CKD-FIX Trial: Controlled trial of slowing of Kidney Disease progression From the Inhibition of Xanthine oxidase*, will involve 620 patients with CKD stage 3 or 4 and albuminuria and decline in GFR of at least 3 ml/min/1.73 m2 in the preceding 12 months. Hyperuricaemia is not mandatory for inclusion. The intervention will be allopurinol dose escalated from 100 mg daily to 300 mg daily in a stepwise manner according to patient tolerance. The follow up period will be 104 weeks and the primary outcome measure of changes in GFR will be assessed. The second trial, *FEATHER Trial: FEbuxostat versus placebo rAndomized controlled Trial regarding reduced renal function in patients with Hyperuricemia complicated by chRonic kidney disease stage 3,* will titrate febuxostat dose from 10 to 40 mg in the first 9 weeks in 400 participants with CKD stage 3. Hyperuricaemia with serum uric acid 7.1–10 mg/dL is required for inclusion in the study. The primary outcome measure of GFR slope will be assessed. The third trial, *The Preventing Early Renal Function Loss (PERL) Allopurinol Study,* will include subjects with type1 diabetes with albuminuria, GFR 45–100 ml/min/1.73 m2 and serum uric acid ≥4.5 mg/dL. The primary outcome measure of GFR will be measured at the end of the three-year study period. At the conclusion of the current trials, it is hoped that more robust evidence will be obtained regarding the effect of uric acid lowering therapy on progression of renal disease in patients with pre-existing CKD.

#### *4.3.3. N-acetyl cysteine*

N-acetyl cysteine provides L-cysteine, which is the rate-limiting precursor to glutathione synthesis, thereby enhancing antioxidant defenses. It also acts as a scavenger of free radicals. Even though there is evidence of reduction in oxidative activity in animal models and dialysis patients [95–97], a small randomized controlled trial in patients with proteinuria and early CKD showed no difference in proteinuria between patients treated with N-acetyl cysteine and placebo [98].

an additional role in improving systemic inflammation. In a meta-analysis by Deng et al. [109], nine randomized controlled trials involving 3098 dialysis patients were analysed. Three studies assessed CRP and six studies utilized hs-CRP. One study also included IL-6 and TNF-α in the assessment. All the studies found significant reductions in CRP and hsCRP in the patients treated with statins, while the control group experienced an increase or no change in inflammatory markers. IL-6 levels did not change but there was a significant decrease in TNF-α in one study. Overall, there is evidence that improvement in systematic inflammatory status achieved by statins may play a contributory role in primary and secondary prevention of atherosclerosis.

The Role of Oxidative Stress and Systemic Inflammation in Kidney Disease and Its Associated…

http://dx.doi.org/10.5772/intechopen.73239

51

Attempts have been made to directly target pro-inflammatory cytokines by investigating the role of anti-IL-1, anti-IL-6 and TNF-α. In a small, randomized controlled trial with 22 haemodialysis patients, administration of an IL-1β antagonist improved the levels of hs-CRP and IL-6 and anti-inflammatory adiponectin after 4 weeks [110]. Currently, the available evidence

A number of small trials have been conducted using antioxidants of various types, such as vitamins, naturally occurring dietary extracts, and trace elements. Of the current available information, the Cochrane systematic review in 2012, prior to the BEACON study, concluded that antioxidants confer a significant reduction in serum creatinine, changes in GFR and risk of ESKD but no difference in cardiovascular outcomes [111]. However, there was a high degree of heterogeneity in the meta-analysis. In another meta-analysis on diabetic kidney disease, it was found that there was a

The vitamin E family consists of tocophenols in saturated form, and tocotrienols in unsaturated form with a side chain of an isoprenoid. They both play a role in reducing oxidative stress by scavenging free radicals, inhibiting pro-inflammatory pathways and increasing levels of other antioxidants. Meta-analyses of studies in haemodialysis patients using vitamin-E coated dialysis membranes, and oral supplements in CKD patients failed to improve any clinical outcomes although some studies did show improvements in oxidative markers [79, 112, 113]. In a randomized controlled study, *Secondary prevention with antioxidants of cardiovascular disease in end stage renal disease* [114]*,* 196 haemodialysis patients were randomized to vitamin E group receiving 800 IU/day or placebo and followed up for 519 days. At the end of the study period, there was a 40% reduction in cardiovascular end points in the group receiving vitamin E, mainly driven by a reduction in the incidence of myocardial infarction. In contrast, a posthoc analysis of another randomized controlled study, *Heart Outcomes Prevention Evaluation* [115], involving 993 patients with mild to moderate renal insufficiency, found no benefit from administration of vitamin E 400 IU/day (RR 1.03, CI 0.79–1.34, p- = 0.82). The apparent disparity in findings between the two studies may be due to the differences in the degree of renal impairment and dose of vitamin E. Nevertheless, the role of vitamin E in mitigating cardio-

significant reduction in albuminuria, but no evidence in other renal outcomes [112].

vascular risk in CKD patients remains uncertain at this point in time.

*4.3.7. Cytokine therapies*

**4.4. Antioxidants**

*4.4.1. Vitamin E*

of the direct anticytokine therapy is limited.

#### *4.3.4. Omega-3-polyunsaturated fatty acids*

Eicosapentaenoic and docosahexaenoic acids are the two major bioactive omega-3 fatty acids mainly derived from dietary sources. In animal models, they have been shown to improve the anti-oxidant systems and reduce inflammation and tubulointerstitial fibrosis [99]. Numerous studies have been conducted in dialysis patients investigating their effects on inflammatory markers, nutritional status and lipid profile. In haemodialysis patients, there is evidence of inhibition of up-regulation of endothelial chemokines [100] and reduction of all-cause mortality [101]. However, a study conducted in continuous ambulatory peritoneal dialysis (CAPD) patients did not show any significant changes in SOD and reduced glutathione (GH) levels [102]. In non-dialysis CKD patients, IL-1β and TBARS were reduced and SOD and GH levels were improved, but no effect on IL-6 and TNF-α was noted [103, 104].

#### *4.3.5. Bardoxolone*

Nuclear factor erythroid 2-related factor (Nrf-2) is a nuclear transcription factor which generates the production of antioxidant enzymes via induction of antioxidant response element (ARE) genes. It is activated by increased oxidative stress, such as reactive oxygen and nitrogen species, and induces the ARE gene. This in turn results in production of reducing factors such as NADPH and the elimination of reactive oxygen species by antioxidant enzymes such as SOD and GSH-Px [105]. In animal models, reduced activity of Nrf-2 produced tubular injury and progressive fibrosis which can be ameliorated by administration of Nrf-2 activators [106]. In a randomized controlled trial involving 227 diabetic CKD patients, improvement in the renal function was noted at 24 weeks which persisted at 52 weeks in the intervention group with Nrf-2 activator, bardoxolone methyl [107]. Based on these initial findings, a prospective, randomized controlled trial was conducted in 2185 patients with type 2 diabetes mellitus and stage 4 CKD with the intervention of bardoxolone methyl (20 mg daily per os) versus placebo [108]. At 9 months, the patients receiving bardoxolone methyl experienced a significantly higher rate of heart failure-related hospitalizations or deaths (hazard ratio 1.83, 95% CI 1.32–2.55, p < 0.001) prompting premature termination of the trial. Bardoxolone methyl did not reduce the risk of the primary composite end-point of ESKD or death from cardiovascular causes (HR 0.98, 95% CI 0.70—1.37, p = 0.92). Although further trials are underway addressing this aspect, the potential therapeutic role of bardoxolone methyl in patients with CKD appears limited.

#### *4.3.6. Statins*

Statins, 3-hydroxyl-3-methylglutaryl coenzyme A reductase inhibitors, are well-established treatment for hypercholesterolemia. New evidence has recently emerged that statins may have an additional role in improving systemic inflammation. In a meta-analysis by Deng et al. [109], nine randomized controlled trials involving 3098 dialysis patients were analysed. Three studies assessed CRP and six studies utilized hs-CRP. One study also included IL-6 and TNF-α in the assessment. All the studies found significant reductions in CRP and hsCRP in the patients treated with statins, while the control group experienced an increase or no change in inflammatory markers. IL-6 levels did not change but there was a significant decrease in TNF-α in one study. Overall, there is evidence that improvement in systematic inflammatory status achieved by statins may play a contributory role in primary and secondary prevention of atherosclerosis.

#### *4.3.7. Cytokine therapies*

*4.3.3. N-acetyl cysteine*

*4.3.5. Bardoxolone*

*4.3.6. Statins*

*4.3.4. Omega-3-polyunsaturated fatty acids*

50 Novel Prospects in Oxidative and Nitrosative Stress

N-acetyl cysteine provides L-cysteine, which is the rate-limiting precursor to glutathione synthesis, thereby enhancing antioxidant defenses. It also acts as a scavenger of free radicals. Even though there is evidence of reduction in oxidative activity in animal models and dialysis patients [95–97], a small randomized controlled trial in patients with proteinuria and early CKD showed no difference in proteinuria between patients treated with N-acetyl cysteine and placebo [98].

Eicosapentaenoic and docosahexaenoic acids are the two major bioactive omega-3 fatty acids mainly derived from dietary sources. In animal models, they have been shown to improve the anti-oxidant systems and reduce inflammation and tubulointerstitial fibrosis [99]. Numerous studies have been conducted in dialysis patients investigating their effects on inflammatory markers, nutritional status and lipid profile. In haemodialysis patients, there is evidence of inhibition of up-regulation of endothelial chemokines [100] and reduction of all-cause mortality [101]. However, a study conducted in continuous ambulatory peritoneal dialysis (CAPD) patients did not show any significant changes in SOD and reduced glutathione (GH) levels [102]. In non-dialysis CKD patients, IL-1β and TBARS were reduced and SOD and GH levels

Nuclear factor erythroid 2-related factor (Nrf-2) is a nuclear transcription factor which generates the production of antioxidant enzymes via induction of antioxidant response element (ARE) genes. It is activated by increased oxidative stress, such as reactive oxygen and nitrogen species, and induces the ARE gene. This in turn results in production of reducing factors such as NADPH and the elimination of reactive oxygen species by antioxidant enzymes such as SOD and GSH-Px [105]. In animal models, reduced activity of Nrf-2 produced tubular injury and progressive fibrosis which can be ameliorated by administration of Nrf-2 activators [106]. In a randomized controlled trial involving 227 diabetic CKD patients, improvement in the renal function was noted at 24 weeks which persisted at 52 weeks in the intervention group with Nrf-2 activator, bardoxolone methyl [107]. Based on these initial findings, a prospective, randomized controlled trial was conducted in 2185 patients with type 2 diabetes mellitus and stage 4 CKD with the intervention of bardoxolone methyl (20 mg daily per os) versus placebo [108]. At 9 months, the patients receiving bardoxolone methyl experienced a significantly higher rate of heart failure-related hospitalizations or deaths (hazard ratio 1.83, 95% CI 1.32–2.55, p < 0.001) prompting premature termination of the trial. Bardoxolone methyl did not reduce the risk of the primary composite end-point of ESKD or death from cardiovascular causes (HR 0.98, 95% CI 0.70—1.37, p = 0.92). Although further trials are underway addressing this aspect, the poten-

tial therapeutic role of bardoxolone methyl in patients with CKD appears limited.

Statins, 3-hydroxyl-3-methylglutaryl coenzyme A reductase inhibitors, are well-established treatment for hypercholesterolemia. New evidence has recently emerged that statins may have

were improved, but no effect on IL-6 and TNF-α was noted [103, 104].

Attempts have been made to directly target pro-inflammatory cytokines by investigating the role of anti-IL-1, anti-IL-6 and TNF-α. In a small, randomized controlled trial with 22 haemodialysis patients, administration of an IL-1β antagonist improved the levels of hs-CRP and IL-6 and anti-inflammatory adiponectin after 4 weeks [110]. Currently, the available evidence of the direct anticytokine therapy is limited.

#### **4.4. Antioxidants**

A number of small trials have been conducted using antioxidants of various types, such as vitamins, naturally occurring dietary extracts, and trace elements. Of the current available information, the Cochrane systematic review in 2012, prior to the BEACON study, concluded that antioxidants confer a significant reduction in serum creatinine, changes in GFR and risk of ESKD but no difference in cardiovascular outcomes [111]. However, there was a high degree of heterogeneity in the meta-analysis. In another meta-analysis on diabetic kidney disease, it was found that there was a significant reduction in albuminuria, but no evidence in other renal outcomes [112].

#### *4.4.1. Vitamin E*

The vitamin E family consists of tocophenols in saturated form, and tocotrienols in unsaturated form with a side chain of an isoprenoid. They both play a role in reducing oxidative stress by scavenging free radicals, inhibiting pro-inflammatory pathways and increasing levels of other antioxidants. Meta-analyses of studies in haemodialysis patients using vitamin-E coated dialysis membranes, and oral supplements in CKD patients failed to improve any clinical outcomes although some studies did show improvements in oxidative markers [79, 112, 113]. In a randomized controlled study, *Secondary prevention with antioxidants of cardiovascular disease in end stage renal disease* [114]*,* 196 haemodialysis patients were randomized to vitamin E group receiving 800 IU/day or placebo and followed up for 519 days. At the end of the study period, there was a 40% reduction in cardiovascular end points in the group receiving vitamin E, mainly driven by a reduction in the incidence of myocardial infarction. In contrast, a posthoc analysis of another randomized controlled study, *Heart Outcomes Prevention Evaluation* [115], involving 993 patients with mild to moderate renal insufficiency, found no benefit from administration of vitamin E 400 IU/day (RR 1.03, CI 0.79–1.34, p- = 0.82). The apparent disparity in findings between the two studies may be due to the differences in the degree of renal impairment and dose of vitamin E. Nevertheless, the role of vitamin E in mitigating cardiovascular risk in CKD patients remains uncertain at this point in time.

#### *4.4.2. Vitamin C*

Vitamin C exerts its antioxidant properties by acting as an electron donor to free radicals. A number of small studies have found improvements in inflammatory markers such as CRP [116], hsCRP [117], and 8-OHdG [118] in haemodialysis patients. In a meta-analysis of randomized controlled trials examining use of antioxidants in diabetic kidney disease, vitamin C reduced albuminuria in some studies but had no effect on GFR [111]. All the studies were small (n = 14–29) with short durations of follow up (4 weeks to 12 months) and generally were of suboptimal methodologic quality. Consequently, no conclusions can currently be drawn regarding the safety and efficacy of vitamin C therapy in patients with CKD.

**5. Future directions**

evaluate such potential therapies.

**Author details**

**References**

**80**(1):17-28

David W. Johnson1,2,3,5\*

Brisbane, Queensland, Australia

CKD constitutes a state of increased oxidative stress and systemic inflammation. These processes are pathogenetically interrelated and there is increasing evidence that they may contribute to CKD progression and a disproportionately increased cardiovascular risk through the promotion of endothelial dysfunction, atherosclerosis and vascular calcification. The various mechanisms of action of this increased oxidative stress are increasingly being elucidated. Interventions to reduce oxidative stress and inflammation in these patients present novel, and potentially effective approaches to add to the currently available traditional risk-modification strategies. Due to the complex interrelation between reactive oxygen species and inflammatory markers, it is possible that simultaneous, multiple targeted approaches may be required to effectively address the pathological changes in CKD and its associated cardiovascular risk. Larger trials with meaningful clinical outcomes and longer follow up are required to further

The Role of Oxidative Stress and Systemic Inflammation in Kidney Disease and Its Associated…

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53

Aye San1,2, Magid Fahim1,2,3, Katrina Campbell1,3,4, Carmel M. Hawley1,2,3,5 and

2 School of Medicine, University of Queensland, Brisbane, Queensland, Australia

3 Australasian Kidney Trials Network, School of Medicine, University of Queensland,

1 Department of Nephrology, Princess Alexandra Hospital, Brisbane, Queensland, Australia

[1] Levey AS, De Jong PE, Coresh J, Nahas ME, Astor BC, Matsushita K, Gansevoort RT, Kasiske BL, Eckardt KU. The definition, classification, and prognosis of chronic kidney disease: A KDIGO Controversies Conference Report. Kidney International. 2011;

[2] Bello AK, Levin A, Tonelli M, Okpechi IG, Feehally J, Harris D, Jindal K, Salako BL, Rateb A, Osman MA, Qarni B. Assessment of global kidney health care status. Journal of

the American Medical Association. 2017 May 9;**317**(18):1864-1881

\*Address all correspondence to: david.johnson2@health.qld.gov.au

4 Faculty of Health Sciences and Medicine, Bond University, Australia

5 Translational Research Institute, Brisbane, Queensland, Australia

#### *4.4.3. L-carnitine*

Carnitine is an endogenous product of amino acid metabolism produced in the liver. It acts as a transporter of long chain fatty acids across the mitochondrial membrane by reversibly substituting the acyl group of Coenzyme A, forming acyl-carnitine. Once the fatty acids have been transported into the mitochondrial matrix, acyl-carnitine dissociates to form L-carnitine again and coenzyme A is regenerated. L-carnitine has been noted to reduce the oxidative stress through increased glutathione levels, increased GSH-Px activity, and a decreased MDA levels. In a meta-analysis by Chen et al. [119] of 49 RCTs involving 1734 haemodialysis patients, L-carnitine was found to improve CRP and LDL levels. However, in another meta-analysis by Yang et al. [120] involving 25 RCTs, contradictory evidence was found in that L-carnitine did not appreciably alter inflammation, oxidative stress, hyperlipidaemia or quality of life. Currently, there is insufficient evidence to support L-carnitine administration to CKD patients.

#### *4.4.4. Coenzyme Q10*

Coenzyme Q10 is a ubiquinone which contains one quinine group and 10 isoprenyl units. It acts as an enzyme co-factor in inner mitochondrial cell membranes protecting against damage from free radicals produced by oxidative phosphorylation. It also stabilizes the cell membranes as an electron and proton carrier and restores vitamin E in its antioxidant form. Early studies in rat models of diabetic nephropathy showed reduced mesangial expansion and tubulointerstitial fibrosis after administration of mitochondrial-targeted coenzyme Q10 [121]. Two recent studies in dialysis patients showed significant reductions in F2-isoprostanes and isofurans at high doses of 1200 g and 1800 g respectively [122, 123]. Further studies exploring the effects of Coenzyme Q10 on disease progression in CKD patients and cardiovascular complications in dialysis patients will be valuable.

#### *4.4.5. Miscellaneous antioxidants*

Studies have been conducted to explore the role of other antioxidants, such as vitamin A, selenium, zinc, methionine, alpha-lipoic acid, and curcumin. However, thus far, there is no strong evidence to support their routine use in clinical practice.

### **5. Future directions**

*4.4.2. Vitamin C*

52 Novel Prospects in Oxidative and Nitrosative Stress

*4.4.3. L-carnitine*

to CKD patients.

*4.4.4. Coenzyme Q10*

complications in dialysis patients will be valuable.

strong evidence to support their routine use in clinical practice.

*4.4.5. Miscellaneous antioxidants*

Vitamin C exerts its antioxidant properties by acting as an electron donor to free radicals. A number of small studies have found improvements in inflammatory markers such as CRP [116], hsCRP [117], and 8-OHdG [118] in haemodialysis patients. In a meta-analysis of randomized controlled trials examining use of antioxidants in diabetic kidney disease, vitamin C reduced albuminuria in some studies but had no effect on GFR [111]. All the studies were small (n = 14–29) with short durations of follow up (4 weeks to 12 months) and generally were of suboptimal methodologic quality. Consequently, no conclusions can currently be drawn

Carnitine is an endogenous product of amino acid metabolism produced in the liver. It acts as a transporter of long chain fatty acids across the mitochondrial membrane by reversibly substituting the acyl group of Coenzyme A, forming acyl-carnitine. Once the fatty acids have been transported into the mitochondrial matrix, acyl-carnitine dissociates to form L-carnitine again and coenzyme A is regenerated. L-carnitine has been noted to reduce the oxidative stress through increased glutathione levels, increased GSH-Px activity, and a decreased MDA levels. In a meta-analysis by Chen et al. [119] of 49 RCTs involving 1734 haemodialysis patients, L-carnitine was found to improve CRP and LDL levels. However, in another meta-analysis by Yang et al. [120] involving 25 RCTs, contradictory evidence was found in that L-carnitine did not appreciably alter inflammation, oxidative stress, hyperlipidaemia or quality of life. Currently, there is insufficient evidence to support L-carnitine administration

Coenzyme Q10 is a ubiquinone which contains one quinine group and 10 isoprenyl units. It acts as an enzyme co-factor in inner mitochondrial cell membranes protecting against damage from free radicals produced by oxidative phosphorylation. It also stabilizes the cell membranes as an electron and proton carrier and restores vitamin E in its antioxidant form. Early studies in rat models of diabetic nephropathy showed reduced mesangial expansion and tubulointerstitial fibrosis after administration of mitochondrial-targeted coenzyme Q10 [121]. Two recent studies in dialysis patients showed significant reductions in F2-isoprostanes and isofurans at high doses of 1200 g and 1800 g respectively [122, 123]. Further studies exploring the effects of Coenzyme Q10 on disease progression in CKD patients and cardiovascular

Studies have been conducted to explore the role of other antioxidants, such as vitamin A, selenium, zinc, methionine, alpha-lipoic acid, and curcumin. However, thus far, there is no

regarding the safety and efficacy of vitamin C therapy in patients with CKD.

CKD constitutes a state of increased oxidative stress and systemic inflammation. These processes are pathogenetically interrelated and there is increasing evidence that they may contribute to CKD progression and a disproportionately increased cardiovascular risk through the promotion of endothelial dysfunction, atherosclerosis and vascular calcification. The various mechanisms of action of this increased oxidative stress are increasingly being elucidated. Interventions to reduce oxidative stress and inflammation in these patients present novel, and potentially effective approaches to add to the currently available traditional risk-modification strategies. Due to the complex interrelation between reactive oxygen species and inflammatory markers, it is possible that simultaneous, multiple targeted approaches may be required to effectively address the pathological changes in CKD and its associated cardiovascular risk. Larger trials with meaningful clinical outcomes and longer follow up are required to further evaluate such potential therapies.

