**1. Introduction**

Oxidative stress is defined as an imbalance between oxidants and antioxidants leading to excessive levels of reactive oxygen species (ROS). Oxidative damage includes oxidative modification of cellular macromolecules, cell death by apoptosis or necrosis, as well as structural tissue damage. DNA, proteins and lipids, are the natural targets of oxidation [1].

In recent years, ROS have gained more attention, because of their central role in the progression of many inflammatory diseases [2]. The radical groups include hydroxyl radical (OH**.** ), nitric oxide (NO**.** ) and superoxide (O2 **.−**). Non-radical compounds can also be highly reactive, which includes peroxynitrite (ONOO<sup>−</sup> ), hydrogen peroxide (H2O2) and hypochlorous acid (HOCl) [3]. ROS can be described as oxygen free radicals and other non-radical oxygen derivatives involved in oxygen radical production [4], which are generated by the cells in most tissues and involved in normal cellular metabolism [5].

ROS can react with DNA and cause damage to purines and pyrimidines [6], resulting in the formation of 8-Hydroxy-deoxyguanosine (8-OHdG) [7]. ROS also cause polypeptide chain fragmentation and covalent crosslinking that results in changes in its protein functional activity [8]. The covalent modifications of proteins induced by ROS or by reacting with secondary products of oxidative stress is termed as protein oxidation. These changes lead to many consequences such as, inhibition of enzyme activity, binding activity, aggregation, proteolysis, increased or decreased cell uptake, altered immunogenicity.

Protein oxidation serves as marker for determination of the levels of OS *in vitro*. There are many mechanisms that can induce protein oxidation as all the amino acyl side chains can be oxidatively modified. Cysteine and Methionine are the two amino acids that are most susceptible to oxidative attack due to the presence of sulfur atoms. Oxidation of cysteine leads to the formation of disulfide bonds, mixed disulfides and thiyl radicals, whereas modification of Methionine produces Methionine sulfoxide [8].

Lipids are important constituents of the lipid bilayer of the cellular membrane. Unsaturated fatty acids, which are easily oxidized, initiate the chain reactions, resulting in further oxidative damage. Lipids are susceptible to oxidation and reacts with molecular oxygen to form lipid peroxyl radicals which further oxidizes the neighboring lipids and propagates the oxidative damage [3]. Lipid peroxidation results in the changes of structural integrity and functioning of cell membranes. Lipid peroxidation markers such as malondialdehyde (MDA), 4-hydroxyl-2-nonenal (HNE), and isoprostane are used to evaluate oxidative damage.

Another category of substances called antioxidants exist in the cells and can effectively delay or inhibit ROS-induced oxidation. Antioxidants present in erythrocytes can be broadly classified into enzymatic and non-enzymatic antioxidants. There are three main groups of enzymatic antioxidants that play a significant role in protecting the cells from OS. Superoxide Dismutase (SOD) catalyzes the conversion of superoxides into hydrogen peroxide (H2O2) & oxygen (O2). H2O2 is less toxic than compared to superoxides. SOD are metal containing enzymes that depend on a bound Mg, Cu or Zn for their antioxidant activity. There are 3 major families of SOD: Cu/Zn SOD, Fe/Mn SOD and Ni SOD [9–11]. Catalase (CAT) is the most common enzyme that is found in nearly all living organisms. It is found in the peroxisome of eukaryotic cells. It degrades H2O2 to H2O & O2. Hence it finishes the detoxification reaction started by SOD [12]. Glutathione peroxidase (GPx) contain Selenium, with peroxidase activity whose main activity is to protect the organism against OS. These enzymes like catalase degrade H2O2 to H2O. They also reduce organic peroxides to their corresponding alcohol, thus provides another route for detoxification [13].

The extracellular endogenous antioxidants generally include the transition metal binding proteins i.e. ceruloplasmin, transferrin, hepatoglobin and albumin [14] and Vitamin C, α-tocopherol and Glutathione [15].


**35**

[22]. O2

patients [35–44].