### **Author details**


3 Australasian Kidney Trials Network, School of Medicine, University of Queensland, Brisbane, Queensland, Australia


### **References**


[3] Thomas B, Matsushita K, Abate KH, Al-Aly Z, Ärnlöv J, Asayama K, Atkins R, Badawi A, Ballew SH, Banerjee A, Barregård L. Global cardiovascular and renal outcomes of reduced. Journal of the American Society of Nephrology. Jul 1, 2017;**28**(7):2167-2179

[15] Nafar M, Sahraei Z, Salamzadeh J, Samavat S, Vaziri ND. Oxidative stress in kidney transplantation: Causes, consequences, and potential treatment. Iranian Journal of Kidney

The Role of Oxidative Stress and Systemic Inflammation in Kidney Disease and Its Associated…

http://dx.doi.org/10.5772/intechopen.73239

55

[16] Rossi M, Campbell KL, Johnson DW. Indoxyl sulphate and p-cresyl sulphate: Thera-

[17] Mahmoodpoor F, Saadat YR, Barzegari A, Ardalan M, Vahed SZ. The impact of gut microbiota on kidney function and pathogenesis. Biomedicine & Pharmacotherapy. Sep

[18] Briskey D, Tucker P, Johnson DW, Coombes JS. The role of the gastrointestinal tract and microbiota on uremic toxins and chronic kidney disease development. Clinical and

[19] Vaziri ND, Zhao YY, Pahl MV. Altered intestinal microbial flora and impaired epithelial barrier structure and function in CKD: The nature, mechanisms, consequences and potential treatment. Nephrology Dialysis Transplantation. Apr 16, 2015;**31**(5):737-746 [20] Lau WL, Kalantar-Zadeh K, Vaziri ND. The gut as a source of inflammation in chronic

[21] Chen Z, Zhu S, Targeting XG. ut microbiota: A potential promising therapy for diabetic kidney disease. American Journal of Translational Research. 2016;**8**(10):4009

[22] Mahmoodnia L, Aghadavod E, Beigrezaei S, Rafieian-Kopaei M. An update on diabetic kidney disease, oxidative stress and antioxidant agents. Journal of Renal Injury

[23] Wu CC, Chen JS, Wu WM, Liao TN, Chu P, Lin SH, Chuang CH, Lin YF. Myeloperoxidase serves as a marker of oxidative stress during single haemodialysis session using two different biocompatible dialysis membranes. Nephrology Dialysis Transplantation. Apr 5,

[24] Yang CC, Hsu SP, Wu MS, Hsu SM, Chien CT. Effects of vitamin C infusion and vitamin E-coated membrane on hemodialysis-induced oxidative stress. Kidney International.

[25] Susantitaphong P, Riella C, Jaber BL. Effect of ultrapure dialysate on markers of inflammation, oxidative stress, nutrition and anemia parameters: A meta-analysis. Nephrology

[26] Morena M, Cristol JP, Bosc JY, Tetta C, Forret G, Leger CL, Delcourt C, Papoz L, Descomps B, Canaud B. Convective and diffusive losses of vitamin C during haemodiafiltration session: A contributive factor to oxidative stress in haemodialysis patients. Nephrology

[27] Becker BN, Himmelfarb J, Henrich WL, Reassessing HRM. he cardiac risk profile in chronic hemodialysis patients: A hypothesis on the role of oxidant stress and other nontraditional cardiac risk factors. Journal of the American Society of Nephrology. Mar 1,

peutically modifiable nephrovascular toxins. OA Nephrology. 2013;**1**(2):13

Diseases. Nov 1, 2011;**5**(6):357

Experimental Nephrology. Feb 1, 2017;**21**(1):7-15

kidney disease. Nephron. 2015;**130**(2):92-98

Dialysis Transplantation. Jan 4, 2013;**28**(2):438-446

Dialysis Transplantation. Mar 1, 2002;**17**(3):422-427

Prevention. 2017;**6**(2):153

2005;**20**(6):1134-1139

Feb 2, 2006;**69**(4):706-714

1997;**8**(3):475-486

1, 2017;**93**:412-419


[15] Nafar M, Sahraei Z, Salamzadeh J, Samavat S, Vaziri ND. Oxidative stress in kidney transplantation: Causes, consequences, and potential treatment. Iranian Journal of Kidney Diseases. Nov 1, 2011;**5**(6):357

[3] Thomas B, Matsushita K, Abate KH, Al-Aly Z, Ärnlöv J, Asayama K, Atkins R, Badawi A, Ballew SH, Banerjee A, Barregård L. Global cardiovascular and renal outcomes of reduced. Journal of the American Society of Nephrology. Jul 1, 2017;**28**(7):2167-2179 [4] Feigin V. Global, regional, and national life expectancy, all-cause mortality, and causespecific mortality for 249 causes of death, 1980-2015: A systematic analysis for the global

[5] Tonelli M, Muntner P, Lloyd A, Manns BJ, Klarenbach S, Pannu N, James MT, Hemmelgarn BR, Alberta Kidney Disease Network. Risk of coronary events in people with chronic kidney disease compared with those with diabetes: A population-level cohort

[6] Kennedy R, Case C, Fathi R, Johnson D, Isbel N, Marwick TH. Does renal failure cause an atherosclerotic milieu in patients with end-stage renal disease? The American Journal

[7] Galli F, Varga Z, Balla J, Ferraro B, Canestrari F, Floridi A, Kakuk G, Buoncristiani U, Vitamin E. ipid profile, and peroxidation in hemodialysis patients. Kidney International.

[8] Tonelli M, Wiebe N, Thompson S, Kinniburgh D, Klarenbach SW, Walsh M, Bello AK, Faruque L, Field C, Manns BJ, Trace HBR. lement supplementation in hemodialysis

[9] Yilmaz MI, Saglam M, Caglar K, Cakir E, Sonmez A, Ozgurtas T, Aydin A, Eyileten T, Ozcan O, Acikel C, Tasar M. The determinants of endothelial dysfunction in CKD: Oxidative stress and asymmetric dimethylarginine. American Journal of Kidney Diseases.

[10] Miranda-Díaz AG, Pazarín-Villaseñor L, Yanowsky-Escatell FG, Andrade-Sierra J. Oxidative stress in diabetic nephropathy with early chronic kidney disease. Journal of

[11] Wanner C, Zimmermann J, Schwedler S, Metzger T. Inflammation and cardiovascular

[12] Barreto DV, Barreto FC, Liabeuf S, Temmar M, Lemke HD, Tribouilloy C, Choukroun G, Vanholder R, Massy ZA, European Uremic Toxin Work Group (EUTox). Plasma interleukin-6 is independently associated with mortality in both hemodialysis and pre-dialysis patients with chronic kidney disease. Kidney International. Mar 2, 2010;**77**(6):550-556

[13] Fortuño A, Beloqui O, San José G, Moreno MU, Zalba G, Díez J. Increased phagocytic nicotinamide adenine dinucleotide phosphate oxidase-dependent superoxide production in patients with early chronic kidney disease. Kidney International. Dec 31, 2005;

[14] Kumar S, Sharma U, Sharma A, Kenwar DB, Singh S, Prasad R, Minz M. Evaluation of oxidant and antioxidant status in living donor renal allograft transplant recipients.

risk in dialysis patients. Kidney International. May 31, 2002;**61**:S99-102

Diabetes Research. Jul 20, 2016;**2016**:1-7. Article ID: 7047238

Molecular and Cellular Biochemistry. Feb 1, 2016;**413**(1-2):1-8

patients: A randomized controlled trial. BMC Nephrology. Apr 11, 2015;**16**(1):52

burden of disease study 2015. The Lancet. 2016;**388**(10053):1459-1544

study. The Lancet. Sep 7, 2012;**380**(9844):807-814

of Medicine. Feb 15, 2001;**110**(3):198-204

Feb 28, 2001;**59**:S148-S154

54 Novel Prospects in Oxidative and Nitrosative Stress

Jan 31, 2006;**47**(1):42-50

**68**:S71-S75


[28] Maggi E, Bellazzi R, Falaschi F, Frattoni A, Perani G, Finardi G, Gazo A, Nai M, Romanini D, Bellomo G. Enhanced LDL oxidation in uremic patients: An additional mechanism for accelerated atherosclerosis? Kidney International. Mar 1, 1994;**45**(3):876-883

[40] Putri AY, Thaha M. Role of oxidative stress on chronic kidney disease progression. Acta

The Role of Oxidative Stress and Systemic Inflammation in Kidney Disease and Its Associated…

http://dx.doi.org/10.5772/intechopen.73239

57

[41] Arici M, Walls J. End-stage renal disease, atherosclerosis, and cardiovascular mortality: Is C-reactive protein the missing link? Kidney International. Feb 28, 2001;**59**(2):407-414

[42] Gupta J, Mitra N, Kanetsky PA, Devaney J, Wing MR, Reilly M, Shah VO, Balakrishnan VS, Guzman NJ, Girndt M, Periera BG. Association between albuminuria, kidney function, and inflammatory biomarker profile in CKD in CRIC. Clinical Journal of the

[43] Cohen SD, Phillips TM, Khetpal P, Kimmel PL. Cytokine patterns and survival in haemodialysis patients. Nephrology Dialysis Transplantation. Dec 11, 2009;**25**(4):1239-1243

[44] Pupim LB, Himmelfarb J, McMonagle E, Shyr Y, Ikizler TA. Influence of initiation of maintenance hemodialysis on biomarkers of inflammation and oxidative stress. Kidney

[45] Akchurin M, Kaskel F. Update on inflammation in chronic kidney disease. Blood Puri-

[46] De Smet R, David F, Sandra P, Van Kaer J, Lesaffer G, Dhondt A, Lameire N, Vanholder R. A sensitive HPLC method for the quantification of free and total p-cresol in patients

[47] Sharma P, Dietrich T, Ferro CJ, Cockwell P, Chapple IL. Association between periodontitis and mortality in stages 3-5 chronic kidney disease: NHANES III and linked mortality

[48] Sitter T, Bergner A, Schiffl H. Dialysate related cytokine induction and response to recombinant human erythropoietin in haemodialysis patients. Nephrology Dialysis

[49] Kaysen GA, Dubin JA, Müller HG, Rosales LM, Levin NW. The acute-phase response varies with time and predicts serum albumin levels in hemodialysis patients. Kidney

[50] Schindler R, Boenisch O, Fischer C, Frei U. Effect of the hemodialysis membrane on the

[51] Libetta C, De Nicola L, Rampino T, De Simone W, Memoli B. Inflammatory effects of peritoneal dialysis: Evidence of systemic monocyte activation. Kidney International. Feb

[52] Dobre M, Yang W, Chen J, Drawz P, Hamm LL, Horwitz E, Hostetter T, Jaar B, Lora CM, Nessel L, Ojo A. Association of serum bicarbonate with risk of renal and cardiovascular outcomes in CKD: A report from the chronic renal insufficiency cohort (CRIC) study.

[53] Zahed NS, Chehrazi S. The evaluation of the relationship between serum levels of Interleukin-6 and Interleukin-10 and metabolic acidosis in hemodialysis patients. Saudi

inflammatory reaction in vivo. Clinical Nephrology. Jun 2000;**53**(6):452-459

American Journal of Kidney Diseases. Oct 31, 2013;**62**(4):670-678

Journal of Kidney Diseases and Transplantation. Jan 1, 2017;**28**(1):23

with chronic renal failure. Clinica Chimica Acta. Nov 1, 1998;**278**(1):1-21

study. Journal of Clinical Periodontology. Feb 1, 2016;**43**(2):104-113

Medica Indonesiana. May 16, 2016;**46**(3):244-252

International. Jun 30, 2004;**65**(6):2371-2379

Transplantation. Aug 1, 2000;**15**(8):1207-1211

International. Jul 31, 2000;**58**(1):346-352

1, 1996;**49**(2):506-511

fication. 2015;**39**(1-3):84-92

American Society of Nephrology. Dec 1, 2012;**7**(12):1938-1946


[40] Putri AY, Thaha M. Role of oxidative stress on chronic kidney disease progression. Acta Medica Indonesiana. May 16, 2016;**46**(3):244-252

[28] Maggi E, Bellazzi R, Falaschi F, Frattoni A, Perani G, Finardi G, Gazo A, Nai M, Romanini D, Bellomo G. Enhanced LDL oxidation in uremic patients: An additional mechanism

[29] Valgimigli M, Merli E, Malagutti P, Soukhomovskaia O, Cicchitelli G, Macrı̀ G, Ferrari R. Endothelial dysfunction in acute and chronic coronary syndromes: Evidence for a pathogenetic role of oxidative stress. Archives of Biochemistry and Biophysics. Dec 15,

[30] Gosmanova EO, Le NA. Cardiovascular complications in CKD patients: Role of oxidative stress. Cardiology Research and Practice. Jan 2, 2011;**2011**:1-8. Article ID: 156326 [31] Zalba G, Beloqui O, San José G, Moreno MU, Fortuño A, Díez J. NADPH oxidase–dependent superoxide production is associated with carotid intima-media thickness in subjects free of clinical atherosclerotic disease. Arteriosclerosis, Thrombosis, and Vascular

[32] Meenakshi Sundaram SP, Nagarajan S, Manjula Devi AJ. Chronic kidney disease—Effect of oxidative stress. Chinese Journal of Biology. Jan 21, 2014;**2014**:1-6. Article ID: 216210

[33] Drüeke T, Witko-Sarsat V, Massy Z, Descamps-Latscha B, Guerin AP, Marchais SJ, Gausson V, London GM. Iron therapy, advanced oxidation protein products, and carotid artery intima-media thickness in end-stage renal disease. Circulation. Oct 22, 2002;

[34] Ari E, Kaya Y, Demir H, Cebi A, Alp HH, Bakan E, Odabasi D, Keskin S. Oxidative DNA damage correlates with carotid artery atherosclerosis in hemodialysis patients.

[35] Stopper H, Boullay F, Heidland A, Bahner U. Comet-assay analysis identifies genomic damage in lymphocytes of uremic patients. American Journal of Kidney Diseases. Aug

[36] Small DM, Bennett NC, Roy S, Gabrielli BG, Johnson DW, Gobe GC. Oxidative stress and cell senescence combine to cause maximal renal tubular epithelial cell dysfunction and loss in an in vitro model of kidney disease. Nephron Experimental Nephrology.

[37] Rossi M, Campbell K, Johnson D, Stanton T, Pascoe E, Hawley C, Dimeski G, McWhinney B, Ungerer J, Isbel N. Uraemic toxins and cardiovascular disease across the chronic kidney disease spectrum: An observational study. Nutrition, Metabolism and Cardiovascular

[38] Rossi M, Campbell KL, Johnson DW, Stanton T, Vesey DA, Coombes JS, Weston KS, Hawley CM, McWhinney BC, Ungerer JP, Isbel N. Protein-bound uremic toxins, inflammation and oxidative stress: A cross-sectional study in stage 3-4 chronic kidney disease.

[39] Mafra D, Lobo JC, Barros AF, Koppe L, Vaziri ND, Fouque D. Role of altered intestinal microbiota in systemic inflammation and cardiovascular disease in chronic kidney dis-

for accelerated atherosclerosis? Kidney International. Mar 1, 1994;**45**(3):876-883

2003;**420**(2):255-261

56 Novel Prospects in Oxidative and Nitrosative Stress

**106**(17):2212-2217

31, 2001;**38**(2):296-301

2012;**122**(3-4):123-130

Diseases. Sep 30, 2014;**24**(9):1035-1042

Archives of Medical Research. May 31, 2014;**45**(4):309-317

ease. Future Microbiology. Mar 2014;**9**(3):399-410

Biology. Jul 1, 2005;**25**(7):1452-1457

Hemodialysis International. Oct 1, 2011;**15**(4):453-459


[54] Ori Y, Bergman M, Bessler H, Zingerman B, Levy-Drummer RS, Gafter U, Salman H. Cytokine secretion and markers of inflammation in relation to acidosis among chronic hemodialysis patients. Blood Purification. 2013;**35**(1-3):181-186

[66] Hassan MO, Duarte R, Dix-Peek T, Vachiat A, Naidoo S, Dickens C, Grinter S, Manga P, Naicker S. Correlation between volume overload, chronic inflammation, and left ventricular dysfunction in chronic kidney disease patients. Clinical Nephrology. 2016;**86**(7):131

The Role of Oxidative Stress and Systemic Inflammation in Kidney Disease and Its Associated…

http://dx.doi.org/10.5772/intechopen.73239

59

[67] Mitsides N, Cornelis T, Broers NJ, Diederen NM, Brenchley P, van der Sande FM, Schalkwijk CG, Kooman JP, Mitra S. Extracellular overhydration linked with endothelial dysfunction in the context of inflammation in haemodialysis dependent chronic kidney

[68] Ortega O, Gallar P, Muñoz M, Rodríguez I, Carreño A, Ortiz M, Molina A, Oliet A, Lozano L, Vigil A. Association between C-reactive protein levels and N-terminal pro-B-type natriuretic peptide in pre-dialysis patients. Nephron Clinical Practice. 2004;

[69] Telini LS, de Carvalho Beduschi G, Caramori JC, Castro JH, Martin LC, Barretti P. Effect of dietary sodium restriction on body water, blood pressure, and inflammation in hemodialysis patients: A prospective randomized controlled study. International Urology

[70] Campbell KL, Johnson DW, Bauer JD, Hawley CM, Isbel NM, Stowasser M, Whitehead JP, Dimeski G, McMahon EA. andomized trial of sodium-restriction on kidney function, fluid volume and adipokines in CKD patients. BMC Nephrology. Apr 4, 2014;**15**(1):57

[71] Danesi F, Ferguson LR. Could pomegranate juice help in the control of inflammatory

[72] Shema-Didi L, Sela S, Ore L, Shapiro G, Geron R, Moshe G, Kristal B. One year of pomegranate juice intake decreases oxidative stress, inflammation, and incidence of infections in hemodialysis patients: A randomized placebo-controlled trial. Free Radical Biology

[73] McGraw NJ, Krul ES, Grunz-Borgmann E, Parrish AR. Soy-based renoprotection. World

[74] Rossi M, Johnson DW, Morrison M, Pascoe EM, Coombes JS, Forbes JM, Szeto CC, McWhinney BC, Ungerer JP, Campbell KL. Synbiotics easing renal failure by improving gut microbiology (SYNERGY): A randomized trial. Clinical Journal of the American

[75] Gould DW, Graham-Brown MP, Watson EL, Viana JL, Smith AC. Physiological benefits of exercise in pre-dialysis chronic kidney disease. Nephrology. Sep 1, 2014;**19**(9):519-527

[76] Viana JL, Kosmadakis GC, Watson EL, Bevington A, Feehally J, Bishop NC, Smith AC. Evidence for anti-inflammatory effects of exercise in CKD. Journal of the American

[77] Castaneda C, Gordon PL, Parker RC, Uhlin KL, Roubenoff R, Levey AS. Resistance training to reduce the malnutrition-inflammation complex syndrome of chronic kidney dis-

ease. American Journal of Kidney Diseases. Apr 30, 2004;**43**(4):607-616

disease. PLoS One. Aug 22, 2017;**12**(8):e0183281

and Nephrology. Jan 1, 2014;**46**(1):91-97

diseases? Nutrients. Aug 30, 2017;**9**(9):958

and Medicine. Jul 15, 2012;**53**(2):297-304

Journal of Nephrology. May 6, 2016;**5**(3):233

Society of Nephrology. Feb 5, 2016;**11**(2):223-231

Society of Nephrology. Sep 1, 2014;**25**(9):2121-2130

**97**(4):c125-c130


[66] Hassan MO, Duarte R, Dix-Peek T, Vachiat A, Naidoo S, Dickens C, Grinter S, Manga P, Naicker S. Correlation between volume overload, chronic inflammation, and left ventricular dysfunction in chronic kidney disease patients. Clinical Nephrology. 2016;**86**(7):131

[54] Ori Y, Bergman M, Bessler H, Zingerman B, Levy-Drummer RS, Gafter U, Salman H. Cytokine secretion and markers of inflammation in relation to acidosis among chronic

[55] Pedraza-Chaverri J, Sánchez-Lozada LG, Osorio-Alonso H, Tapia E, Scholze A. New pathogenic concepts and therapeutic approaches to oxidative stress in chronic kidney disease. Oxidative Medicine and Cellular Longevity. Jun 27, 2016;**2016**:1-21. Article ID:

[56] Stenvinkel P, Heimbürger O, Paultre F, Diczfalusy U, Wang T, Berglund L, Jogestrand T. Strong association between malnutrition, inflammation, and atherosclerosis in chronic

[57] Yeun JY, Levine RA, Mantadilok V, Kaysen GA. C-reactive protein predicts all-cause and cardiovascular mortality in hemodialysis patients. American Journal of Kidney

[58] Noh H, Lee SW, Kang SW, Shin SK, Choi KH, Lee HY, Han DS. Serum C-reactive protein: A predictor of mortality in continuous ambulatory peritoneal dialysis patients.

[59] Zimmermann J, Herrlinger S, Pruy A, Metzger T, Wanner C. Inflammation enhances cardiovascular risk and mortality in hemodialysis patients. Kidney International. Feb

[60] Antunovic T, Stefanovic A, Gligorovic Barhanovic N, Miljkovic M, Radunovic D, Ivanisevic J, Prelevic V, Bulatovic N, Ratkovic M, Stojanov M. Prooxidant–antioxidant balance, hsTnI and hsCRP: Mortality prediction in haemodialysis patients, two-year

[61] Rossi M, Johnson DW, Xu H, Carrero JJ, Pascoe E, French C, Campbell KL. Dietary protein-fiber ratio associates with circulating levels of indoxyl sulfate and p-cresyl sulfate in chronic kidney disease patients. Nutrition, Metabolism and Cardiovascular Diseases.

[62] Sirich TL, Plummer NS, Gardner CD, Hostetter TH, Meyer TW. Effect of increasing dietary fiber on plasma levels of colon-derived solutes in hemodialysis patients. Clinical

[63] Meijers BK, De Preter V, Verbeke K, Vanrenterghem Y, Evenepoel P. p-Cresyl sulfate serum concentrations in haemodialysis patients are reduced by the prebiotic oligofructose-enriched inulin. Nephrology Dialysis Transplantation. Aug 19, 2009;**25**(1):219-224

[64] Xu H, Huang X, Risérus U, et al. Dietary fiber, kidney function, inflammation, and mortality risk. Clinical Journal of the American Society of Nephrology: CJASN.

[65] Chiavaroli L, Mirrahimi A, Sievenpiper JL, Jenkins DJ, Darling PB. Dietary fiber effects in chronic kidney disease: A systematic review and meta-analysis of controlled feeding

trials. European Journal of Clinical Nutrition. Jul 1, 2015;**69**(7):761-768

Journal of the American Society of Nephrology. Sep 5, 2014;**9**(9):1603-1610

hemodialysis patients. Blood Purification. 2013;**35**(1-3):181-186

renal failure. Kidney International. May 31, 1999;**55**(5):1899-1911

Peritoneal Dialysis International. Jan 1, 1998;**18**(4):387-394

follow-up. Renal Failure. Jan 1, 2017;**39**(1):491-499

Diseases. Mar 31, 2000;**35**(3):469-476

28, 1999;**55**(2):648-658

Sep 30, 2015;**25**(9):860-865

2014;**9**(12):2104-2110

6043601

58 Novel Prospects in Oxidative and Nitrosative Stress


[78] Hakim RM. Clinical implications of hemodialysis membrane biocompatibility. Kidney International. Sep 1, 1993;**44**(3):484-494

[91] Goicoechea M, de Vinuesa SG, Verdalles U, Ruiz-Caro C, Ampuero J, Rincón A, Arroyo D, Luño J. Effect of allopurinol in chronic kidney disease progression and cardiovascular risk. Clinical Journal of the American Society of Nephrology. Aug 1, 2010;**5**(8):1388-1393

The Role of Oxidative Stress and Systemic Inflammation in Kidney Disease and Its Associated…

http://dx.doi.org/10.5772/intechopen.73239

61

[92] Siu YP, Leung KT, Tong MK, Kwan TH. Use of allopurinol in slowing the progression of renal disease through its ability to lower serum uric acid level. American Journal of

[93] Bose B, Badve SV, Hiremath SS, Boudville N, Brown FG, Cass A, De Zoysa JR, Fassett RG, Faull R, Harris DC, Hawley CM. Effects of uric acid-lowering therapy on renal outcomes: A systematic review and meta-analysis. Nephrology Dialysis Transplantation.

[94] Goicoechea M, de Vinuesa SG, Verdalles U, Verde E, Macias N, Santos A, de Jose AP, Cedeño S, Linares T, Luño J. Allopurinol and progression of CKD and cardiovascular events: Long-term follow-up of a randomized clinical trial. American Journal of Kidney

[95] Ribeiro G, Roehrs M, Bairros A, Moro A, Charão M, Araújo F, Valentini J, Arbo M, Brucker N, Moresco R, Leal M. N-acetylcysteine on oxidative damage in diabetic rats.

[96] Hsu SP, Chiang CK, Yang SY, Chien CT. N-acetylcysteine for the management of anemia and oxidative stress in hemodialysis patients. Nephron Clinical Practice. 2010;

[97] Nascimento MM, Suliman ME, Silva M, Chinaglia T, Marchioro J, Hayashi SY, Riella MC, Lindholm B, Anderstam B. Effect of oral N-acetylcysteine treatment on plasma inflammatory and oxidative stress markers in peritoneal dialysis patients: A placebo-

[98] Renke M, Tylicki L, Rutkowski P, Larczyński W, Aleksandrowicz E, Łysiak-Szydłowska W, Rutkowski B. The effect of N-acetylcysteine on proteinuria and markers of tubular injury in non-diabetic patients with chronic kidney disease. Kidney and Blood Pressure

[99] Peake JM, Gobe GC, Fassett RG, Coombes JS. The effects of dietary fish oil on inflammation, fibrosis and oxidative stress associated with obstructive renal injury in rats.

[100] Hung AM, Booker C, Ellis CD, Siew ED, Graves AJ, Shintani A, Abumrad NN, Himmelfarb J, Ikizler TA. Omega-3 fatty acids inhibit the up-regulation of endothelial chemokines in maintenance hemodialysis patients. Nephrology Dialysis Transplantation.

[101] Inoue T, Okano K, Tsuruta Y, Tsuruta Y, Tsuchiya K, Akiba T, Nitta K. Eicosapentaenoic acid (EPA) decreases the all-cause mortality in hemodialysis patients. Internal Medicine.

Molecular Nutrition & Food Research. Mar 1, 2011;**55**(3):400-410

controlled study. Peritoneal Dialysis International. May 1, 2010;**30**(3):336-342

Kidney Diseases. Jan 31, 2006;**47**(1):51-59

Sep 15, 2013;**29**(2):406-413

**116**(3):c207-c216

Research. 2008;**31**(6):404-410

Sep 9, 2014;**30**(2):266-274

2015;**54**(24):3133-3137

Diseases. Apr 30, 2015;**65**(4):543-549

Drug and Chemical Toxicology. Oct 1, 2011;**34**(4):467-474


[91] Goicoechea M, de Vinuesa SG, Verdalles U, Ruiz-Caro C, Ampuero J, Rincón A, Arroyo D, Luño J. Effect of allopurinol in chronic kidney disease progression and cardiovascular risk. Clinical Journal of the American Society of Nephrology. Aug 1, 2010;**5**(8):1388-1393

[78] Hakim RM. Clinical implications of hemodialysis membrane biocompatibility. Kidney

[79] D'Arrigo G, Baggetta R, Tripepi G, Galli F, Bolignano D. Effects of vitamin E-coated versus conventional membranes in chronic hemodialysis patients: A systematic review and

[80] Den Hoedt CH, Bots ML, Grooteman MP, Van Der Weerd NC, Mazairac AH, Penne EL, Levesque R, Ter Wee PM, Nubé MJ, Blankestijn PJ, Van Den Dorpel MA. Online hemodiafiltration reduces systemic inflammation compared to low-flux hemodialysis. Kidney

[81] Ayus JC, Mizani MR, Achinger SG, Thadhani R, Go AS, Lee S. Effects of short daily versus conventional hemodialysis on left ventricular hypertrophy and inflammatory markers: A prospective, controlled study. Journal of the American Society of Nephrology. Sep

[82] Susantitaphong P, Koulouridis I, Balk EM, Madias NE, Jaber BL. Effect of frequent or extended hemodialysis on cardiovascular parameters: A meta-analysis. American Jour-

[83] Palleschi S, Ghezzi PM, Palladino G, Rossi B, Ganadu M, Casu D, Cossu M, Mattana G, Pinna AM, Contu B, Ghisu T. Vitamins (A, C and E) and oxidative status of hemodialysis patients treated with HFR and HFR-supra. BMC Nephrology. Aug 26, 2016;**17**(1):120 [84] Schulman G, Berl T, Beck GJ, Remuzzi G, Ritz E, Arita K, Kato A, Shimizu M. Randomized placebo-controlled EPPIC trials of AST-120 in CKD. Journal of the American Society of

[85] Perlstein TS, Gumieniak O, Hopkins PN, Murphey LJ, Brown NJ, Williams GH, Hollenberg NK, Fisher ND. Uric acid and the state of the intrarenal renin-angiotensin

[86] Grayson PC, Kim SY, LaValley M, Choi HK. Hyperuricemia and incident hypertension: A systematic review and meta-analysis. Arthritis Care & Research. Jan 2011;**63**(1, 1):

[87] Ramirez-Sandoval JC, Sanchez-Lozada LG, Madero M. Uric acid, vascular stiffness, and chronic kidney disease: Is there a link? Blood Purification. 2017;**43**(1-3):189-195 [88] Short RA, Johnson RJ, Tuttle KR. Uric acid, microalbuminuria and cardiovascular events in high-risk patients. American Journal of Nephrology. 2005;**25**(1):36-44 [89] Madero M, Sarnak MJ, Wang X, Greene T, Beck GJ, Kusek JW, Collins AJ, Levey AS, Menon V. Uric acid and long-term outcomes in CKD. American Journal of Kidney

[90] Petreski T, Bevc S, Ekart R, Hojs R. Hyperuricemia and long-term survival in patients with chronic kidney disease undergoing hemodialysis. Clinical Nephrology.

system in humans. Kidney International. Oct 31, 2004;**66**(4):1465-1470

International. Sep 1, 1993;**44**(3):484-494

60 Novel Prospects in Oxidative and Nitrosative Stress

International. Aug 31, 2014;**86**(2):423-432

Nephrology. Jul 1, 2015;**26**(7):1732-1746

Diseases. May 31, 2009;**53**(5):796-803

2017;**88**(13):s69-72

1, 2005;**16**(9):2778-2788

102-110

meta-analysis. Blood Purification. 2017;**43**(1-3):101-122

nal of Kidney Diseases. May 31, 2012;**59**(5):689-699


[102] Taheri S, Keyvandarian N, Moeinzadeh F, Mortazavi M, Naini AE. The effect of omega-3 fatty acid supplementation on oxidative stress in continuous ambulatory peritoneal dialysis patients. Advanced Biomedical Research. 2014;**3**:1-9. PMCID: PMC4139980

[115] Mann JF, Lonn EV, Yi Q, Gerstein HC, Hoogwerf BJ, Pogue J, Bosch J, Dagenais GR, Yusuf S, Investigators HOPE. Effects of vitamin E on cardiovascular outcomes in people with mild-to-moderate renal insufficiency: Results of the HOPE study. Kidney

The Role of Oxidative Stress and Systemic Inflammation in Kidney Disease and Its Associated…

http://dx.doi.org/10.5772/intechopen.73239

63

[116] Biniaz V, Shermeh MS, Ebadi A, Tayebi A, Einollahi B. Effect of vitamin C supplementation on C-reactive protein levels in patients undergoing hemodialysis: A randomized, double blind, placebo-controlled study. Nephro-Urology Monthly. 2014;**6**(1):e13351:1-6

[117] Zhang K, Li Y, Cheng X, Liu L, Bai W, Guo W. Wu L, Zuo L. cross-over study of influence of oral vitamin C supplementation on inflammatory status in maintenance hemo-

[118] Tarng DC, Liu TY, Huang TP. Protective effect of vitamin C on 8-hydroxy-2′-deoxyguanosine level in peripheral blood lymphocytes of chronic hemodialysis patients.

[119] Chen Y, Abbate M, Tang L, Cai G, Gong Z, Wei R, Zhou J, Chen X. L-Carnitine supplementation for adults with end-stage kidney disease requiring maintenance hemodialysis: A systematic review and meta-analysis. The American Journal of Clinical Nutrition.

[120] Yang SK, Xiao L, Song PA, Xu X, Liu FY, Sun L. Effect of L-carnitine therapy on patients in maintenance hemodialysis: A systematic review and meta-analysis. Journal of Neph-

[121] Chacko BK, Reily C, Srivastava A, Johnson MS, Ye Y, Ulasova E, Agarwal A, Zinn KR, Murphy MP, Kalyanaraman B, Darley-Usmar V. Prevention of diabetic nephropathy in Ins2+/− AkitaJ mice by the mitochondria-targeted therapy MitoQ. Biochemical Journal.

[122] Rivara MB, Yeung CK, Robinson-Cohen C, Phillips BR, Ruzinski J, Rock D, Linke L, Shen DD, Ikizler TA, Himmelfarb J. Effect of coenzyme Q 10 on biomarkers of oxidative stress and cardiac function in Hemodialysis patients: The CoQ 10 biomarker trial.

[123] Yeung CK, Billings FT, Claessens AJ, Roshanravan B, Linke L, Sundell MB, Ahmad S, Shao B, Shen DD, Ikizler TA, Coenzyme HJ. Q 10 dose-escalation study in hemodialysis patients: Safety, tolerability, and effect on oxidative stress. BMC Nephrology. Nov 3,

American Journal of Kidney Diseases. Mar 31, 2017;**69**(3):389-399

dialysis patients. BMC Nephrology. Nov 14, 2013;**14**(1):252

Kidney International. Aug 31, 2004;**66**(2):820-831

Feb 1, 2014;**99**(2):408-422

Nov 15, 2010;**432**(1):9-19

2015;**16**(1):183

rology. Jun 1, 2014;**27**(3):317-329

International. Apr 30, 2004;**65**(4):1375-1380


[115] Mann JF, Lonn EV, Yi Q, Gerstein HC, Hoogwerf BJ, Pogue J, Bosch J, Dagenais GR, Yusuf S, Investigators HOPE. Effects of vitamin E on cardiovascular outcomes in people with mild-to-moderate renal insufficiency: Results of the HOPE study. Kidney International. Apr 30, 2004;**65**(4):1375-1380

[102] Taheri S, Keyvandarian N, Moeinzadeh F, Mortazavi M, Naini AE. The effect of omega-3 fatty acid supplementation on oxidative stress in continuous ambulatory peritoneal dialysis patients. Advanced Biomedical Research. 2014;**3**:1-9. PMCID: PMC4139980 [103] Deike E, Bowden RG, Moreillon JJ, Griggs JO, Wilson RL, Cooke M, Shelmadine BD, Beaujean AA. The effects of fish oil supplementation on markers of inflammation in chronic kidney disease patients. Journal of Renal Nutrition. Nov 30, 2012;**22**(6):572-577

[104] Bouzidi N, Mekki K, Boukaddoum A, Dida N, Kaddous A, Bouchenak M. Effects of omega-3 polyunsaturated fatty-acid supplementation on redox status in chronic renal failure patients with dyslipidemia. Journal of Renal Nutrition. Sep 30, 2010;**20**(5):321-328

[105] Ma Q. Role of nrf2 in oxidative stress and toxicity. Annual Review of Pharmacology

[106] Nezu M, Suzuki N, Yamamoto M. Targeting the KEAP1-NRF2 system to prevent kidney disease progression. American Journal of Nephrology. 2017;**45**(6):473-483

[107] Pergola PE, Raskin P, Toto RD, Meyer CJ, Huff JW, Grossman EB, Krauth M, Ruiz S, Audhya P, Christ-Schmidt H, Wittes J. Bardoxolone methyl and kidney function in CKD with type 2 diabetes. New England Journal of Medicine. Jul 28, 2011;**365**(4):327-336 [108] De Zeeuw D, Akizawa T, Audhya P, Bakris GL, Chin M, Christ-Schmidt H, Goldsberry A, Houser M, Krauth M, Lambers Heerspink HJ, McMurray JJ. Bardoxolone methyl in type 2 diabetes and stage 4 chronic kidney disease. New England Journal of Medicine.

[109] Deng J, Wu Q, Liao Y, Huo D, Yang Z. Effect of statins on chronic inflammation and nutrition status in renal dialysis patients: A systematic review and meta-analysis.

[110] Hung AM, Ellis CD, Shintani A, Booker C, Ikizler TA. IL-1β receptor antagonist reduces inflammation in hemodialysis patients. Journal of the American Society of Nephrology.

[111] Jun M, Venkataraman V, Razavian M, Cooper B, Zoungas S, Ninomiya T, Webster AC, Antioxidants PV. or chronic kidney disease. The Cochrane Library. Jan 1, 2012

[112] Bolignano D, Cernaro V, Gembillo G, Baggetta R, Buemi M, D'Arrigo G. Antioxidant agents for delaying diabetic kidney disease progression: A systematic review and meta-

[113] Huang J, Yi B, Li AM, Zhang H. Effects of vitamin E-coated dialysis membranes on anemia, nutrition and dyslipidemia status in hemodialysis patients: A meta-analysis.

[114] Boaz M, Smetana S, Weinstein T, Matas Z, Gafter U, Iaina A, Knecht A, Weissgarten Y, Brunner D, Fainaru M, Green MS. Secondary prevention with antioxidants of cardiovascular disease in endstage renal disease (SPACE): Randomised placebo-controlled

and Toxicology. Jan 6, 2013;**53**:401-426

62 Novel Prospects in Oxidative and Nitrosative Stress

Dec 26, 2013;**369**(26):2492-2503

Mar 1, 2011;**22**(3):437-442

Nephrology. Aug 1, 2012;**17**(6):545-551

analysis. PLoS One. Jun 1, 2017;**12**(6):e0178699

trial. The Lancet. Oct 7, 2000;**356**(9237):1213-1218

Renal Failure. Mar 16, 2015;**37**(3):398-407


**Chapter 3**

**Provisional chapter**

**Role of Oxidative/Nitrosative Stress in Diarrhea and**

**Role of Oxidative/Nitrosative Stress in Diarrhea and** 

Oxidative/nitrosative stress, a pervasive condition of increased amounts of reactive and nitrogen species, is responsible for a variety of degenerative processes in some human diseases such as gastrointestinal affections. Diarrhea is one such infection that has long been recognized as one of the most important health problems in developing countries. Constipation is a delay or difficulty in evacuating the stool. In this respect, several studies were performed and have shown that the diarrhea pathophysiology and constipation were accompanied by accumulation of biomarkers of oxidative/nitrosative stress as well as the depletion of antioxidant system. In this chapter, we discuss about the recent advances that propose a major role of oxidative/nitrosative stress on diarrhea pathogen-

**Keywords:** oxidative/nitrosative stress, reactive and nitrogen species, diarrhea

Reactive oxygen species (ROS)/reactive nitrogen species (RNS) are produced as the by-products of the normal metabolic mechanism in all aerobic organisms [1]. The augmentation of oxidative/nitrosative stress normally describes a situation in which cellular antioxidant capacities are incapable to scavenge the ROS and RNS engendered as a result of massive generation of ROS/ RNS, loss of antioxidant defenses, or both. The ROS/RNS cause disruptions in the cellular macromolecules such as the oxidative degradation of lipids, DNA lesion and proteins alteration [2].

> © 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use,

distribution, and reproduction in any medium, provided the original work is properly cited.

DOI: 10.5772/intechopen.74788

Kaïs Rtibi, Hichem Sebai and Lamjed Marzouki

Kaïs Rtibi, Hichem Sebai and Lamjed Marzouki

Additional information is available at the end of the chapter

Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.74788

**Constipation**

**Abstract**

esis and constipation.

pathogenesis

**1. Introduction**

**Constipation**

#### **Role of Oxidative/Nitrosative Stress in Diarrhea and Constipation Role of Oxidative/Nitrosative Stress in Diarrhea and Constipation**

DOI: 10.5772/intechopen.74788

Kaïs Rtibi, Hichem Sebai and Lamjed Marzouki Kaïs Rtibi, Hichem Sebai and Lamjed Marzouki

Additional information is available at the end of the chapter Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.74788

#### **Abstract**

Oxidative/nitrosative stress, a pervasive condition of increased amounts of reactive and nitrogen species, is responsible for a variety of degenerative processes in some human diseases such as gastrointestinal affections. Diarrhea is one such infection that has long been recognized as one of the most important health problems in developing countries. Constipation is a delay or difficulty in evacuating the stool. In this respect, several studies were performed and have shown that the diarrhea pathophysiology and constipation were accompanied by accumulation of biomarkers of oxidative/nitrosative stress as well as the depletion of antioxidant system. In this chapter, we discuss about the recent advances that propose a major role of oxidative/nitrosative stress on diarrhea pathogenesis and constipation.

**Keywords:** oxidative/nitrosative stress, reactive and nitrogen species, diarrhea pathogenesis

### **1. Introduction**

Reactive oxygen species (ROS)/reactive nitrogen species (RNS) are produced as the by-products of the normal metabolic mechanism in all aerobic organisms [1]. The augmentation of oxidative/nitrosative stress normally describes a situation in which cellular antioxidant capacities are incapable to scavenge the ROS and RNS engendered as a result of massive generation of ROS/ RNS, loss of antioxidant defenses, or both. The ROS/RNS cause disruptions in the cellular macromolecules such as the oxidative degradation of lipids, DNA lesion and proteins alteration [2].

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Constipation is defined as infrequent or difficult evacuation of feces leading to water absorption, hardening of stool in colon, and excessive straining. This gastrointestinal disorder is a risk factor of colorectal cancer [3].

Diarrhea is usually a result of gastrointestinal infection, which can be induced by various microorganisms such as viruses, bacteria, and parasites. Despite different pathophysiological changes in different types of diarrheas, there are four major mechanisms responsible for this gastrointestinal disruption in electrolyte and water exchange, that is, elevated luminal osmolarity, increased electrolyte secretion, decreased electrolyte absorption, and accelerated intestinal motility [4].

Therefore, the objective of this chapter is to discuss, based on the literature, the contribution of oxidative/nitrosative stress in gastrointestinal disorders such as constipation and diarrhea.

### **2. Oxidative/nitrosative stress and gastrointestinal disorders**

Alterations in the digestive tract such as constipation and diarrhea are caused by many external agents and factors. These disturbances are accompanied by the installation of oxidative/nitrosative stress, which can cause various disruptions in gastrointestinal intestinal function (**Figure 1**).