*Modulations in Oxidative Stress of Erythrocytes during Bacterial and Viral Infections*

the C-2 and C-3 double bond to act as antioxidants that result in the formation of semi dehydroascorbic acid, an intermediate free radical. This then reduces to a neutral ascorbate molecule [14]. In cells, ascorbic acid is maintained in its reduced form by reacting with glutathione, catalyzed by protein disulfide

c.Vitamin E is a collective name that is given for a set of eight related tocopherols and tocotrienols that are fat soluble. α-tocopherol is the most commonly occurring natural antioxidant and has a phytyl chain that is attached to its chromanol nucleus [14]. It is a lipid soluble antioxidant and acts in the glutathione peroxidase pathway [19]. It protects the cell membranes against lipid peroxidation chain reaction by reacting with the lipid radicals produced due to OS. It removes the free radical intermediates, thus preventing the cascade and

Bacterial infections cause OS, which trigger ROS production leading to organ

Periodontitis is a common bacterial infection, caused *Fusobacterium nucleatum,* resulting in the destruction of teeth supporting tissues. Periodontitis is associated with overproduction of ROS by neutrophils. It is characterized by increased

way called "respiratory burst" catalyzed by NADPH oxidase during phagocytosis

and non-radical derivatives, such as hydrogen peroxide (H2O2), hypochlorous

ies have shown that not only neutrophils, but also other phagocytes and cells of periodontal tissues such as monocytes, gingival fibroblast and periodontal ligament cells exhibit enhanced ROS production upon stimulation by periodontal pathogens and their components [23–25]. This was evident in the results of lipid peroxidation and protein oxidation products. Higher levels of TBARS, MDA and 8-isoprostanes were found in blood plasma, erythrocytes, gingival crevicular fluid (GCF) [26], saliva, GCF [27–30] respectively in periodontitis patients compared to healthy controls. Higher levels of protein carbonyl groups (PC) were found in GCF, saliva and serum of periodontitis patients [31–34] and 8-Hydroxy-deoxyguanosine (8-OHdG) in GCF and saliva of periodontitis

Variations in antioxidant enzymes were also observed in many studies. SOD, CAT, GPx and glutathione reductase activities decreased in saliva of periodontitis patients [36, 45]. However, SOD and CAT in plasma, erythrocytes and gingival tissues were elevated, whereas activities of non-enzymatic antioxidants (vitamins E, vitamin C, and reduced glutathione) decreased in periodontitis [26]. Periodontitis

Mycobacteria initiate infection at oxygen rich lung microenvironments, generating oxidative radicals. These toxic radicals kill the pathogens by causing

is associated with decreased Total antioxidant capacity (TAC) [46–52].

*1.1.2 Tuberculosis- mycobacterium tuberculosis*

**.−**can be released into phagosome and gets converted to different radical

) and singlet oxygen (1O2). *In vitro* stud-

**.−** via the metabolic path-

metabolites of lipid peroxidation, DNA damage and protein damage.

After pathogen stimulation, neutrophils produce O2

*DOI: http://dx.doi.org/10.5772/intechopen.98236*

isomerase and glutaredoxins [17].

**1.1 Bacterial and viral and infections cause OS**

damage by altering the metabolic pathway [21].

*1.1.1 Periodontitis-*Fusobacterium nucleatum

acid (HOCl), hydroxyl radical (OH**.**

further damage [20].

the C-2 and C-3 double bond to act as antioxidants that result in the formation of semi dehydroascorbic acid, an intermediate free radical. This then reduces to a neutral ascorbate molecule [14]. In cells, ascorbic acid is maintained in its reduced form by reacting with glutathione, catalyzed by protein disulfide isomerase and glutaredoxins [17].

c.Vitamin E is a collective name that is given for a set of eight related tocopherols and tocotrienols that are fat soluble. α-tocopherol is the most commonly occurring natural antioxidant and has a phytyl chain that is attached to its chromanol nucleus [14]. It is a lipid soluble antioxidant and acts in the glutathione peroxidase pathway [19]. It protects the cell membranes against lipid peroxidation chain reaction by reacting with the lipid radicals produced due to OS. It removes the free radical intermediates, thus preventing the cascade and further damage [20].

## **1.1 Bacterial and viral and infections cause OS**

Bacterial infections cause OS, which trigger ROS production leading to organ damage by altering the metabolic pathway [21].