#### **2.1. Oxidative stress and diarrhea**

Many literature studies suggest the involvement of oxidative stress in the aggravation of diverse perturbations, including gastrointestinal infectious diseases produced by pathogens. These results indicate that Rotavirus induces a generation of ROS and deficiency in the reduced glutathione (GSH)/oxidized glutathione (GSSG) ratio [5]. Added to that, it has been shown also that diarrhea induced by bacterial infections was combined with an oxidative injury. Indeed, during the steps of salmonellosis, ROS are also generated which provokes a depletion of glutathione in intestinal epithelial cells [6].

H2 O2 to H2

O and O2

**2.2. Oxidative stress and constipation**

children and depletion of antioxidant enzyme activities [16].

duction of ROS [12, 13].

and constipation pathogenesis.

[11]. In addition, glutathione peroxidase has a high affinity for hydrogen

Role of Oxidative/Nitrosative Stress in Diarrhea and Constipation

http://dx.doi.org/10.5772/intechopen.74788

67

peroxide; it therefore allows for the removal of hydrogen peroxide, even when present at a low concentration. In this respect, numerous studies have reported that castor oil-induced diarrhea causes a depletion of antioxidant activities of SOD, CAT, and GPx, which explains the overpro-

**Figure 1.** Contribution of oxidative/nitrosative stress in gastrointestinal disorders including diarrhea pathophysiology

On the other hand, several studies have reported an increased oxidative stress and imbalance in antioxidant enzymes following the administration of antineoplastic agents that induced the constipation. In this respect, the use of vinblastine was provoked by the installation of constipation which is associated with a disturbance in the balance between the production of reactive oxygen species (free radicals) and antioxidant defenses in intestinal mucosal barrier. This mechanism was evaluated by lipid peroxidation, protein oxidation, and damaging actions on sulfhydryl groups. Disorders in the normal redox state of cells can induce toxic activities through the generation of free radical reactive oxygen species that induce cell injury and alter these cellular macromolecules [14]. These obtained results are in agreement with those found by Li et al. [15] who revealed that the level of MDA augmented in constipated rats. In addition, other previous reports indicate that chronic constipation can cause potential oxidative stress in

Other researches have shown the implication of oxidative stress in castor oil-induced diarrhea. Therefore, recent studies have shown that acute administration of castor increased the formation of malondialdehyde (MDA) in the gastrointestinal tract mucosa indicating an increase in lipid peroxidation. This process presents a possible mechanism of tissue alteration by oxygen reactive derivatives [7, 8]. Furthermore, current findings showed that intestinal hypersecretion was also accompanied by H2 O2 generation in mucosal intestine. H<sup>2</sup> O2 can lead to the formation of toxic (•OH) which oxidizes important cellular components and induces the depletion of glutathione. Oxidative damage of lipids provokes a membrane fluidity alteration, disruption in ion transport, loss of membrane integrity, and finally, cellular function disturbance [9].

Other studies reported that diarrhea was able to induce deleterious effects on the sulfhydryl (─SH) group and generation of protein carbonyls. These effects can be explained by the proteins oxidation process, which leads to the dysfunction of many enzymes [10].

Enzymatic antioxidants including superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) have an important role in the prevention of oxidative damage by reactive oxygen species. SOD plays a crucial action in dismutation of superoxide radicals to H<sup>2</sup> O and oxygen. On the other hand, catalase protects the cells from toxic effects of ROS by transforming

**Figure 1.** Contribution of oxidative/nitrosative stress in gastrointestinal disorders including diarrhea pathophysiology and constipation pathogenesis.

H2 O2 to H2 O and O2 [11]. In addition, glutathione peroxidase has a high affinity for hydrogen peroxide; it therefore allows for the removal of hydrogen peroxide, even when present at a low concentration. In this respect, numerous studies have reported that castor oil-induced diarrhea causes a depletion of antioxidant activities of SOD, CAT, and GPx, which explains the overproduction of ROS [12, 13].

#### **2.2. Oxidative stress and constipation**

Constipation is defined as infrequent or difficult evacuation of feces leading to water absorption, hardening of stool in colon, and excessive straining. This gastrointestinal disorder is a

Diarrhea is usually a result of gastrointestinal infection, which can be induced by various microorganisms such as viruses, bacteria, and parasites. Despite different pathophysiological changes in different types of diarrheas, there are four major mechanisms responsible for this gastrointestinal disruption in electrolyte and water exchange, that is, elevated luminal osmolarity, increased electrolyte secretion, decreased electrolyte absorption, and accelerated intestinal motility [4].

Therefore, the objective of this chapter is to discuss, based on the literature, the contribution of oxidative/nitrosative stress in gastrointestinal disorders such as constipation and diarrhea.

Alterations in the digestive tract such as constipation and diarrhea are caused by many external agents and factors. These disturbances are accompanied by the installation of oxidative/nitrosative stress, which can cause various disruptions in gastrointestinal intestinal function (**Figure 1**).

Many literature studies suggest the involvement of oxidative stress in the aggravation of diverse perturbations, including gastrointestinal infectious diseases produced by pathogens. These results indicate that Rotavirus induces a generation of ROS and deficiency in the reduced glutathione (GSH)/oxidized glutathione (GSSG) ratio [5]. Added to that, it has been shown also that diarrhea induced by bacterial infections was combined with an oxidative injury. Indeed, during the steps of salmonellosis, ROS are also generated which provokes a

Other researches have shown the implication of oxidative stress in castor oil-induced diarrhea. Therefore, recent studies have shown that acute administration of castor increased the formation of malondialdehyde (MDA) in the gastrointestinal tract mucosa indicating an increase in lipid peroxidation. This process presents a possible mechanism of tissue alteration by oxygen reactive derivatives [7, 8]. Furthermore, current findings showed that intestinal hypersecretion

generation in mucosal intestine. H<sup>2</sup>

of toxic (•OH) which oxidizes important cellular components and induces the depletion of glutathione. Oxidative damage of lipids provokes a membrane fluidity alteration, disruption in ion transport, loss of membrane integrity, and finally, cellular function disturbance [9].

Other studies reported that diarrhea was able to induce deleterious effects on the sulfhydryl (─SH) group and generation of protein carbonyls. These effects can be explained by the pro-

Enzymatic antioxidants including superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) have an important role in the prevention of oxidative damage by reactive

oxygen. On the other hand, catalase protects the cells from toxic effects of ROS by transforming

oxygen species. SOD plays a crucial action in dismutation of superoxide radicals to H<sup>2</sup>

teins oxidation process, which leads to the dysfunction of many enzymes [10].

O2

can lead to the formation

O and

**2. Oxidative/nitrosative stress and gastrointestinal disorders**

risk factor of colorectal cancer [3].

66 Novel Prospects in Oxidative and Nitrosative Stress

**2.1. Oxidative stress and diarrhea**

was also accompanied by H2

depletion of glutathione in intestinal epithelial cells [6].

O2

On the other hand, several studies have reported an increased oxidative stress and imbalance in antioxidant enzymes following the administration of antineoplastic agents that induced the constipation. In this respect, the use of vinblastine was provoked by the installation of constipation which is associated with a disturbance in the balance between the production of reactive oxygen species (free radicals) and antioxidant defenses in intestinal mucosal barrier. This mechanism was evaluated by lipid peroxidation, protein oxidation, and damaging actions on sulfhydryl groups. Disorders in the normal redox state of cells can induce toxic activities through the generation of free radical reactive oxygen species that induce cell injury and alter these cellular macromolecules [14]. These obtained results are in agreement with those found by Li et al. [15] who revealed that the level of MDA augmented in constipated rats. In addition, other previous reports indicate that chronic constipation can cause potential oxidative stress in children and depletion of antioxidant enzyme activities [16].

#### **2.3. Nitrosative stress and diarrhea**

The castor oil-induced diarrhea model and intestinal mucosal injury responses may involve the nitric oxide that caused an enhancement of epithelial layer permeability to calcium ions, leading to an accumulation of intracellular Ca2+ and improvement of calmudin activation of NO synthetase action. At this level, the NO could cause the hypersecretion process in the small bowel. It was later proved in many research studies that NO and prostaglandins are strongly involved in the inflammatory pathway produced by castor oil [17].

[6] Marleen TJVA, Arjan JS, Carolien V, Robert JB, Roelof VM, et al. Intestinal barrier function in response to abundant or depleted mucosal glutathione in salmonella-infected

Role of Oxidative/Nitrosative Stress in Diarrhea and Constipation

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69

[7] Sebai H, Jabri MA, Souli A, Rtibi K, Selmi S, et al. Antidiarrheal and antioxidant activities of chamomile (Matricariarecutita) decoction in rats. Journal of Ethnopharmacology.

[8] Rao CV, Vijayakumar M, Sairam K, Kumar V. Antidiarrhoeal activity of the standardised extract of Cinnamomumtamala in experimental rats. Journal of Natural Medicines.

[9] Tandon R, Khanna HD, Dorababu M, Goel RK. Oxidative stress and antioxidants status in peptic ulcer and gastric carcinoma. Indian Journal of Physiology and Pharmacology.

[10] Jabri MA, Rtibi K, Saklya M, Marzouki L, Sebai H. Role of gastrointestinal motility inhibition and antioxidant properties of myrtle berries (Myrtuscommunis L.) juice in diar-

[11] Jówko E, Długołęcka B, Makaruk B, Cieśliński I. The effect of green tea extract supplementation on exercise-induced oxidative stress parameters in male sprinters. European

[12] Michael S, Navdeep SC. ROS function in redox signaling and oxidative stress. Current

[13] Rtibi K, Selmi S, Grami D, Sebaia H, Amri M, Marzouki L. Irinotecan chemotherapyinduced intestinal oxidative stress:Underlying causes of disturbed mucosal water and

[14] Rtibi K, Grami D, Selmi S, Amri M, Sebai H, Marzouki L. Vinblastine, an anticancer drug, causes constipation and oxidative stress as well as others disruptions in intestinal

[15] Li Y, Zong Y, Qi J, Liu K. Prebiotics and oxidative stress in constipated rats. Journal of

[16] Jun-Fu Z, Jian-Guo L, Sheng-Li Z, Ji-Yue W. Potential oxidative stress in childrenwith

[17] Sharma P, Vidyasagar G, Bhandari A, Singh S, Bhadoriya U, Ghule S, Dubey N. A pharmacological evaluation of antidiarrhoeal activity of leaves extract of *Murrayakoenigii* in experimentally induced diarrhoea in rats. Asian Pacific Journal of Tropical Medicine.

chronic constipation. World Journal of Gastroenterology. 2005;**11**:368-371

electrolyte transport. Pathophysiology. 2017;**07**:002. DOI: 10.1016/j.pathophys

rats. BMC Physiology. 2009;**9**:6. DOI: 10.1186/1472-6793-9-6

2014;**152**(2):327-332. DOI: 10.1016/j.jep.2014.01.015

rhea treatment. Biomed Pharma. 2016;**84**:1937-1944

tract in rat. Toxicology Reports. 2017;**4**:221-225

Pediatric Gastroenterology and Nutrition. 2011;**53**:447-452

Journal of Nutrition. 2015;**54**:783-791

Biology. 2014;**24**:R453-R462

2012;**2**:230-233

2008;**62**:396-402

2004;**48**:115-118

### **3. Conclusion**

These data clearly demonstrate the implication of oxidative/nitrosative stress in gastrointestinal disorders such as diarrhea and constipation.

### **Author details**

Kaïs Rtibi\*, Hichem Sebai and Lamjed Marzouki

\*Address all correspondence to: rtibikais@yahoo.fr

Laboratory of Functional Physiology and Valorization of Bioresources, Higher Institute of Biotechnology of Beja, Beja, Tunisia

### **References**


[6] Marleen TJVA, Arjan JS, Carolien V, Robert JB, Roelof VM, et al. Intestinal barrier function in response to abundant or depleted mucosal glutathione in salmonella-infected rats. BMC Physiology. 2009;**9**:6. DOI: 10.1186/1472-6793-9-6

**2.3. Nitrosative stress and diarrhea**

68 Novel Prospects in Oxidative and Nitrosative Stress

nal disorders such as diarrhea and constipation.

Kaïs Rtibi\*, Hichem Sebai and Lamjed Marzouki \*Address all correspondence to: rtibikais@yahoo.fr

Biotechnology of Beja, Beja, Tunisia

Research. 2013;**7**:580-588

PMID:22267761]

2008;**116**:125-130

**3. Conclusion**

**Author details**

**References**

The castor oil-induced diarrhea model and intestinal mucosal injury responses may involve the nitric oxide that caused an enhancement of epithelial layer permeability to calcium ions, leading to an accumulation of intracellular Ca2+ and improvement of calmudin activation of NO synthetase action. At this level, the NO could cause the hypersecretion process in the small bowel. It was later proved in many research studies that NO and prostaglandins are

These data clearly demonstrate the implication of oxidative/nitrosative stress in gastrointesti-

Laboratory of Functional Physiology and Valorization of Bioresources, Higher Institute of

[1] Pandey R, Singh M, Singhal U, Gupta KB, Aggarwrwal SK. Nitrosative stress and the pathobiology of chronic obstructive pulmonary disease. Journal of Clinical and Diagnostic

[2] Aoshiba K, Zhou F, Tsuji T, Nagai A. DNA damage as a molecular link in the pathogenesis of COPD in smokers. European Respiratory Journal. 2012 [Epub ahead of print,

[3] Tashiro N, Budhathoki S, Ohnaka K, Toyomura K, Kono S, Ueki T, Tanaka M, Kakeji Y, Maehara Y, Okamura T, Ikejiri K, Futami K, Maekawa T, Yasunami Y, Takenaka K, Ichimiya H, Terasaka R. Constipation and colorectal cancer risk: The Fukuoka colorectal

[4] Suleiman MM, Dzenda T, Sani CA. Antidiarrhoeal activity of the methanol stem-bark extract of Annona senegalensis Pers. (Annonaceae). Journal of Ethnopharmacology.

[5] Buccigrossi V, Laudiero G, Russo C, Miele E, Sofia M, et al. Chloride secretion induced by rotavirus is oxidative stress-dependent and inhibited by Saccharomyces boulardii in

human enterocytes. PLoS One. 2014;**9**:e99830. DOI: 10.1371/journal.pone.0099830

cancer study. Asian Pacific Journal of Cancer Prevention. 2011;**12**:2025-2030

strongly involved in the inflammatory pathway produced by castor oil [17].


**Chapter 4**

**Provisional chapter**

**Role of Reactive Oxygen Species in Male Reproduction**

The production of reactive oxygen species (ROS) is a normal physiological event in the male germ line. ROS are a double-edged sword, despite its role as key signaling molecules in physiological processes such as capacitation and hyperactivation, its overproduction which overwhelms the body's antioxidant defenses is thought to affect male fertility and normal embryonic development. The excess generation of ROS in semen by exogenous and endogenous factors has been recognized as detrimental etiologies for male infertilities. Spermatozoa are vulnerable to ROS attack because they are rich in mitochondria, have abundance of substrates for free radical attack and their capacity to protect themselves from oxidative stress is limited. The cytotoxic aldehydes generated as a result of lipid peroxidation are known to form adduct with the mitochondrial protein involved in electron transport chain and stimulate generation of ROS in mitochondria. ROS and their metabolites can lead to oxidative DNA damage in mitochondria and nucleus that eventually culminates in DNA fragmentation. The presence for large amount of damaged DNA is a major characteristic of defective human spermatozoa, which affect the fertility and pregnancy outcome. Thus, as a comprehensive approach, treatment of oxidative stress should involve strategies to reduce stress-provoking conditions to help reverse sperm dysfunction.

Infertility is a disorder affecting 10–15% couples of reproductive age worldwide [1, 2]. It is defined as the inability of a couple to achieve spontaneous pregnancy after 1 year of regular, unprotected sexual intercourse [3]. The inability to have children affects the infertile couples psychologically and it may lead to depression, suicidal tendencies and other pathological and psychological conditions [4, 5]. Although, fertility may decrease with increase in age, but often occurs as a result of anatomic defects, endocrinopathies, immunologic problems, gene

**Keywords:** ROS, oxidative stress, male infertility, DNA damage

**Role of Reactive Oxygen Species in Male Reproduction**

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use,

distribution, and reproduction in any medium, provided the original work is properly cited.

DOI: 10.5772/intechopen.74763

Sabiha Fatima

Sabiha Fatima

**Abstract**

**1. Introduction**

Additional information is available at the end of the chapter

Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.74763

#### **Role of Reactive Oxygen Species in Male Reproduction Role of Reactive Oxygen Species in Male Reproduction**

DOI: 10.5772/intechopen.74763

#### Sabiha Fatima Sabiha Fatima

Additional information is available at the end of the chapter Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/intechopen.74763

#### **Abstract**

The production of reactive oxygen species (ROS) is a normal physiological event in the male germ line. ROS are a double-edged sword, despite its role as key signaling molecules in physiological processes such as capacitation and hyperactivation, its overproduction which overwhelms the body's antioxidant defenses is thought to affect male fertility and normal embryonic development. The excess generation of ROS in semen by exogenous and endogenous factors has been recognized as detrimental etiologies for male infertilities. Spermatozoa are vulnerable to ROS attack because they are rich in mitochondria, have abundance of substrates for free radical attack and their capacity to protect themselves from oxidative stress is limited. The cytotoxic aldehydes generated as a result of lipid peroxidation are known to form adduct with the mitochondrial protein involved in electron transport chain and stimulate generation of ROS in mitochondria. ROS and their metabolites can lead to oxidative DNA damage in mitochondria and nucleus that eventually culminates in DNA fragmentation. The presence for large amount of damaged DNA is a major characteristic of defective human spermatozoa, which affect the fertility and pregnancy outcome. Thus, as a comprehensive approach, treatment of oxidative stress should involve strategies to reduce stress-provoking conditions to help reverse sperm dysfunction.

**Keywords:** ROS, oxidative stress, male infertility, DNA damage

#### **1. Introduction**

Infertility is a disorder affecting 10–15% couples of reproductive age worldwide [1, 2]. It is defined as the inability of a couple to achieve spontaneous pregnancy after 1 year of regular, unprotected sexual intercourse [3]. The inability to have children affects the infertile couples psychologically and it may lead to depression, suicidal tendencies and other pathological and psychological conditions [4, 5]. Although, fertility may decrease with increase in age, but often occurs as a result of anatomic defects, endocrinopathies, immunologic problems, gene

Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons

mutation, ejaculatory failures or radiation, chemotherapy and environmental exposures [6–9]. In approximately half of all the cases of infertility, male factor is the sole or major contributing factor with no identifiable cause found in over 25% of infertile males [10, 11]. In approximately 40–50% of the male infertility cases, oxidative stress-related mechanisms are found to be responsible for the impairment of the sperm function and fertilization [12]. Oxidative stress is a disturbance in the balance between the systemic manifestation of reactive oxygen species (ROS) and the ability of the body to counteract their harmful effects through neutralization by antioxidant defense mechanism [13]. ROS such as superoxide anion (O2 − ), hydrogen peroxide (H2 O2 ), and hydroxyl radical (HO•) are highly reactive oxidizing agents produced continuously during metabolic processes [14]. Oxidative processes related to spermatozoa are particularly of interest as they exhibit a double-edged sword role in these cells (**Figure 1**). The physiological level of ROS is necessary to regulate a critical redox-sensitive processes such as capacitation and hyperactivation without which fertilization is impossible [15]. While its supraphysiological level affects normal spermatogenesis and sperm functions such as motility, capacitation, acrosome reaction, egg penetration and decondensation of sperm head, which is essential to achieve fertilization. Spermatogenesis is a metabolically active biological process during which haploid spermatozoa are produced in the seminiferous tubules. During this process O2 − are generated as a natural by-product of cellular respiration. The germ cells undergoing differentiation to spermatids in testes are protected from oxidative stress by its nurse cells called sertoli cells which possess high level of antioxidant enzymes such as superoxide dismutase (SOD) as well as the reductase, transferase, and peroxidase activities of the glutathione cycle [16]. Once the spermatozoa are released from the germinal epithelium, they become vulnerable to oxidative attack as they are no longer protected by defense mechanism of sertoli cells [13, 17]. Excess ROS can lead to cellular injury by damaging DNA, lipids, and

proteins in the cells [18]. Thus, the ROS must be maintained at physiological levels for optimal sperm function, the maintenance of cellular homeostasis, and redox-sensitive signal transduc-

During their transit through the epididymis, spermatozoa progressively acquire the ability to move but lack fertilizing capacity [19]. They acquire the ability to fertilize in the female tract through a series of physiological changes called 'capacitation' which involves hyperactivation, acrosomal reaction, and sperm-oocyte fusion. Mammalian sperm capacitation is a redox regulated process which requires the production of different types of ROS to promote the fertilization of spermatozoa to the mature oocytes [20, 21]. The primary ROS generated in human

reduction product of oxygen generated reacts with itself via dismutation reaction, which is

lations of mammalian spermatozoa generate ROS mainly by two mechanisms: the membrane nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, an enzyme complex that is contained in the plasma membrane; and the mitochondrial nicotinamide adenine dinucleotide (NADH)-dependent oxido-reductase [23, 24]. The NADPH required by NADPH oxidase can be supplied by dehydrogenases located both in the plasma membrane and the cytosol. Studies have suggested the activation of sperm plasma membrane oxidase during capacitation and acrosome reaction [25, 26]. In mammalian spermatozoa, NADPH oxidase 5 (NOX5)

of the sperm generates a low level of ROS during steady-state respiration but have the potential to accelerate this activity when these gametes enter the intrinsic apoptotic pathway [28,

oxide synthase (NOS) and result in the formation of powerful oxidant ONOO−, which mediates the oxidation of cholesterol to oxysterols. The oxysterols then exit the plasma membrane dramatically to enhance membrane fluidity [31, 32]. Further, the combined action of ONOO−

cyclase, thereby stimulating cAMP production and the activation of protein kinase A (PKA) [33–35]. Activated PKA phosphorylates and inhibits protein phosphatase and activates tyrosine kinase that leads to an increase in actin polymerization, an essential process required for the development of hyperactivated motility [36, 37]. Only hyperactivated spermatozoa have increased motility to undergo acrosome reaction and acquire the characteristics required for

tion in vitro has been well documented [38]. The hyperactivated spermatozoon traverse the cumulus oophorus surrounding ovulated eggs, it then binds and penetrate to the zonapellucida (ZP) of the oocyte and initiates an exocytotic release of proteolytic enzymes, creating a

generated from these two sources is thought to combine with •NO produced by nitric

concomitantly lead to the inhibition of tyrosine phosphatase activity while the

to stimulate sperm capitation includes nitric oxide (•NO) and peroxynitrite (ONOO<sup>−</sup>

−

O2

−

which appears to play a role in this process [22]. This one-electron

. It has been reported that the capacitating popu-

Role of Reactive Oxygen Species in Male Reproduction http://dx.doi.org/10.5772/intechopen.74763 73

[27]. The mitochondria located in the mid-piece region

), and calcium ions (Ca2+) activates soluble adenylyl

in the initiation of hyperactiva-

) [30, 31].

, a variety of secondary cytotoxic radicals which are reported

tion mechanisms affecting fertility.

**2. ROS and sperm physiology**

−

greatly accelerated by SOD, to generate H2

are actively involved in generating O2

−

O2

 and O2 −

, bicarbonate (HCO3

successful fertilization. The role of low concentrations of OH<sup>−</sup>

spermatozoa is the O2

29]. In addition to H2

The O2 −

and H2

O2

combination of O2

**Figure 1.** Physiological and pathological role of ROS in male reproduction.

proteins in the cells [18]. Thus, the ROS must be maintained at physiological levels for optimal sperm function, the maintenance of cellular homeostasis, and redox-sensitive signal transduction mechanisms affecting fertility.

### **2. ROS and sperm physiology**

mutation, ejaculatory failures or radiation, chemotherapy and environmental exposures [6–9]. In approximately half of all the cases of infertility, male factor is the sole or major contributing factor with no identifiable cause found in over 25% of infertile males [10, 11]. In approximately 40–50% of the male infertility cases, oxidative stress-related mechanisms are found to be responsible for the impairment of the sperm function and fertilization [12]. Oxidative stress is a disturbance in the balance between the systemic manifestation of reactive oxygen species (ROS) and the ability of the body to counteract their harmful effects through neutraliza-

continuously during metabolic processes [14]. Oxidative processes related to spermatozoa are particularly of interest as they exhibit a double-edged sword role in these cells (**Figure 1**). The physiological level of ROS is necessary to regulate a critical redox-sensitive processes such as capacitation and hyperactivation without which fertilization is impossible [15]. While its supraphysiological level affects normal spermatogenesis and sperm functions such as motility, capacitation, acrosome reaction, egg penetration and decondensation of sperm head, which is essential to achieve fertilization. Spermatogenesis is a metabolically active biological process during which haploid spermatozoa are produced in the seminiferous tubules. During

undergoing differentiation to spermatids in testes are protected from oxidative stress by its nurse cells called sertoli cells which possess high level of antioxidant enzymes such as superoxide dismutase (SOD) as well as the reductase, transferase, and peroxidase activities of the glutathione cycle [16]. Once the spermatozoa are released from the germinal epithelium, they become vulnerable to oxidative attack as they are no longer protected by defense mechanism of sertoli cells [13, 17]. Excess ROS can lead to cellular injury by damaging DNA, lipids, and

), and hydroxyl radical (HO•) are highly reactive oxidizing agents produced

are generated as a natural by-product of cellular respiration. The germ cells

−

), hydrogen

tion by antioxidant defense mechanism [13]. ROS such as superoxide anion (O2

peroxide (H2

this process O2

O2

72 Novel Prospects in Oxidative and Nitrosative Stress

−

**Figure 1.** Physiological and pathological role of ROS in male reproduction.

During their transit through the epididymis, spermatozoa progressively acquire the ability to move but lack fertilizing capacity [19]. They acquire the ability to fertilize in the female tract through a series of physiological changes called 'capacitation' which involves hyperactivation, acrosomal reaction, and sperm-oocyte fusion. Mammalian sperm capacitation is a redox regulated process which requires the production of different types of ROS to promote the fertilization of spermatozoa to the mature oocytes [20, 21]. The primary ROS generated in human spermatozoa is the O2 − which appears to play a role in this process [22]. This one-electron reduction product of oxygen generated reacts with itself via dismutation reaction, which is greatly accelerated by SOD, to generate H2 O2 . It has been reported that the capacitating populations of mammalian spermatozoa generate ROS mainly by two mechanisms: the membrane nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, an enzyme complex that is contained in the plasma membrane; and the mitochondrial nicotinamide adenine dinucleotide (NADH)-dependent oxido-reductase [23, 24]. The NADPH required by NADPH oxidase can be supplied by dehydrogenases located both in the plasma membrane and the cytosol. Studies have suggested the activation of sperm plasma membrane oxidase during capacitation and acrosome reaction [25, 26]. In mammalian spermatozoa, NADPH oxidase 5 (NOX5) are actively involved in generating O2 − [27]. The mitochondria located in the mid-piece region of the sperm generates a low level of ROS during steady-state respiration but have the potential to accelerate this activity when these gametes enter the intrinsic apoptotic pathway [28, 29]. In addition to H2 O2 and O2 − , a variety of secondary cytotoxic radicals which are reported to stimulate sperm capitation includes nitric oxide (•NO) and peroxynitrite (ONOO<sup>−</sup> ) [30, 31]. The O2 − generated from these two sources is thought to combine with •NO produced by nitric oxide synthase (NOS) and result in the formation of powerful oxidant ONOO−, which mediates the oxidation of cholesterol to oxysterols. The oxysterols then exit the plasma membrane dramatically to enhance membrane fluidity [31, 32]. Further, the combined action of ONOO− and H2 O2 concomitantly lead to the inhibition of tyrosine phosphatase activity while the combination of O2 − , bicarbonate (HCO3 − ), and calcium ions (Ca2+) activates soluble adenylyl cyclase, thereby stimulating cAMP production and the activation of protein kinase A (PKA) [33–35]. Activated PKA phosphorylates and inhibits protein phosphatase and activates tyrosine kinase that leads to an increase in actin polymerization, an essential process required for the development of hyperactivated motility [36, 37]. Only hyperactivated spermatozoa have increased motility to undergo acrosome reaction and acquire the characteristics required for successful fertilization. The role of low concentrations of OH<sup>−</sup> in the initiation of hyperactivation in vitro has been well documented [38]. The hyperactivated spermatozoon traverse the cumulus oophorus surrounding ovulated eggs, it then binds and penetrate to the zonapellucida (ZP) of the oocyte and initiates an exocytotic release of proteolytic enzymes, creating a pore in ZP's extracellular matrix. For successful fertilization, the spermatozoa then penetrate this physical zona barrier and fuse with the oocyte [39, 40]. Thus, ROS during the capacitation and acrosome reaction has been shown to increase the membrane fluidity and rates of spermoocyte fusion (**Figure 1**).

antioxidant mechanisms. These mechanisms compensate for the deficiency in cytoplasmic enzymes in sperm [54, 55]. Thus the sperms which spend long period as an isolated cells both in male and female genital tracts (approximately 3 weeks), these limited defenses can be eas-

Spermatozoa like other aerobic cells are dependent on cellular respiration process which supports its life. But excessive generation of its metabolites, such as ROS, can modify cell functions. Hence, under normal condition male reproductive system must continuously inactivate ROS to maintain a balance between ROS production and its scavenging mechanism in order to keep only the small amount necessary to maintain normal cell function. Thus, in order to maintain the redox homeostasis, the mature spermatozoa with limited antioxidant defense capacity are mainly dependent on seminal plasma which is well endowed with an array of

The main enzymatic antioxidants in the semen include superoxide dismutase (SOD), catalase, and glutathione peroxidase/glutathione reductase (GPX/GRD) system [59]. SOD is metalloenzymes which is present in both intracellular and extracellular forms [60]. SOD spontane-

enzyme of the antioxidant system in the semen is glutathione peroxidase (GPX), which catalyzes the reduction of hydrogen peroxide and organic peroxides, including the peroxides of phospholipids [61]. Spermatozoa have limited supply of catalase and GPX, while SOD is the main enzymatic antioxidant which protects it from oxidative stress [62]. Beside the enzymes antioxidant protective mechanism, seminal plasma is also employed by the low molecular weight, non-enzymatic antioxidants that assist enzyme activity. These include ascorbic acid (vitamin C), tocopherol (vitamin E), vitamin A, pantothenic acid, coenzyme Q10, carnitine, amino acids (taurine, hypotaurine) zinc, selenium albumin, and urate. These agents principally act by directly neutralizing free radical activity chemically and some of these antioxidants are reported to enhance sperm viability/motility as well as normal sperm morphology and required for spermatogenesis, development of spermatozoa [63, 64]. The seminal plasma antioxidants concentrations have been shown to be significantly higher in fertile men than

Oxidants in seminal plasma originate from numerous extrinsic and intrinsic sources. The human ejaculate is composed of various types of cells, which include mature and immature cells, round cells from extraordinary degrees of spermatogenesis, leukocytes, and epithelial cells [67, 68]. Of those, leukocytes, specially neutrophils and macrophages and immature

and catalase catalyzes the decomposition of H2

O) thus preventing the lipid peroxidation of the sperm plasma membrane. Another

O2 to O2

Role of Reactive Oxygen Species in Male Reproduction http://dx.doi.org/10.5772/intechopen.74763

and

75

effective enzymatic and non-enzymatic antioxidant defense mechanisms [57, 58].

ily overwhelmed with an increased generation of ROS [56].

**4. ROS scavenging capacity of semen**

ously dismutase O2

water (H2

−

those in infertile men [65, 66].

**5. Sources of ROS in seminal plasma**

to form H2

O2

### **3. Human spermatozoa are vulnerable to oxidative stress**

During spermatogenesis, germ cells produce high levels of reactive oxygen species, but fortunately a complex of antioxidant defense system and DNA repair system exists in the testis that protects genome integrity in differentiating sperm [16]. In normal spermatogenesis, the developing spermatozoa extrude most of the cytoplasm by the action of sertoli cells to change to a condensed, elongated form [41]. The lack of cytoplasm results in decreased intrinsic antioxidant defense due to the loss of most of antioxidant enzymes, rendering the cells less protected against ROS by the time they are discharged into the epididymis [42, 43]. Further, they also lack the necessary cytoplasmic-enzyme repair systems, thus they have very limited capacity for detection and repair of DNA damage [44]. Therefore, during their transit and storage into the epididymis or post-ejaculation they have no DNA repair mechanism, and thus cannot synthesize DNA, RNA, or translate proteins (such as repair enzymes) [45, 46]. The mammalian spermatozoa are vulnerable to oxidative stress not only because of their inherent free radical generating activity and lack of endogenous antioxidant protection, but also due to the abundant substrates that these cells possess for free radical attack. In mature spermatozoa, the small cytoplasm with limited defense remains confined to the mid-piece region in the vicinity of the mitochondria. As a result, the plasma membrane richly endowed with high concentrations of polyunsaturated fatty acids (PUFAs), particularly docosahexaenoic acid (DHA) (22:6) and arachidonic acids (20:4) containing six and four carbon-carbon double bonds per molecule, surrounding the acrosome and the tail are not protected by the intracellular antioxidants [47–49]. In human spermatozoa, approximately 50% of the fatty acids are composed of DHA which is thought to play a major role in regulating spermatogenesis and membrane fluidity [18]. The presence of double bond in PUFAs adjacent to a methylene group weakens the methyl carbon-hydrogen bonds and makes hydrogen more susceptible to abstraction and thus vulnerable to oxidation [44, 50].

Sperm mitochondrial DNA has long been postulated as a sources and often likely target of ROS oxidation as they are not protected by histones and has a very limited capacity for DNA repair with complete lack of nucleotide-excision repair pathways [51]. It is estimated that the mitochondrial DNA exhibits the mutation rate two orders of magnitude higher than that of nuclear DNA. Thus any quantitative or qualitative aberrations in mitochondrial DNA will result in the increased ROS generation which will affect the cellular functioning of the cell [52].

Despite its vulnerability to oxidative stress, maturing sperms spontaneously generate ROS during their progress through the epididymis, as its normal metabolite that aids it to acquire full fertility competence [53]. The lack of intrinsic antioxidant protection forces these cells to dependent on defense provided by seminal and epididymal enzymatic and non-enzymatic antioxidant mechanisms. These mechanisms compensate for the deficiency in cytoplasmic enzymes in sperm [54, 55]. Thus the sperms which spend long period as an isolated cells both in male and female genital tracts (approximately 3 weeks), these limited defenses can be easily overwhelmed with an increased generation of ROS [56].

### **4. ROS scavenging capacity of semen**

pore in ZP's extracellular matrix. For successful fertilization, the spermatozoa then penetrate this physical zona barrier and fuse with the oocyte [39, 40]. Thus, ROS during the capacitation and acrosome reaction has been shown to increase the membrane fluidity and rates of sperm-

During spermatogenesis, germ cells produce high levels of reactive oxygen species, but fortunately a complex of antioxidant defense system and DNA repair system exists in the testis that protects genome integrity in differentiating sperm [16]. In normal spermatogenesis, the developing spermatozoa extrude most of the cytoplasm by the action of sertoli cells to change to a condensed, elongated form [41]. The lack of cytoplasm results in decreased intrinsic antioxidant defense due to the loss of most of antioxidant enzymes, rendering the cells less protected against ROS by the time they are discharged into the epididymis [42, 43]. Further, they also lack the necessary cytoplasmic-enzyme repair systems, thus they have very limited capacity for detection and repair of DNA damage [44]. Therefore, during their transit and storage into the epididymis or post-ejaculation they have no DNA repair mechanism, and thus cannot synthesize DNA, RNA, or translate proteins (such as repair enzymes) [45, 46]. The mammalian spermatozoa are vulnerable to oxidative stress not only because of their inherent free radical generating activity and lack of endogenous antioxidant protection, but also due to the abundant substrates that these cells possess for free radical attack. In mature spermatozoa, the small cytoplasm with limited defense remains confined to the mid-piece region in the vicinity of the mitochondria. As a result, the plasma membrane richly endowed with high concentrations of polyunsaturated fatty acids (PUFAs), particularly docosahexaenoic acid (DHA) (22:6) and arachidonic acids (20:4) containing six and four carbon-carbon double bonds per molecule, surrounding the acrosome and the tail are not protected by the intracellular antioxidants [47–49]. In human spermatozoa, approximately 50% of the fatty acids are composed of DHA which is thought to play a major role in regulating spermatogenesis and membrane fluidity [18]. The presence of double bond in PUFAs adjacent to a methylene group weakens the methyl carbon-hydrogen bonds and makes hydrogen more susceptible to abstraction and

Sperm mitochondrial DNA has long been postulated as a sources and often likely target of ROS oxidation as they are not protected by histones and has a very limited capacity for DNA repair with complete lack of nucleotide-excision repair pathways [51]. It is estimated that the mitochondrial DNA exhibits the mutation rate two orders of magnitude higher than that of nuclear DNA. Thus any quantitative or qualitative aberrations in mitochondrial DNA will result in the increased ROS generation which will affect the cellular functioning of the cell [52]. Despite its vulnerability to oxidative stress, maturing sperms spontaneously generate ROS during their progress through the epididymis, as its normal metabolite that aids it to acquire full fertility competence [53]. The lack of intrinsic antioxidant protection forces these cells to dependent on defense provided by seminal and epididymal enzymatic and non-enzymatic

**3. Human spermatozoa are vulnerable to oxidative stress**

oocyte fusion (**Figure 1**).

74 Novel Prospects in Oxidative and Nitrosative Stress

thus vulnerable to oxidation [44, 50].

Spermatozoa like other aerobic cells are dependent on cellular respiration process which supports its life. But excessive generation of its metabolites, such as ROS, can modify cell functions. Hence, under normal condition male reproductive system must continuously inactivate ROS to maintain a balance between ROS production and its scavenging mechanism in order to keep only the small amount necessary to maintain normal cell function. Thus, in order to maintain the redox homeostasis, the mature spermatozoa with limited antioxidant defense capacity are mainly dependent on seminal plasma which is well endowed with an array of effective enzymatic and non-enzymatic antioxidant defense mechanisms [57, 58].

The main enzymatic antioxidants in the semen include superoxide dismutase (SOD), catalase, and glutathione peroxidase/glutathione reductase (GPX/GRD) system [59]. SOD is metalloenzymes which is present in both intracellular and extracellular forms [60]. SOD spontaneously dismutase O2 − to form H2 O2 and catalase catalyzes the decomposition of H2 O2 to O2 and water (H2 O) thus preventing the lipid peroxidation of the sperm plasma membrane. Another enzyme of the antioxidant system in the semen is glutathione peroxidase (GPX), which catalyzes the reduction of hydrogen peroxide and organic peroxides, including the peroxides of phospholipids [61]. Spermatozoa have limited supply of catalase and GPX, while SOD is the main enzymatic antioxidant which protects it from oxidative stress [62]. Beside the enzymes antioxidant protective mechanism, seminal plasma is also employed by the low molecular weight, non-enzymatic antioxidants that assist enzyme activity. These include ascorbic acid (vitamin C), tocopherol (vitamin E), vitamin A, pantothenic acid, coenzyme Q10, carnitine, amino acids (taurine, hypotaurine) zinc, selenium albumin, and urate. These agents principally act by directly neutralizing free radical activity chemically and some of these antioxidants are reported to enhance sperm viability/motility as well as normal sperm morphology and required for spermatogenesis, development of spermatozoa [63, 64]. The seminal plasma antioxidants concentrations have been shown to be significantly higher in fertile men than those in infertile men [65, 66].

### **5. Sources of ROS in seminal plasma**

Oxidants in seminal plasma originate from numerous extrinsic and intrinsic sources. The human ejaculate is composed of various types of cells, which include mature and immature cells, round cells from extraordinary degrees of spermatogenesis, leukocytes, and epithelial cells [67, 68]. Of those, leukocytes, specially neutrophils and macrophages and immature spermatozoa are taken into consideration as the primary endogenous assets of ROS [69], while numerous life style elements including immoderate smoking and alcohol intake, and environmental elements inclusive of radiation and pollution can contribute as exogenous sources of ROS (**Figure 2**) [70, 71]. Exposure to radiation and toxins induces ROS production which impairs spermatogenesis and leads to DNA damage in human spermatozoa, which further decreases the motility and vitality of sperm cells as well as their concentration depending on the duration of exposure [72]. Cigarette smoking is found to be correlated with leukocytospermia. It has been reported that smoking can elevate the leukocyte concentration by 48% and ROS by 107% in seminal plasma [73].

against infections as well as inflammation [78]. However, the high concentrations of ROS may overwhelm seminal antioxidant defenses and damage the sperm cell [79]. Essentially, the cellular mechanisms for the generation of ROS within leukocytes and spermatozoa are same, in leukocytes, the release of the large amounts of superoxide into phagocytic vesicles

Role of Reactive Oxygen Species in Male Reproduction http://dx.doi.org/10.5772/intechopen.74763 77

The exact mechanism of oxidative stress-induced decline in sperm function remains unknown but is mainly attributed to peroxidative damage to axoneme and depletion of intracellular ATP levels, followed by generation of 4-hydroxynonenal (4-HNE) and malondialdehyde (MDA) owing to the oxidation of lipid membrane components and oxidation of DNA fol-

In PUFAs, the hydroxyl radicals attack lipids containing carbon-carbon double bond and promote the hydrogen abstraction from carbon to generate a carbon-centered lipid radical (2CH−•) that then combines with oxygen to generate lipid peroxyl radicals (ROO•) [83]. The ROO• radicals subsequently attacks another lipid molecule, abstract a hydrogen atom in order to stabilize itself as the lipid hydroperoxide but in the process generates another carbon-centered lipid radical that perpetuates the cascade of chemical reactions called lipid peroxidation. The process results in the generation of small molecular mass electrophilic lipid aldehydes such as 4-hydroxynonenal (4HNE), acrolein, and malondialdehyde [84]. Lipid peroxidation (LPO) is extremely harmful to spermatozoa, having a dramatic effect on both sperm movement and the competence of these cells for fertilization (**Figure 3**). Immature human sperm cells contain high levels of DHA in the cytoplasmatic droplet and showed more sus-

Added to this vulnerability, it has been shown that cytotoxic aldehydes generated as the result of oxidative stress has the ability to of triggering ROS generation by the sperm mitochondria in a self-perpetuating cycle; the greater the level of unsaturation, the greater the level of the stimulatory effect. The defective human spermatozoa contain abnormally high cellular contents of free polyunsaturated fatty acids, the levels of which are positively correlated with mitochondrial superoxide generation. The lipid aldehydes, 4HNE or acrolein bind covalently to the nucleophilic centers of vulnerable proteins, such as succinic acid dehydrogenase and form a protein adducts in the mitochondrial electron transport chain (ETC) that results in the leakage of electrons which disturbs the normal flow of electrons and reduction of oxygen to water [86, 87]. The leakage of electrons from the ETC results in the reduction of oxygen to gen-

, which then by mitochondrial superoxide dismutase rapidly dismutates to H2

The excess of cytoplasm in the immature or defective spermatozoa contain superabundance of cytoplasmic enzymes. The retention of excess of SOD can only be an asset for any cell seeking to protect itself from oxidative stress if it is accompanied by a corresponding increase in the

O2 [88].

for killing the pathogens [80, 81].

**6.1. Lipid peroxidation**

erate O2 −

**6. Impact of oxidative stress on spermatozoa**

lowed by fragmentation of both nuclear and mitochondrial DNA [82].

ceptibility to LPO than normal matured sperm with lower DHA levels [85].