#### *1.1.1 Periodontitis-*Fusobacterium nucleatum

Periodontitis is a common bacterial infection, caused *Fusobacterium nucleatum,* resulting in the destruction of teeth supporting tissues. Periodontitis is associated with overproduction of ROS by neutrophils. It is characterized by increased metabolites of lipid peroxidation, DNA damage and protein damage.

After pathogen stimulation, neutrophils produce O2 **.−** via the metabolic pathway called "respiratory burst" catalyzed by NADPH oxidase during phagocytosis [22]. O2 **.−**can be released into phagosome and gets converted to different radical and non-radical derivatives, such as hydrogen peroxide (H2O2), hypochlorous acid (HOCl), hydroxyl radical (OH**.** ) and singlet oxygen (1O2). *In vitro* studies have shown that not only neutrophils, but also other phagocytes and cells of periodontal tissues such as monocytes, gingival fibroblast and periodontal ligament cells exhibit enhanced ROS production upon stimulation by periodontal pathogens and their components [23–25]. This was evident in the results of lipid peroxidation and protein oxidation products. Higher levels of TBARS, MDA and 8-isoprostanes were found in blood plasma, erythrocytes, gingival crevicular fluid (GCF) [26], saliva, GCF [27–30] respectively in periodontitis patients compared to healthy controls. Higher levels of protein carbonyl groups (PC) were found in GCF, saliva and serum of periodontitis patients [31–34] and 8-Hydroxy-deoxyguanosine (8-OHdG) in GCF and saliva of periodontitis patients [35–44].

Variations in antioxidant enzymes were also observed in many studies. SOD, CAT, GPx and glutathione reductase activities decreased in saliva of periodontitis patients [36, 45]. However, SOD and CAT in plasma, erythrocytes and gingival tissues were elevated, whereas activities of non-enzymatic antioxidants (vitamins E, vitamin C, and reduced glutathione) decreased in periodontitis [26]. Periodontitis is associated with decreased Total antioxidant capacity (TAC) [46–52].

#### *1.1.2 Tuberculosis- mycobacterium tuberculosis*

Mycobacteria initiate infection at oxygen rich lung microenvironments, generating oxidative radicals. These toxic radicals kill the pathogens by causing

*Erythrocyte - A Peripheral Biomarker for Infection and Inflammation*

decreased cell uptake, altered immunogenicity.

ROS can react with DNA and cause damage to purines and pyrimidines [6], resulting in the formation of 8-Hydroxy-deoxyguanosine (8-OHdG) [7]. ROS also cause polypeptide chain fragmentation and covalent crosslinking that results in changes in its protein functional activity [8]. The covalent modifications of proteins induced by ROS or by reacting with secondary products of oxidative stress is termed as protein oxidation. These changes lead to many consequences such as, inhibition of enzyme activity, binding activity, aggregation, proteolysis, increased or

Protein oxidation serves as marker for determination of the levels of OS *in vitro*. There are many mechanisms that can induce protein oxidation as all the amino acyl side chains can be oxidatively modified. Cysteine and Methionine are the two amino acids that are most susceptible to oxidative attack due to the presence of sulfur atoms. Oxidation of cysteine leads to the formation of disulfide bonds, mixed disulfides and thiyl radicals, whereas modification of Methionine produces Methionine sulfoxide [8]. Lipids are important constituents of the lipid bilayer of the cellular membrane. Unsaturated fatty acids, which are easily oxidized, initiate the chain reactions, resulting in further oxidative damage. Lipids are susceptible to oxidation and reacts with molecular oxygen to form lipid peroxyl radicals which further oxidizes the neighboring lipids and propagates the oxidative damage [3]. Lipid peroxidation results in the changes of structural integrity and functioning of cell membranes. Lipid peroxidation markers such as malondialdehyde (MDA), 4-hydroxyl-2-none-