#### **5.1. Immature/abnormal spermatozoa**

One of the major cellular sources of ROS in the semen is sperm cells [74]. When spermatogenesis is impaired, the cytoplasmic extrusion mechanisms are defective, and spermatozoa are released from the germinal epithelium carrying surplus cytoplasmic residues in the midpiece [75]. These residues are rich in the cytoplasmic enzymes such as superoxide dismutase, lactic acid dehydrogenase, glucose-6-phosphate dehydrogenase (G6PDH), and creatine kinase [69, 76]. However among these enzymes, the key enzyme was thought to be G6PDH, which would be expected to enhance the intracellular availability of NADPH via the hexose monophosphate shunt. NADPH is used to fuel the generation of ROS via NADPH oxidase activity [27, 77]

#### **5.2. Leukocytes**

The main source of ROS inside semen is leukocytes. Infection or chronic inflammation may activate the leukocytes to release 1000-times more ROS than spermatozoa [78]. This high production of ROS by leukocytes plays an important role in the cellular defense system

**Figure 2.** Extrinsic and intrinsic factors of ROS generation in seminal plasma.

against infections as well as inflammation [78]. However, the high concentrations of ROS may overwhelm seminal antioxidant defenses and damage the sperm cell [79]. Essentially, the cellular mechanisms for the generation of ROS within leukocytes and spermatozoa are same, in leukocytes, the release of the large amounts of superoxide into phagocytic vesicles for killing the pathogens [80, 81].

### **6. Impact of oxidative stress on spermatozoa**

The exact mechanism of oxidative stress-induced decline in sperm function remains unknown but is mainly attributed to peroxidative damage to axoneme and depletion of intracellular ATP levels, followed by generation of 4-hydroxynonenal (4-HNE) and malondialdehyde (MDA) owing to the oxidation of lipid membrane components and oxidation of DNA followed by fragmentation of both nuclear and mitochondrial DNA [82].

#### **6.1. Lipid peroxidation**

spermatozoa are taken into consideration as the primary endogenous assets of ROS [69], while numerous life style elements including immoderate smoking and alcohol intake, and environmental elements inclusive of radiation and pollution can contribute as exogenous sources of ROS (**Figure 2**) [70, 71]. Exposure to radiation and toxins induces ROS production which impairs spermatogenesis and leads to DNA damage in human spermatozoa, which further decreases the motility and vitality of sperm cells as well as their concentration depending on the duration of exposure [72]. Cigarette smoking is found to be correlated with leukocytospermia. It has been reported that smoking can elevate the leukocyte concentration

One of the major cellular sources of ROS in the semen is sperm cells [74]. When spermatogenesis is impaired, the cytoplasmic extrusion mechanisms are defective, and spermatozoa are released from the germinal epithelium carrying surplus cytoplasmic residues in the midpiece [75]. These residues are rich in the cytoplasmic enzymes such as superoxide dismutase, lactic acid dehydrogenase, glucose-6-phosphate dehydrogenase (G6PDH), and creatine kinase [69, 76]. However among these enzymes, the key enzyme was thought to be G6PDH, which would be expected to enhance the intracellular availability of NADPH via the hexose monophosphate shunt. NADPH is used to fuel the generation of ROS via NADPH oxidase

The main source of ROS inside semen is leukocytes. Infection or chronic inflammation may activate the leukocytes to release 1000-times more ROS than spermatozoa [78]. This high production of ROS by leukocytes plays an important role in the cellular defense system

by 48% and ROS by 107% in seminal plasma [73].

**Figure 2.** Extrinsic and intrinsic factors of ROS generation in seminal plasma.

**5.1. Immature/abnormal spermatozoa**

76 Novel Prospects in Oxidative and Nitrosative Stress

activity [27, 77]

**5.2. Leukocytes**

In PUFAs, the hydroxyl radicals attack lipids containing carbon-carbon double bond and promote the hydrogen abstraction from carbon to generate a carbon-centered lipid radical (2CH−•) that then combines with oxygen to generate lipid peroxyl radicals (ROO•) [83]. The ROO• radicals subsequently attacks another lipid molecule, abstract a hydrogen atom in order to stabilize itself as the lipid hydroperoxide but in the process generates another carbon-centered lipid radical that perpetuates the cascade of chemical reactions called lipid peroxidation. The process results in the generation of small molecular mass electrophilic lipid aldehydes such as 4-hydroxynonenal (4HNE), acrolein, and malondialdehyde [84]. Lipid peroxidation (LPO) is extremely harmful to spermatozoa, having a dramatic effect on both sperm movement and the competence of these cells for fertilization (**Figure 3**). Immature human sperm cells contain high levels of DHA in the cytoplasmatic droplet and showed more susceptibility to LPO than normal matured sperm with lower DHA levels [85].

Added to this vulnerability, it has been shown that cytotoxic aldehydes generated as the result of oxidative stress has the ability to of triggering ROS generation by the sperm mitochondria in a self-perpetuating cycle; the greater the level of unsaturation, the greater the level of the stimulatory effect. The defective human spermatozoa contain abnormally high cellular contents of free polyunsaturated fatty acids, the levels of which are positively correlated with mitochondrial superoxide generation. The lipid aldehydes, 4HNE or acrolein bind covalently to the nucleophilic centers of vulnerable proteins, such as succinic acid dehydrogenase and form a protein adducts in the mitochondrial electron transport chain (ETC) that results in the leakage of electrons which disturbs the normal flow of electrons and reduction of oxygen to water [86, 87]. The leakage of electrons from the ETC results in the reduction of oxygen to generate O2 − , which then by mitochondrial superoxide dismutase rapidly dismutates to H2 O2 [88]. The excess of cytoplasm in the immature or defective spermatozoa contain superabundance of cytoplasmic enzymes. The retention of excess of SOD can only be an asset for any cell seeking to protect itself from oxidative stress if it is accompanied by a corresponding increase in the

and dramatic remodeling of the chromatin during which most of the histones are removed from the DNA and are first replaced by transition proteins TP1 and TP2, and then by protamines P1 and P2 which are approximately half the size of histones. P1 and P2 are normally expressed in a 1:1 ratio in human sperm, and provide a tight packaging of the sperm DNA. The chromatin remodeling is facilitated by the coordinated loosening of the chromatin by histone hyperacetylation and by the DNA topoisomerase II (topo II), which produce temporary stand breaks in the sperm DNA to relieve torsional stress that results from supercoiling [96, 97]. This forms the basic packaging unit of sperm chromatin, a toroid, which is further compacted by the intramolecular and intermolecular disulfide cross-links between cysteine residues present in protamines. The tight packaging of the sperm DNA enables the entire haploid genome to be condensed and packed in a sperm head measuring 5 × 2.5 μm. This level of protect and ensures that the paternal genome is delivered in a form that allows developing embryo to accurately express genetic information Normally, these temporary strand breaks are repaired by nuclear poly (ADP-ribose) polymersases (PARP) and topoisomerase II prior to completion of spermiogenesis and ejaculation [98]. However in pathological cases, the error in chromatin remodeling and repair mechanism leads to the generation of high level of nicked and poorly protaminated nuclear DNA with relatively high nucleohistone content or abnormally high and low P1/P2 ratios [99–101]. Thus, defect in the chromatin remodeling process causes DNA damage in spermatids during spermiogenesis, this creates a state of vulnerability whereby spermatozoa become increasingly suscep-

Role of Reactive Oxygen Species in Male Reproduction http://dx.doi.org/10.5772/intechopen.74763 79

When the protection of DNA in spermatozoa, which is dependent on its close association with cysteine rich protaminesis is lost, the cells become very susceptible to oxidative DNA damage induced by several extrinsic and intrinsic factors. Deoxygenated guanine (dG) is more susceptible to oxidation than other nucleosides in DNA due to its low oxidation potential [102]. The enzyme 8-oxoguanine glycosylase 1 (OGG1) immediately clips the 8OHdG residues out of the DNA generating an abasic site, But due to the absence of base excision repair enzyme, the spermatozoa are ejaculated carrying a abasic sites in their DNA [103]. Studies have reported that the spermatozoa of subfertile patients contain particularly high levels of 8-hydroxy-2′ deoxyguanosine (8OHdG), the major oxidized base adduct formed when DNA is subjected

DNA repair does occur during spermiogenesis but stops post-spermiogenesis because spermatozoa are transcriptionally and translationally silent. They cannot undergo programmed cell death called apoptosis, due to their inherent physical architecture, the endonucleases released from the mitochondria have no access to the DNA. Thus, abortive apoptosis initiated post-meiotically, when the ability to drive the spermiogenesis process to completion is declined and the stand breaks are not repaired due to impairment in the repair process results in high levels of DNA fragmented sperm in the ejaculate [105]. Sperm with DNA fragmentation still has thepotential to fertilize and some types of stand DNA breaks in sperm can be repaired by oocytes, before the initiation of the first cleavage division, and generate normal offspring, but that

tible to oxidative damage.

to attack by ROS [104].

**7. Causes of DNA damage in spermatozoa**

**Figure 3.** (A) ROS-induced initiation and propagation of lipid peroxidation (LPO) generates lipid hydroperoxides plus a new carbon-centered radical that continues the chain reaction. (B) Mitochondrial ROS generation. The electrophilic aldehydes generated as a by-product of LPO process bind to protein of electron transport chain and further promote mitochondrial ROS generation. This process results in loss of mitochondrial membrane potential, low ATP generation, loss of sperm motility, oxidative DNA damage followed by DNA fragmentation.

presence of enzymes such as glutathione peroxidase or catalase that can scavenge H2 O2 . But excess of SOD and limited supply of glutathione peroxidase or catalase in human spermatozoa simply turns a short-lived, membrane-impermeant, relatively inert free radical O2 − into a long-lived, membrane-permeate reactive oxidant, H2 O2 [89, 90]. The damage of protein and membrane lipids due to elevated levels of ROS in mitochondria might affect the process of oxidative phosphorylation causing depletion of intracellular ATP levels leading to axonemal damage, decreased sperm viability, and increased mid-piece sperm morphological defects with deleterious effects on sperm capacitation and acrosome reaction and decline of motility and fertility [91]. The mitochondrial function as a measure of inner mitochondrial membrane potential is found to be decreased in the spermatozoa of infertile men with elevated levels of ROS production and is positively correlated with the sperm concentration [92].

#### **6.2. DNA damage in spermatozoa**

Mitochondrial DNA is particularly vulnerable to free radical attack because it is essentially unprotected and has a very limited capacity for DNA repair [93]. Sperm nuclear DNA, on the other hand, is much resistant to damage because it is tightly compacted by replacing histones with small, positively charged molecules known as protamine [94, 95]. Sperm DNA maturation and appropriate packaging are vital steps in the proper development of spermatozoa.

During the late spermatogenesis in the mammalian germinal epithelium, the differentiating spertamids are highly susceptible to DNA damage due to important changes in the cytoarchitecture and dramatic remodeling of the chromatin during which most of the histones are removed from the DNA and are first replaced by transition proteins TP1 and TP2, and then by protamines P1 and P2 which are approximately half the size of histones. P1 and P2 are normally expressed in a 1:1 ratio in human sperm, and provide a tight packaging of the sperm DNA. The chromatin remodeling is facilitated by the coordinated loosening of the chromatin by histone hyperacetylation and by the DNA topoisomerase II (topo II), which produce temporary stand breaks in the sperm DNA to relieve torsional stress that results from supercoiling [96, 97]. This forms the basic packaging unit of sperm chromatin, a toroid, which is further compacted by the intramolecular and intermolecular disulfide cross-links between cysteine residues present in protamines. The tight packaging of the sperm DNA enables the entire haploid genome to be condensed and packed in a sperm head measuring 5 × 2.5 μm. This level of protect and ensures that the paternal genome is delivered in a form that allows developing embryo to accurately express genetic information Normally, these temporary strand breaks are repaired by nuclear poly (ADP-ribose) polymersases (PARP) and topoisomerase II prior to completion of spermiogenesis and ejaculation [98]. However in pathological cases, the error in chromatin remodeling and repair mechanism leads to the generation of high level of nicked and poorly protaminated nuclear DNA with relatively high nucleohistone content or abnormally high and low P1/P2 ratios [99–101]. Thus, defect in the chromatin remodeling process causes DNA damage in spermatids during spermiogenesis, this creates a state of vulnerability whereby spermatozoa become increasingly susceptible to oxidative damage.

### **7. Causes of DNA damage in spermatozoa**

presence of enzymes such as glutathione peroxidase or catalase that can scavenge H2

zoa simply turns a short-lived, membrane-impermeant, relatively inert free radical O2

ROS production and is positively correlated with the sperm concentration [92].

long-lived, membrane-permeate reactive oxidant, H2

loss of sperm motility, oxidative DNA damage followed by DNA fragmentation.

**6.2. DNA damage in spermatozoa**

78 Novel Prospects in Oxidative and Nitrosative Stress

spermatozoa.

excess of SOD and limited supply of glutathione peroxidase or catalase in human spermato-

**Figure 3.** (A) ROS-induced initiation and propagation of lipid peroxidation (LPO) generates lipid hydroperoxides plus a new carbon-centered radical that continues the chain reaction. (B) Mitochondrial ROS generation. The electrophilic aldehydes generated as a by-product of LPO process bind to protein of electron transport chain and further promote mitochondrial ROS generation. This process results in loss of mitochondrial membrane potential, low ATP generation,

membrane lipids due to elevated levels of ROS in mitochondria might affect the process of oxidative phosphorylation causing depletion of intracellular ATP levels leading to axonemal damage, decreased sperm viability, and increased mid-piece sperm morphological defects with deleterious effects on sperm capacitation and acrosome reaction and decline of motility and fertility [91]. The mitochondrial function as a measure of inner mitochondrial membrane potential is found to be decreased in the spermatozoa of infertile men with elevated levels of

Mitochondrial DNA is particularly vulnerable to free radical attack because it is essentially unprotected and has a very limited capacity for DNA repair [93]. Sperm nuclear DNA, on the other hand, is much resistant to damage because it is tightly compacted by replacing histones with small, positively charged molecules known as protamine [94, 95]. Sperm DNA maturation and appropriate packaging are vital steps in the proper development of

During the late spermatogenesis in the mammalian germinal epithelium, the differentiating spertamids are highly susceptible to DNA damage due to important changes in the cytoarchitecture

O2

O2 . But

− into a

[89, 90]. The damage of protein and

When the protection of DNA in spermatozoa, which is dependent on its close association with cysteine rich protaminesis is lost, the cells become very susceptible to oxidative DNA damage induced by several extrinsic and intrinsic factors. Deoxygenated guanine (dG) is more susceptible to oxidation than other nucleosides in DNA due to its low oxidation potential [102]. The enzyme 8-oxoguanine glycosylase 1 (OGG1) immediately clips the 8OHdG residues out of the DNA generating an abasic site, But due to the absence of base excision repair enzyme, the spermatozoa are ejaculated carrying a abasic sites in their DNA [103]. Studies have reported that the spermatozoa of subfertile patients contain particularly high levels of 8-hydroxy-2′ deoxyguanosine (8OHdG), the major oxidized base adduct formed when DNA is subjected to attack by ROS [104].

DNA repair does occur during spermiogenesis but stops post-spermiogenesis because spermatozoa are transcriptionally and translationally silent. They cannot undergo programmed cell death called apoptosis, due to their inherent physical architecture, the endonucleases released from the mitochondria have no access to the DNA. Thus, abortive apoptosis initiated post-meiotically, when the ability to drive the spermiogenesis process to completion is declined and the stand breaks are not repaired due to impairment in the repair process results in high levels of DNA fragmented sperm in the ejaculate [105]. Sperm with DNA fragmentation still has thepotential to fertilize and some types of stand DNA breaks in sperm can be repaired by oocytes, before the initiation of the first cleavage division, and generate normal offspring, but that

### depends on the type and level of chromatin damage and the capacity of the oocyte to repair it [106]. DNA-strand breaks are extremely harmful lesions if not repaired and can lead to genomic instability and cell death. In natural conception, percentage of DNA damage has been negatively correlated to the rate of fertilization. If post fertilization oocyte make mistake in the repair process, deletions or sequence errors may be introduced, then it fabricates the possibility for *de novo* mutations, which could have a profound impact on the health and well-being of the offspring [107]. Sperm DNA damage in context to assisted reproductive technique (ART) has important clinical implications. Sperm selected for ART mostly originates from environment experiencing oxidative stress and high percentage of these sperms may have damaged DNA. If such sperms are used clinically in the form of therapy then can lead to substantial risk in pregnancy outcome. In case of intrauterine insemination (IUI) and in vitro fertilization (IVF), the use of these spermatozoa may not be cause of concern. But in case of intracellular sperm injection (ICSI), this natural selection barrier is bypassed and the spermatozoa with damaged DNA are directly injected into oocytes. Studies have reported that DNA damaged spermatozoa used in ICSI have some capacity for fertilization, but percentage of DNA damage has been negatively correlated to the rate of fertilization [108]. ROS-mediated DNA damage may be linked to an increase in early embryo death, infertility in the offspring, and high incidence of childhood cancer [109, 110]. We propose that extrinsic and intrinsic sources of ROS could make a significant contribution to the induction of OS and DNA damage in spermatozoa which can decrease pregnancy rate and affect the fertility outcome, further additional studies are clearly needed to validate this concept.

**8.2. Dietary antioxidants**

potential of these cells [119].

**9. Conclusion**

**9.1. Suggestion**

As many studies suggested that oxidative stress is a major cause of unexplained male infertility, antioxidant therapy would be expected to have a therapeutic effect in such cases. There are evidences which have suggested that oral antioxidants and herbal products can also boost male reproductive functions [116, 117]. But, despite of known effect of antioxidant on oxidative stress, very few studies conducted have any validity due to small sample size, difference in dosage and duration of therapy, and lack controls [118]. In order to make the study valid, patient's selection criteria for the trial should be based on the evidence indication oxidative stress as a key element in their pathology, a thorough diagnosis is required to determine patients that need to be supplemented. However, if this strategy is pursued, great care must be taken in selecting the most appropriate antioxidants for clinical use. Since ROS plays an important role in regulating the signal transduction cascades that drive sperm capacitation, we should ensure that any antioxidants employed in vitro do not compromise the fertilizing

Role of Reactive Oxygen Species in Male Reproduction http://dx.doi.org/10.5772/intechopen.74763 81

The study of *in vitro* antioxidants is highly relevant in the era of assisted reproduction because sperm preparation techniques in ART are potential generators of exogenous stresses that

Oxidative stress has been recognized as a major contributory factor to male infertility. Spermatozoa are professional generator of ROS as physiological level of ROS is necessary to regulate critical redox-sensitive processes such as capacitation, hyperactivation, acrosome reactions, and signaling processes to ensure appropriate fertilization. On the other hand, many endogenous and exogenous factors can elevate ROS production which can overwhelm their antioxidant mechanism. This results in male infertility via mechanisms involving the induction of peroxidative damage to the sperm plasma membrane, DNA damage, which significantly impairs sperm function. Lack of repair mechanism and abortive apoptosis in mature spermatozoa results in high levels of DNA fragmented sperm in the ejaculate. In natural conception, oocytes can repair some of stand DNA breaks, but that depends on the type and level of chromatin damage and the capacity of the oocyte to repair it. If post fertilization oocyte make mistake in the repair process it may lead to failure in fertilization. But if fertilization occurs, then it creates the possibility for *de novo* mutations, which could have a profound impact on the health and well-being of the offspring. When the natural balance between ROS and antioxidants is disturbed, the first restorative measure to be taken should be changes in lifestyle, maintaining a healthy and balanced diet, and antioxidant supplementation may then

The conventional seminological parameter in infertile cases reflects the functional competence of the spermatozoa and the fertilizing potential of the ejaculate, but the underlying mechanisms of male fertility is not known. Thus in order to enrich the diagnostic value of this

make human spermatozoa vulnerable to oxidative stress and DNA damage.

be taken together to improve the patient's health outcomes.

### **8. Management of infertility caused by oxidative stress**

Oxidative stress plays an important role in the pathophysiology of male infertility, which is caused due to pathological level of ROS and the loss of antioxidant protection for the spermatozoa. There are many factors which can induce oxidative stress and can alter seminal parameters and rate of fertilization. Thorough examination and management of some of these factors may protect the ROS-induced DNA damage and improve a couple's chances of conception either naturally or via assisted reproduction.

#### **8.1. Behavior and life style modification**

Various behaviors and lifestyles factors such as alcohol consumption, cigarette smoking, obesity, excess exposure to environmental toxicants, and psychological stress are negatively correlated with spermatogenesis and may cause oxidative stress and reduction in sperm quality [111]. The increased consumption of simple sugars and high-fat food and physical inactivity are leading causes of the growing obesity. It is suggested that abnormal hormonal regulation, dysregulation of adipocytokine, and ROS generation lead to suboptimal semen quality these patients [112]. Several systemic diseases, such as diabetes mellitus, infection, and cancer are known to cause oxidative stress-induced male infertility [113, 114]. There are studies which have shown positive correlation of exercise with improvements in semen parameters, sperm DNA integrity, and pregnancy rate [115]. Nevertheless, modification in behavior and unhealthy living, regular exercise, stress free jobs, and treatment of a patient's underlying pathology should be the first steps to reduce or eliminate stress-provoking conditions to reverse sperm dysfunction.

#### **8.2. Dietary antioxidants**

depends on the type and level of chromatin damage and the capacity of the oocyte to repair it [106]. DNA-strand breaks are extremely harmful lesions if not repaired and can lead to genomic instability and cell death. In natural conception, percentage of DNA damage has been negatively correlated to the rate of fertilization. If post fertilization oocyte make mistake in the repair process, deletions or sequence errors may be introduced, then it fabricates the possibility for *de novo* mutations, which could have a profound impact on the health and well-being of the offspring [107]. Sperm DNA damage in context to assisted reproductive technique (ART) has important clinical implications. Sperm selected for ART mostly originates from environment experiencing oxidative stress and high percentage of these sperms may have damaged DNA. If such sperms are used clinically in the form of therapy then can lead to substantial risk in pregnancy outcome. In case of intrauterine insemination (IUI) and in vitro fertilization (IVF), the use of these spermatozoa may not be cause of concern. But in case of intracellular sperm injection (ICSI), this natural selection barrier is bypassed and the spermatozoa with damaged DNA are directly injected into oocytes. Studies have reported that DNA damaged spermatozoa used in ICSI have some capacity for fertilization, but percentage of DNA damage has been negatively correlated to the rate of fertilization [108]. ROS-mediated DNA damage may be linked to an increase in early embryo death, infertility in the offspring, and high incidence of childhood cancer [109, 110]. We propose that extrinsic and intrinsic sources of ROS could make a significant contribution to the induction of OS and DNA damage in spermatozoa which can decrease pregnancy rate and affect the fertil-

ity outcome, further additional studies are clearly needed to validate this concept.

Oxidative stress plays an important role in the pathophysiology of male infertility, which is caused due to pathological level of ROS and the loss of antioxidant protection for the spermatozoa. There are many factors which can induce oxidative stress and can alter seminal parameters and rate of fertilization. Thorough examination and management of some of these factors may protect the ROS-induced DNA damage and improve a couple's chances of con-

Various behaviors and lifestyles factors such as alcohol consumption, cigarette smoking, obesity, excess exposure to environmental toxicants, and psychological stress are negatively correlated with spermatogenesis and may cause oxidative stress and reduction in sperm quality [111]. The increased consumption of simple sugars and high-fat food and physical inactivity are leading causes of the growing obesity. It is suggested that abnormal hormonal regulation, dysregulation of adipocytokine, and ROS generation lead to suboptimal semen quality these patients [112]. Several systemic diseases, such as diabetes mellitus, infection, and cancer are known to cause oxidative stress-induced male infertility [113, 114]. There are studies which have shown positive correlation of exercise with improvements in semen parameters, sperm DNA integrity, and pregnancy rate [115]. Nevertheless, modification in behavior and unhealthy living, regular exercise, stress free jobs, and treatment of a patient's underlying pathology should be the first

steps to reduce or eliminate stress-provoking conditions to reverse sperm dysfunction.

**8. Management of infertility caused by oxidative stress**

ception either naturally or via assisted reproduction.

**8.1. Behavior and life style modification**

80 Novel Prospects in Oxidative and Nitrosative Stress

As many studies suggested that oxidative stress is a major cause of unexplained male infertility, antioxidant therapy would be expected to have a therapeutic effect in such cases. There are evidences which have suggested that oral antioxidants and herbal products can also boost male reproductive functions [116, 117]. But, despite of known effect of antioxidant on oxidative stress, very few studies conducted have any validity due to small sample size, difference in dosage and duration of therapy, and lack controls [118]. In order to make the study valid, patient's selection criteria for the trial should be based on the evidence indication oxidative stress as a key element in their pathology, a thorough diagnosis is required to determine patients that need to be supplemented. However, if this strategy is pursued, great care must be taken in selecting the most appropriate antioxidants for clinical use. Since ROS plays an important role in regulating the signal transduction cascades that drive sperm capacitation, we should ensure that any antioxidants employed in vitro do not compromise the fertilizing potential of these cells [119].

The study of *in vitro* antioxidants is highly relevant in the era of assisted reproduction because sperm preparation techniques in ART are potential generators of exogenous stresses that make human spermatozoa vulnerable to oxidative stress and DNA damage.

### **9. Conclusion**

Oxidative stress has been recognized as a major contributory factor to male infertility. Spermatozoa are professional generator of ROS as physiological level of ROS is necessary to regulate critical redox-sensitive processes such as capacitation, hyperactivation, acrosome reactions, and signaling processes to ensure appropriate fertilization. On the other hand, many endogenous and exogenous factors can elevate ROS production which can overwhelm their antioxidant mechanism. This results in male infertility via mechanisms involving the induction of peroxidative damage to the sperm plasma membrane, DNA damage, which significantly impairs sperm function. Lack of repair mechanism and abortive apoptosis in mature spermatozoa results in high levels of DNA fragmented sperm in the ejaculate. In natural conception, oocytes can repair some of stand DNA breaks, but that depends on the type and level of chromatin damage and the capacity of the oocyte to repair it. If post fertilization oocyte make mistake in the repair process it may lead to failure in fertilization. But if fertilization occurs, then it creates the possibility for *de novo* mutations, which could have a profound impact on the health and well-being of the offspring. When the natural balance between ROS and antioxidants is disturbed, the first restorative measure to be taken should be changes in lifestyle, maintaining a healthy and balanced diet, and antioxidant supplementation may then be taken together to improve the patient's health outcomes.

#### **9.1. Suggestion**

The conventional seminological parameter in infertile cases reflects the functional competence of the spermatozoa and the fertilizing potential of the ejaculate, but the underlying mechanisms of male fertility is not known. Thus in order to enrich the diagnostic value of this fundamental form of investigation, the detailed examination of sperm DNA damage may be incorporated as a potentially valuable tool to investigate the functional integrity of the spermatozoa at the molecular level.

[8] Lavranos G, Balla M, Tzortzopoulou A, Syriou V, Angelopoulou R. Investigating ROS sources in male infertility: A common end for numerous pathways. Reproductive

Role of Reactive Oxygen Species in Male Reproduction http://dx.doi.org/10.5772/intechopen.74763 83

[9] Shiraishi K, Matsuyama H, Takihara H. Pathophysiology of varicocele in male infertility in the era of assisted reproductive technology. International Journal of Urology. 2012

[10] Agarwal A, Virk G, Ong C, du Plessis SS. Effect of oxidative stress on male reproduction.

[11] Borges Jr, E. Total motile sperm count: A better way to rate the severity of male factor infertility? JBRA Assisted Reproduction. 2016;**20**:47-48. DOI: 10.5935/1518-0557.20160012

[12] Agarwal A, Sekhon LH. Oxidative stress and antioxidants for idiopathic Oligoasthenoteratospermia: Is it justified? Indian Journal of Urology. 2011;**27**:74-85. DOI: 10.4103/

[13] Bauché F, Fouchard MH, Jégou B. Antioxidant system in rat testicular cells. FEBS Letters.

[14] Birben E, Sahiner UM, Sackesen C, Erzurum S, Kalayci O. Oxidative stress and antioxidant defense. World Allergy Organization Journal. 2012;**5**:9-19. DOI: 10.1097/

[15] Lee D, Moawad AR, Morielli T, Fernandez MC, O'Flaherty C. Peroxiredoxins prevent oxidative stress during human sperm capacitation. Molecular Human Reproduction.

[16] Baarends WM, van der Laan R, Grootegoed JA. DNA repair mechanisms and gameto-

[17] Aitken RJ. Gpx5 protects the family jewels. Journal of Clinical Investigation.

[18] Aitken RJ, Baker MA, De Iuliis GN, Nixon B. New insights into sperm physiology and pathology. Handbook of Experimental Pharmacology. 2010;**198**:99-115. DOI: 10.1007/

[19] Baker MA, Hetherington L, Weinberg A, Velkov T. Phosphopeptide analysis of rodent epididymal spermatozoa. Journal of Visualized Experiments. 2014;**94**:51546. DOI:

[20] Baker MA, Aitken RJ. Reactive oxygen species in spermatozoa: Methods for monitoring and significance for the origins of genetic disease and infertility. Reproductive Biology

[21] O'Flaherty C. Redox regulation of mammalian sperm capacitation. Asian Journal of

genesis. Reproduction. 2001;**121**:31-39. DOI: 10.1530/rep.0.1210031

and Endocrinology. 2005;**3**:67. DOI: 10.1186/1477-7827-3-67

Andrology. 2015;**17**:583-590. DOI: 10.4103/1008-682X.153303

World Jornal of Mens Health. 2014;**32**:1-17. DOI: 10.5534/wjmh.2014.32.1.1

Toxicology. 2012;**34**:298-307. DOI: 10.1016/j.reprotox.2012.06.007

Jun;**19**:538-550. DOI: 10.1111/j.1442-2042.2012.02982.x

1994;**349**:392-396. DOI: 10.1016/0014-5793(94)00709-8

2017;**23**:106-115. DOI: 10.1093/molehr/gaw081

2009;**119**:1849-1851. DOI: 10.1172/JCI39688

0970-1591.78437

WOX.0b013e3182439613

978-3-642-02062-9\_7

10.3791/51546

### **Author details**

### Sabiha Fatima

Address all correspondence to: sabmehdi@ksu.edu.sa

Department of Clinical Laboratory Sciences, College of Applied Medical Sciences, King Saud University, Riyadh, Saudi Arabia

### **References**


[8] Lavranos G, Balla M, Tzortzopoulou A, Syriou V, Angelopoulou R. Investigating ROS sources in male infertility: A common end for numerous pathways. Reproductive Toxicology. 2012;**34**:298-307. DOI: 10.1016/j.reprotox.2012.06.007

fundamental form of investigation, the detailed examination of sperm DNA damage may be incorporated as a potentially valuable tool to investigate the functional integrity of the sper-

Department of Clinical Laboratory Sciences, College of Applied Medical Sciences, King Saud

[1] Louis JF, Thoma ME, Sorensen DN, McLain AC, King RB, Sundaram R, Keiding N, Buck Louis GM. The prevalence of couple infertility in the United States from a male perspective: Evidence from a nationally representative sample. Andrology. 2013;**1**:741-748. DOI:

[2] Chandra A, Copen CE, Stephen EH. Infertility service use in the United States: Data from the National Survey of family growth, 1982-2010. National Health Statistics Reports.

[3] Catanzariti F, Cantoro U, Lacetera V, Muzzonigro G, Polito M. Comparison between WHO (World Health Organization) 2010 and WHO 1999 parameters for semen analysisinterpretation of 529 consecutive samples. Archivio Italiano di Urologia e Andrologia.

[4] Fassino S, Piero A, Boggio S, Piccioni V, Garzaro L. Anxiety, depression and anger suppression in infertile couples: A controlled study. Human Reproduction. 2002;**17**:2986-

[5] Chachamovich JR, Chachamovich E, Ezer H, Fleck MP, Knauth D, Passos EP. Investigating quality of life and health-related quality of life in infertility: A systematic review. Journal of Psychosomatic Obstetrics and Gynecology. 2010;**31**:101-110. DOI:

[6] Campbell AJ, Irvine DS. Male infertility and intracytoplasmic sperm injection (ICSI).

[7] Jurasović J, Cvitković P, Pizent A, Colak B, Telisman S. Semen quality and reproductive endocrine function with regard to blood cadmium in Croatian male subjects. Biometals.

British Medical Bulletin. 2000;**5**:616-629. DOI: 10.1258/0007142001903427

matozoa at the molecular level.

82 Novel Prospects in Oxidative and Nitrosative Stress

University, Riyadh, Saudi Arabia

10.1111/j.2047-2927.2013.00110.x

2994. DOI: 10.1093/17.11.2986

10.3109/0167482X.2010.4813

2013;**85**:125-129. DOI: 10.4081/aiua.2013.3.125

2004;**17**:735-743. DOI: 10.1007/s10534-004-1689-7

Address all correspondence to: sabmehdi@ksu.edu.sa

**Author details**

Sabiha Fatima

**References**

2014;**73**:1-21


[22] Zhang H, Zheng RL. Promotion of human sperm capacitation by superoxide anion. Free Radical Research. 1996;**24**:261-268. DOI: 10.3109/10715769609088023

[36] Krapf D, Arcelay E, Wertheimer EV, Sanjay A, Pilder SH, Salicioni AM, Visconti PE. Inhibition of Ser/Thr phosphatases induces capacitation-associated signaling in the presence of Src kinase inhibitors. The Journal of biological chemistry. 2010;**285**:7977-

Role of Reactive Oxygen Species in Male Reproduction http://dx.doi.org/10.5772/intechopen.74763 85

[37] Battistone MA, Da Ros VG, Salicioni AM, Navarrete FA, Krapf D, Visconti PE, Cuasnicú PS. Functional human sperm capacitation requires both bicarbonate-dependent PKA activation and down-regulation of Ser/Thr phosphatases by Src family kinases.

[38] Desai N, Sharma R, Makker K, Sabanegh E, Agarwal A. Physiologic and pathologic levels of reactive oxygen species in neat semen of infertile men. Fertility and Sterility.

[39] Avella MA, Dean J. Fertilization with acrosome-reacted mouse sperm: Implications for the site of exocytosis. Proceedings of the National Academy of Sciences of the United

[40] Saldívar-Hernández A, González-González ME, Sánchez-Tusié A, Maldonado-Rosas I, López P, Treviño CL, Larrea F, Chirinos M. Human sperm degradation of zona pellucida proteins contributes to fertilization. Reproductive Biology and Endocrinology.

[41] O'Donnell L. Mechanisms of spermiogenesis and spermiation and how they are disturbed. Spermatogenesis. 2015;**4**:e979623. DOI: 10.4161/21565562.2014.979623

[42] Iwasaki A, Gagnon C. Formation of reactive oxygen species in spermatozoa of infertile patients. Fertility and Sterility. 1992;**57**:409-416. DOI: 10.1016/S0015-0282(16)54855-9 [43] Zini A, de Lamirande E, Gagnon C. Reactive oxygen species in the semen of infertile patients: Levels of superoxide dismutase and catalase-like activities in seminal plasma. International Journal of Andrology. 1993;**16**:183-188. DOI: 10.1111/j.1365-2605.1993.

[44] Saleh RA, Agarwal A. Oxidative stress and male infertility: From research bench to clinical practice. Journal of Andrology. 2002;**23**:737-752. DOI: 10.1002/j.1939-4640.2002.

[45] Sotomayor RE, Sega GA. Unscheduled DNA synthesis assay in mammalian spermatogenic cells: An update. Environmental and molecular mutagenesis. 2000;**36**:255-265.

[46] Gunes S, Al-Sadaan M, Agarwal A. Spermatogenesis, DNA damage and DNA repair mechanisms in male infertility. Reproductive Biomedicine online. 2015;**31**:309-319. DOI:

[47] Ollero M, Powers RD, Alvarez JG. Variation of docosahexaenoic acid content in subsets of human spermatozoa at different stages of maturation: Implications for sperm lipoperoxidative damage. Molecular Reproduction and Development. 2000;**55**:326-334. DOI:

DOI: 10.1002/1098-2280(2000)36:4<255::AID-EM1>3.0.CO;2-O

10.1002/(SICI)1098-2795(200003)55:3<326::AID-MRD11>3.3.CO;2-1

States of America. 2011;**108**:19843-19844. DOI: 10.1073/pnas.1118234109

Molecular Human Reproduction. 2013;**19**:570-580. DOI: 10.1093/molehr/gat033

2009;**92**:1626-1631. DOI: 10.1016/j.fertnstert.2008.08.109

2015;**13**:99. DOI: 10.1186/s12958-015-0094-0

tb01177.x

tb02324.x

10.1016/j.rbmo.2015.06.010

7985. DOI: 10.1074/jbc.M109.085845


[36] Krapf D, Arcelay E, Wertheimer EV, Sanjay A, Pilder SH, Salicioni AM, Visconti PE. Inhibition of Ser/Thr phosphatases induces capacitation-associated signaling in the presence of Src kinase inhibitors. The Journal of biological chemistry. 2010;**285**:7977- 7985. DOI: 10.1074/jbc.M109.085845

[22] Zhang H, Zheng RL. Promotion of human sperm capacitation by superoxide anion. Free

[23] Gavella M, Lipovac V. NADH-dependent oxidoreductase (diaphorase) activity and isozyme pattern of sperm in infertile men. Archives of Andrology. 1992;**28**:135-141. DOI:

[24] Aprioku JS. Pharmacology of free radicals and the impact of reactive oxygen species on

[25] deLamirande E, Gagnon C. Human sperm hyperactivation and capacitation as parts of an oxidative process. Free Radical Biology & Medicine. 1993;**14**:157-166. DOI: 10.1016/

[26] deLamirande E, Gagnon C. Capacitation-associated production of superoxide anion by human spermatozoa. Free Radical Biology & Medicine. 1995;**18**:487-195. DOI:

[27] Musset B, Clark RA, DeCoursey TE, Petheo GL, Geiszt M, Chen Y, Cornell JE, Eddy CA, Brzyski RG, El Jamali A. NOX5 in human spermatozoa: Expression, function, and regulation. Journal of Biological Chemistry. 2012;**287**:9376-9388. DOI: 10.1074/jbc.M111. 314955

[28] Andreyev AY, Kushnareva YE, Starkov AA. Mitochondrial metabolism of reactive oxygen species. Biochemistry (Mosc). 2005;**70**:200-214. DOI: 10.1007/s10541-005-0102-7 [29] Amaral A, Lourenço B, Marques M, Ramalho-Santos J. Mitochondria functionality and sperm quality. Reproduction. 2013;**146**:R163-R174. DOI: 10.1530/REP-13-0178

[30] deLamirande E, Lamothe G, Villemure M. Control of superoxide and nitric oxide formation during human sperm capacitation. Free Radical Biology & Medicine. 2009;**46**:1420-

[31] Aitken RJ. Reactive oxygen species as mediators of sperm capacitation and pathological damage. Molecular reproduction and development. 2017;**84**:1039-1052. DOI: 10.1002/

[32] Baker MA, Hetherington L, Curry B, Aitken RJ. Phosphorylation and consequent stimulation of the tyrosine kinase cAbl by PKA in mouse spermatozoa; its implications during capacitation. Developments in Biologicals. 2009;**33**:57-66. DOI: 10.1016/j.

[33] Herrero MB, Gagnon C. Nitric oxide: A novel mediator of sperm function. Journal of

[34] Ecroyd HW, Jones RC, Aitken RJ. Endogenous redox activity in mouse spermatozoa and its role in regulating the tyrosine phosphorylation events associated with sperm capacitation. Biology of Reproduction. 2003;**69**:347-354. DOI: 10.1095/biolreprod.102.012716 [35] Visconti PE. Understanding the molecular basis of sperm capacitation through kinase design. The molecular basis of sperm capacitation through kinase design. Proceedings of the National Academy of Sciences of the United States of America. 2009;**106**:667-668.

Andrology. 2001;**22**:349-356. DOI: 10.1002/j.1939-4640.2001.tb02188.x

Radical Research. 1996;**24**:261-268. DOI: 10.3109/10715769609088023

the testis. Journal of Reproduction & Infertility. 2013;**14**:158-172

10.3109/ 01485019208987691

84 Novel Prospects in Oxidative and Nitrosative Stress

0891-5849(93)90006-G

mrd. 22871

ydbio.2009.06.022

DOI: 10.1073/pnas.0811895106

10.1016/0891-5849(94)00169-K

1427. DOI: 10.1016/j.freeradbiomed.2009.02.022


[48] Ollero M, Gil-Guzman E, Lopez MC, Sharma RK, Agarwal A, Larson K, Evenson D, Thomas Jr AJ, Alvarez JG. Characterization of subsets of human spermatozoa at different stages of maturation: implications in the diagnosis and treatment of male infertility. Human Reproduction. 2001;**16**:1912-1921. DOI: 10.1093/humrep/16.9.1912

[60] Mruk DD, Cheng CY. In vitro regulation of extracellular superoxide dismutase in sertoli

Role of Reactive Oxygen Species in Male Reproduction http://dx.doi.org/10.5772/intechopen.74763 87

[61] Imai H, Nakagawa Y. Biological significance of phospholipid hydroperoxide glutathione peroxidase (PHGPx, GPx4) in mammalian cells. Free Radical Biology and Medicine.

[62] O'Flaherty C. Peroxiredoxins: Hidden players in the antioxidant defence of human spermatozoa. Basic and Clinical Andrology. 2014;**24**:4. DOI: 10.1186/2051-4190-24-4

[63] Tremellen K. Oxidative stress and male infertility – A clinical perspective. Human

[64] Walczak–Jedrzejowska R, Wolski JK, Slowikowska–Hilczer J. The role of oxidative stress and antioxidants in male fertility. Central European Journal of Urology. 2013;**66**:60-67.

[65] Atig F, Raffa M, Habib BA, Kerkeni A, Saad A, Ajina M. Impact of seminal trace element and glutathione levels on semen quality of Tunisian infertile men. BMC Urology.

[66] Micheli L, Cerretani D, Collodel G, Menchiari A, Moltoni L, Fiaschi AI, Moretti E. Evaluation of enzymatic and non-enzymatic antioxidants in seminal plasma of men with genitourinary infections, varicocele and idiopathic infertility. Andrology. 2016;**4**:456-464.

[67] Arata de Bellabarba G, Tortolero I, Villarroel V, Molina CZ, Bellabarba C, Velazquez E. Nonsperm cells in human semen and their relationship with semen parameters.

[68] Patil PS, Humbarwadi RS, Patil AD, Gune AR.Immature germ cells in semen – Correlation with total sperm count and sperm motility. Journal of Cytology. 2013;**30**:185-189. DOI:

[69] Rengan AK, Agarwal A, van der Linde M, du Plessis SS. An investigation of excess residual cytoplasm in human spermatozoa and its distinction from the cytoplasmic droplet. Reproductive Biology and Endocrinology. 2012;**10**:92. DOI: 10.1186/1477-7827-10-92 [70] Gharagozloo P, Aitken RJ. The role of sperm oxidative stress in male infertility and the significance of oral antioxidant therapy. Human Reproduction. 2011;**26**:1628-1640. DOI:

[71] Harlev A, Agarwal A, Gunes SO, Shetty A, du Plessis SS. Smoking and male infertility: An evidence-based review. World Journal of Mens Health. 2015;**33**:143-160. DOI:

[72] Avendaño C, Mata A, Sanchez Sarmiento CA, Doncel GF. Use of laptop computers connected to internet through Wi-Fi decreases human sperm motility and increases sperm DNA fragmentation. Fertility and Sterility. 2012;**97**:39.e2-45.e2. DOI: 10.1016/j.

Archives of Andrology. 2000;**45**:131-136. DOI: 10.1080/01485010050193896

Reproduction Update. 2008;**14**:243-258. DOI: 10.1093/humupd/dmn004

cells. Life Sciences. 2000;**67**(2):133-145. DOI: 10.1016/S0024-3205(00)00609-3

2003;**34**:145-169. DOI: 10.1016/S0891-5849(02)01197-8

DOI: 10.5173/ceju.2013.01.art19

DOI: 10.1111/andr.12181

10.4103/0970-9371.117682

10.1093/humrep/der132

fertnstert.2011.10.012

10.5534/wjmh. 2015.33.3.143

2012;**12**:6. DOI: 10.1186/1471-2490-12-6


[60] Mruk DD, Cheng CY. In vitro regulation of extracellular superoxide dismutase in sertoli cells. Life Sciences. 2000;**67**(2):133-145. DOI: 10.1016/S0024-3205(00)00609-3

[48] Ollero M, Gil-Guzman E, Lopez MC, Sharma RK, Agarwal A, Larson K, Evenson D, Thomas Jr AJ, Alvarez JG. Characterization of subsets of human spermatozoa at different stages of maturation: implications in the diagnosis and treatment of male infertility.

[49] Aitken RJ, Curry BJ. Redox regulation of human sperm function: From the physiological control of sperm capacitation to the etiology of infertility and DNA damage in the germ line. Antioxidants and Redox signaling. 2011;**14**:367-381. DOI: 10.1089/ars.2010.3186

[50] Saalu LC. The incriminating role of reactive oxygen species in idiopathic male infertility: An evidence based evaluation. Pakistan Journal of Biological Sciences. 2010;**13**:413-422.

[51] Güney AI, Javadova D, Kırac D, Ulucan K, Koc G, Ergec D, Tavukcu H, Tarcan T. Detection of Y chromosome microdeletions and mitochondrial DNA mutations in male infertility

patients. Genetics and Molecular Research. 2012;**11**:1039-1048. DOI: 10.4238/2012

of Urology. 2008;**24**:150-154. DOI: 10.4103/0970-1591.40606

[52] Shamsi MB, Kumar R, Bhatt A, Bamezai RN, Kumar R, Gupta NP, Das TK, Dada R. Mitochondrial DNA mutations in etiopathogenesis of male infertility. Indian Journal

[53] Gil-Guzman E, Ollero M, Lopez MC, Sharma RK, Alvarez JG, Thomas AJ Jr, Agarwal A. Differential production of reactive oxygen species by subsets of human spermatozoa at different stages of maturation. Human Reproduction. 2001;**16**:1922-1930. DOI:

[54] Rhemrev JP, van Overveld FW, Haenen GR, Teerlink T, Bast A, and Vermeiden JP. Quantification of the nonenzymatic fast and slow TRAP in a postaddition assay in human seminal plasma and the antioxidant contributions of various seminal compounds. Journal of Andrology. 2000;**21**:913-920. DOI: 10.1002/j.1939-4640.2000.tb03422.x

[55] Vernet P, Aitken RJ, Drevet JR. Antioxidant strategies in the epididymis. Molecular and

[56] Marchetti F, Wyrobek AJ. Mechanisms and consequences of paternally-transmitted chromosomal abnormalities. Birth Defects Research Part C: Embryo Today. 2005;**75**:112-

[57] Fujii J, Iuchi Y, Matsuki S, Ishii T. Cooperative function of antioxidant and redox systems against oxidative stress in male reproductive tissues. Asian Journal of Andrology.

[58] Doshi SB, Khullar K, Sharma RK, Agarwal A. Role of reactive nitrogen species in male infertility. Reproductive Biology and Endocrinology. 2012;**10**:109. DOI: 10.1186/

[59] Potts RJ, Notarianni LJ, Jefferies TM. Seminal plasma reduces exogenous oxidative damage to human sperm, determined by the measurement of DNA strand breaks and lipid peroxidation. Mutation Research. 2000;**447**:249-256. DOI: 10.1016/S0027-5107(99) 00215-8

Cellular Endocrinology. 2004;**216**:31-39. DOI: 10.1016/j.mce.2003.10.069

Human Reproduction. 2001;**16**:1912-1921. DOI: 10.1093/humrep/16.9.1912

DOI: 10.3923/pjbs.2010.413.422

86 Novel Prospects in Oxidative and Nitrosative Stress

10.1093/humrep/16.9.1922

129. DOI: 10.1002/bdrc.20040

1477-7827-10-109

2003;**5**:231-242. DOI: 10.1.1.526.8507


[73] Saleh RA, Agarwal A, Sharma RK, Nelson DR, Thomas AJ Jr. Effect of cigarette smoking on levels of seminal oxidative stress in infertile men: A prospective study. Fertility and Sterility. 2002;**78**:491-499. DOI: 10.1016/S0015-0282(02)03294-6

[85] Khosrowbeygi A, Zarghami N. Fatty acid composition of human spermatozoa and seminal plasma levels of oxidative stress biomarkers in subfertile males. Prostaglandins Leukotrienes, and Essential Fatty Acids. 2007;**77**:117-121. DOI: 10.1016/j.plefa.2007.08.003

Role of Reactive Oxygen Species in Male Reproduction http://dx.doi.org/10.5772/intechopen.74763 89

[86] Aitken RJ, Whiting S, De Iuliis GN, McClymont S, Mitchell LA, Baker MA. Electrophilic aldehydes generated by sperm metabolism activate mitochondrial reactive oxygen species generation and apoptosis by targeting succinate dehydrogenase. The Journal of

[87] Zhong H, Yin H. Role of lipid peroxidation derived 4-hydroxynonenal (4-HNE) in cancer: Focusing on mitochondria. Redox Biology. 2015;**4**:193-199. DOI: 10.1016/j.redox.

[88] Aitken RJ, Jones KT, Robertson SA. Reactive oxygen species and sperm function--in sickness and in health. Journal of Andrology. 2012;**33**:1096-1106. DOI: 10.2164/jandrol.112.

[89] Gomez E, Buckingham DW, Brindle J, Lanzafame F, Irvine DS, Aitken RJ. Development of an image analysis system to monitor the retention of residual cytoplasm by human spermatozoa, correlation with biochemical markers of the cytoplasmic space, oxidative stress and sperm function. Journal of Andrology. 1996;**17**:276-287. DOI: 10.1002/j.1939-

[90] Sanocka D, Miesel R, Jedrzejczak P, Chełmonska-Soyta AC, Kurpisz M. Effect of reactive oxygen species and the activity of antioxidant systems on human semen; association with male infertility. International Journal of Andrology. 1997;**20**:255-264. DOI:

[91] Bansal AK, Bilaspuri GS. Impacts of oxidative stress and antioxidants on semen functions. Veterinary Medicine InternationalVeterinary Medicine International. Sep 7,

[92] Wang X, Sharma R K, Gupta A, George V, Thomas A J, Falcone T, Agarwal A. Alterations in mitochondria membrane potential and oxidative stress in infertile men: A prospective observational study. Fertility and Sterility. 2003;**80**:844-850. DOI: 10.1016/

[93] Sawyer DE, Roman SD, Aitken RJ. Relative susceptibilities of mitochondrial and nuclear DNA to damage induced by hydrogen peroxide in two mouse germ cell lines. Redox

[94] Sawyer DE, Mercer BG, Wiklendt AM, Aitken RJ. Quantitative analysis of gene-specific DNA damage in human spermatozoa. Mutation Research. 2003;**529**:21-34. DOI: 10.1016/

[95] Bennetts LE, Aitken RJ. A comparative study of oxidative DNA damage in mammalian spermatozoa. Molecular Reproduction and Development. 2005;**71**:77-87. DOI: 10.1002/

Biological Chemistry. 2012;**287**:33048-33060. DOI: 10.1074/jbc.M112.366690

2014.12.011

4640.1996.tb01783.x

S0015-0282(03)00983-X

S0027-5107(03)00101-5

mrd. 20285

10.1046/j.1365-2605.1997.00050.x

2010;**2010**. pii: 686137. doi: 10.4061/2011/686137

Report. 2001;**6**:182-184. DOI: 10.1179/135100001101536157

016535


[85] Khosrowbeygi A, Zarghami N. Fatty acid composition of human spermatozoa and seminal plasma levels of oxidative stress biomarkers in subfertile males. Prostaglandins Leukotrienes, and Essential Fatty Acids. 2007;**77**:117-121. DOI: 10.1016/j.plefa.2007.08.003

[73] Saleh RA, Agarwal A, Sharma RK, Nelson DR, Thomas AJ Jr. Effect of cigarette smoking on levels of seminal oxidative stress in infertile men: A prospective study. Fertility and

[74] Koppers AJ, De Iuliis GN, Finnie JM, McLaughlin EA, Aitken RJ. Significance of mitochondrial reactive oxygen species in the generation of oxidative stress in spermatozoa. The Journal of Clinical Endocrinology and Metabolism. 2008;**93**:3199-3207. DOI: 10.1210/

[75] Aitken RJ, Fisher HM, Fulton N, Gomez E, Knox W, Lewis B, Irvine S. Reactive oxygen species generation by human spermatozoa is induced by exogenous NADPH and inhibited by the flavoprotein inhibitors diphenyleneiodonium and quinacrine. Molecular Reproduction and Development. 1997;**47**:468-482. DOI: 10.1002/

[76] Fisher HM, Aitken RJ. Comparative analysis of the ability of precursor germ cells and epididymal spermatozoa to generate reactive oxygen metabolites. The Journal of Experimental Zoology. 1997;**277**:390-400. DOI: 10.1002/(SICI)1097-010X(19970401)

[77] Shukla S, Jha RK, Laloraya M, Kumar PG. Identification of non-mitochondrial NADPH oxidase and the spatio-temporal organization of its components in mouse spermatozoa. Biochemical and Biophysical Research Communications. 2005;**331**:476-483. DOI:

[78] Plante M, de Lamirande E, Gagnon C. Reactive oxygen species released by activated neutrophils, but not by deficient spermatozoa, are sufficient to affect normal sperm motility. Fertility and Sterility. 1994;**62**:387-393. DOI: 10.1016/S0015-0282(16)56895-2 [79] Ramya T, Misro MM, Sinha D, Nandan D. Sperm function and seminal oxidative stress as tools to identify sperm pathologies in infertile men. Fertility and Sterility. 2010;**93**:297-

[80] Wolff H. The biologic significance of white blood cells in semen. Fertility and Sterility.

[81] Henkel RR. Leukocytes and oxidative stress: Dilemma for sperm function and male fer-

[82] Gavriliouk D, Aitken RJ. Damage to sperm DNA mediated by reactive oxygen species: Its impact on human reproduction and the health trajectory of offspring. Advances in Experimental Medicine nad Biology. 2015;**868**:23-47. DOI: 10.1007/978-3-319-18881-2\_2 [83] Sikka SC. Relative impact of oxidative stress on male reproductive function. Current

[84] Moazamian R, Polhemus A, Connaughton H, Fraser B, Whiting S, Gharagozloo P, Aitken RJ. Oxidative stress and human spermatozoa: Diagnostic and functional significance of aldehydes generated as a result of lipid peroxidation. Molecular Human Reproduction.

tility. Asian Journal of Andrology. 2011;**13**:43-52. DOI: 10.1038/aja.2010.76

Medicinal Chemistry. 2001;**8**:851-862. DOI: 10.2174/0929867013373039

Sterility. 2002;**78**:491-499. DOI: 10.1016/S0015-0282(02)03294-6

(SICI)1098-2795(199708)47:4<468::AID-MRD14>3.0.CO;2-S

277:5<390::AID-JEZ53.0.CO;2-K>

300. DOI: 10.1016/j.fertnstert.2009.05.074

2015;**2**:502-515. DOI: 10.1093/molehr/gav014

1995;**63**:1143-1157. DOI: 10.1016/S0015-0282(16)57588-8

10.1016/j.bbrc.2005.03.198.

jc.2007-2616

88 Novel Prospects in Oxidative and Nitrosative Stress


[96] Marcon L, Boissonneault G. Transient DNA strand breaks during mouse and human spermiogenesis new insights in stage specificity and link to chromatin remodeling. Biology of Reproduction. 2004;**70**:910-918. DOI: 10.1095/biolreprod.103.022541

[109] Hazout A, Menezo Y, Madelenat P, Yazbeck C, Selva J, Cohen-Bacrie P. Causes and clinical implications of sperm DNA damages. Gynécologie, obstétrique and fertilité.

Role of Reactive Oxygen Species in Male Reproduction http://dx.doi.org/10.5772/intechopen.74763 91

[110] Chen SJ, Allam JP, Duan YG, Haidl G. Influence of reactive oxygen species on human sperm, functions and fertilizing capacity including therapeutical approaches. Archives of Gynecology and Obstetrics. 2013;**288**:191-199. DOI: 10.1007/s00404-013-2801-4 [111] Sharma R, Biedenharn KR, Fedor JM, Agarwal A. Lifestyle factors and reproductive health: taking control of your fertility. Reproductive Biology and Endocrinology.

[112] Furukawa S, Fujita T, Shimabukuro M, Iwaki M, Yamada Y, Nakajima Y, Nakayama O, Makishima M, Matsuda M, Shimomura I. Increased oxidative stress in obesity and its impact on metabolic syndrome. The Journal of Clinical Investigation. 2004;**114**:1752-

[113] La Vignera S, Condorelli R, Vicari E, D'Agata R, Calogero AE. Diabetes mellitus and sperm parameters. Journal of Andrology. 2012;**33**:145-153. DOI: 10.2164/

[114] Kumar K, Lewis S, Vinci S, Riera-Escamilla A, Fino MG, Tamburrino L, Muratori M, Larsen P, Krausz C. Evaluation of sperm DNA quality in men presenting with testicular cancer and lymphoma using alkaline and neutral comet assays. Andrology. 2018;**6**:

[115] Hajizadeh Maleki B, Tartibian B. Combined aerobic and resistance exercise training for improving reproductive function in infertile men: A randomized controlled trial. Applied Physiology, Nutrition, and Metabolism. 2017;**42**:1293-1306. DOI: 10.1139/

[116] Gvozdjáková A, Kucharská J, Dubravicky J, Mojto V, Singh RB. Coenzyme Q10, α-tocopherol, and oxidative stress could be important metabolic biomarkers of male

[117] Adewoyin M, Ibrahim M, Roszaman R, Isa MLM, Alewi NAM, Rafa AAA, Anuar MNN Male infertility: The effect of natural antioxidants and phytocompounds on seminal

[118] Zini A, Al-Hathal N. Antioxidant therapy in male infertility: Fact or fiction? Asian

[119] Agarwal A, Nallella KP, Allamaneni SS, Said TM. Role of antioxidants in treatment of male infertility: An overview of the literature. Reproductive Biomedicine Online.

infertility. Disease Markers. 2015;**2015**:827941. DOI: 10.1155/2015/827941

oxidative stress. Diseases. 2017;**5**:pii: E9. DOI: 10.3390/diseases5010009

Journal of Andrology. 2011;**13**:374-381. DOI: 10.1038/aja.2010.182

2004;**8**:616-627. DOI: 10.1016/S1472-6483(10)61641-0

2008;**36**:1109-1117. DOI: 10.1016/j.gyobfe.2008.07.017

2013;**11**:66. DOI: 10.1186/1477-7827-11-66

1761. DOI: 10.1172/JCI21625

230-235. DOI: 10.1111/andr.12429

jandrol.111.013193

apnm-2017-0249


[109] Hazout A, Menezo Y, Madelenat P, Yazbeck C, Selva J, Cohen-Bacrie P. Causes and clinical implications of sperm DNA damages. Gynécologie, obstétrique and fertilité. 2008;**36**:1109-1117. DOI: 10.1016/j.gyobfe.2008.07.017

[96] Marcon L, Boissonneault G. Transient DNA strand breaks during mouse and human spermiogenesis new insights in stage specificity and link to chromatin remodeling.

[98] Meyer-Ficca ML, Lonchar JD, Ihara M, Bader JJ, Meyer RG. Alteration of poly (ADPribose) metabolism affects murine sperm nuclear architecture by impairing pericentric heterochromatin condensation. Chromosoma. 2013;**122**:319-335. DOI: 10.1007/s00412-

[99] Leduc F, Maquennehan V, Nkoma GB, Boissonneault G. DNA damage response during chromatin remodeling in elongating spermatids of mice. Biology of Reproduction.

[100] Aitken RJ, Bronson R, Smith TB, De Iuliis GN. The source and significance of DNA damage in human spermatozoa; acommentary on diagnostic strategies and straw man fallacie. Molecular Human Reproduction. 2013;**19**:475-485. DOI: 10.1093/molehr/gat025

[101] Simon L, Castillo J, Oliva R, Lewis SE. Relationships between human sperm protamines, DNA damage and assisted reproduction outcomes. Reproductive Biomedicine Online.

[102] Milligan JR, Aguilera JA, Nguyen JV, Ward JF. Redox reactivity of guanyl radicals in plasmid DNA. International Journal of Radiation Biology. 2001;**77**:281-293. DOI:

[103] Smith TB, Dun MD, Smith ND, Curry BJ, Connaughton HS, Aitken RJ. The presence of a truncated base excision repair pathway in human spermatozoa that is mediated by

[104] Kodama H, Yamaguchi R, Fukuda J, Kasai H, Tanaka T. Increased oxidative deoxyribonucleic acid damage in the spermatozoa of in fertile male patients. Fertility and

[105] Sakkas D, Seli E, Manicardi GC, Nijs M, Ombelet W, Bizzaro D. The presence of abnormal spermatozoa in the ejaculate: Did apoptosis fail? Human Fertility (Cambridge,

[106] Ahmadi A, Ng SC. Fertilizing ability of DNA-damaged spermatozoa. Journal of Experimental Zoology. 1999;**284**:696-704. DOI: 10.1002/(SICI)1097-010X(19991101)284:6

[107] González-Marín C, Gosálvez J, Roy R. Types, causes, detection and repair of DNA fragmentation in animal and human sperm cells. International Journal of Molecular

[108] Merchant R, Gandhi G, Allahbadia GN. In vitro fertilization/intracytoplasmic sperm injection for male infertility. Indian Journal of Urology. 2011;**27**:121-132. DOI: 10.4103/

OGG1. Journal of Cell Science. 2013;**126**:1488-1497. DOI: 10.1242/jcs.121657

Sterility. 1997;**68**:519-524. DOI: 10.1016/S0015-0282(97)00236-7

England). 2004;**7**:99-103. DOI:10.1080/14647270410001720464

Sciences. 2012;**13**:14026-14052. DOI: 10.3390/ijms131114026

2008;**78**:324-332. DOI: 10.1095/biolreprod.107.064162

201;**23**:724-734. DOI: 10.1016/j.rbmo.2011.08.010

1080/095530000100.13436

<696::AID-JEZ11>3.0.CO;2-E

0970-1591.78430

Biology of Reproduction. 2004;**70**:910-918. DOI: 10.1095/biolreprod.103.022541 [97] Laberge RM, Boissonneault G. On the nature and origin of DNA strand breaks in elongating spermatids. Biology of Reproduction. 2005;**73**:289-296. DOI: 10.1095/biolreprod.

104.036939

90 Novel Prospects in Oxidative and Nitrosative Stress

013-0416-y


**Chapter 5**

**Provisional chapter**

**Particularities of Oxidative Stress in Newborns**

**Particularities of Oxidative Stress in Newborns**

DOI: 10.5772/intechopen.73369

The oxidative stress at newborns is augmented by different conditions like preterm birth, asphyxia, respiratory distress, and intraventricular hemorrhage. Preterm neonates associate a more pronounced oxidative stress than healthy term newborns. Several neonatal conditions like respiratory distress (RDS), asphyxia, intraventricular hemorrhage, bronchopulmonary dysplasia, retinopathy, and necrotizing enterocolitis will increase the oxidative stress. The harmful effects of free radicals are linked to their capacity to react with polyunsaturated fatty acids of cell membranes, proteins, and nucleic acids. Free radicals

Oxidative stress represents all injuries caused by reactive oxygen species (ROS) on biomolecules, inducing the destruction of membranes, enzymes, receptors, as well as alteration of cell function. The consequence of oxidative stress is a disruption of the physiological balance

Newborn possesses defense mechanisms such us molecules protection, limitation of ROS production, and mechanisms for repair and adaptation to endogenous and exogenous ROS

Reactive oxygen species are involved in physiological processes such as physical exercise, hyperbarism, regulation of vascular tone, stimulation of cell growth and proliferation, stimulation of erythropoietin secretion, the learning and memory process, as well as in pathological

will produce protein alteration with function loss and lipid peroxidation.

**Keywords:** oxidative stress, new born, prematurity

between pro-oxidants and antioxidants [1, 2].

processes: inflammation, aging, carcinogenesis [1–3].

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution,

© 2018 The Author(s). Licensee IntechOpen. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use,

distribution, and reproduction in any medium, provided the original work is properly cited.

and reproduction in any medium, provided the original work is properly cited.

Melinda Mátyás and Gabriela Zaharie

Melinda Mátyás and Gabriela Zaharie

http://dx.doi.org/10.5772/intechopen.73369

**Abstract**

**1. Introduction**

overproduction.

Additional information is available at the end of the chapter

Additional information is available at the end of the chapter

**Provisional chapter**