Another category of substances called antioxidants exist in the cells and can effectively delay or inhibit ROS-induced oxidation. Antioxidants present in erythrocytes can be broadly classified into enzymatic and non-enzymatic antioxidants. There are three main groups of enzymatic antioxidants that play a significant role in protecting the cells from OS. Superoxide Dismutase (SOD) catalyzes the conversion of superoxides into hydrogen peroxide (H2O2) & oxygen (O2). H2O2 is less toxic than compared to superoxides. SOD are metal containing enzymes that depend on a bound Mg, Cu or Zn for their antioxidant activity. There are 3 major families of SOD: Cu/Zn SOD, Fe/Mn SOD and Ni SOD [9–11]. Catalase (CAT) is the most common enzyme that is found in nearly all living organisms. It is found in the peroxisome of eukaryotic cells. It degrades H2O2 to H2O & O2. Hence it finishes the detoxification reaction started by SOD [12]. Glutathione peroxidase (GPx) contain Selenium, with peroxidase activity whose main activity is to protect the organism against OS. These enzymes like catalase degrade H2O2 to H2O. They also reduce organic peroxides to their corresponding alcohol, thus provides another route for

The extracellular endogenous antioxidants generally include the transition metal binding proteins i.e. ceruloplasmin, transferrin, hepatoglobin and albumin [14] and

a.Glutathione (GSH), a major thiol antioxidant is a multifunctional intracellular antioxidant It is one of the most important cellular antioxidants due to its high concentration and its role in maintaining the redox state of the cell. GSH is a cysteine containing peptide and possesses antioxidant property due to the thiol group that serves as a reducing agent which can be reversibly oxidized and

b.Vitamin C, also known as L-ascorbic acid or ascorbate, is present naturally in the body and interconverts between each other depending on the pH. Ascorbic acid is well known reducing agent, thus serves as a good antioxidant [18]. Vit C is a water-soluble electron donor vitamin as it donates two of its electrons from

nal (HNE), and isoprostane are used to evaluate oxidative damage.

**34**

detoxification [13].

reduced. [16, 17].

Vitamin C, α-tocopherol and Glutathione [15].

disintegration of bacterial cell membrane, DNA damage, deactivation of metabolic enzymes or proteins [53–56]. After invasion into the host, mycobacteria induce NADPH oxidase 2 (NOX2) expression to generate superoxide radicals (O2 **.−**), which are then converted to more toxic hydrogen peroxide (H2O2) by superoxide dismutase (SOD) and subsequently reduced to water and molecular oxygen by catalase [57, 58]. NADPH oxidase 2 (NOX2) is the key enzyme responsible for the cellular ROS production by using superoxide radicals (O2-) as precursor molecule [59]. Alterations in regulatory components of NOX2 results in generation of phagocytic oxidative stress and phagocytic burst to eliminate enclosed pathogen [58].

However, pathogenic mycobacteria can inhibit oxidative stress mechanisms by modulation of cell signaling mechanisms, up-regulation of antioxidant enzymes and redox buffering systems [59–62].

#### *1.1.3 Pneumococcal meningitis-* streptococcus pneumoniae

Pneumococcal meningitis is a life-threatening disease characterized by acute infection affecting the pia mater, arachnoid, and subarachnoid spaces [63]. *Streptococcus pneumoniae* crosses the blood–brain barrier (BBB) and disrupts the intraepithelial tight junctions. Host polymorphonuclear leukocytes produce nitric oxide, superoxide radicals, and hydrogen peroxide in response to bacterial infection. O2 − ∙ and NO∙ can lead to the formation of peroxynitrite (ONOO), a strong oxidant [64–66]. ONOO<sup>−</sup> can damage neurons and glial cells by lipid peroxidation and cell membrane destabilization, resulting in DNA disintegration and subsequent poly (ADP-ribose) polymerase (PARP) activation. Elevated 4-HNE and MDA levels are found in bacterial meningitis patients [67]. Thus, ROS/ RNS can be considered key players of immune activation, blood–brain barrier disruption, vascular failure, neuronal injury, and cochlear damage during pneumococcal meningitis.

#### *1.1.4 Gastritis/gastric cancer-* Helicobacter pylori

*Helicobacter pylori* is the causative pathogen for human gastritis or gastric cancer, which is characterized with inflammation and ulceration of the stomach and duodenum. Gastric cancer arises from oxidative stress and environmental toxins, which increase DNA mutation rates [68]. The possible sources of ROS/ RNS in *H. pylori* infected stomach, include neutrophils, vascular endothelial cells and gastric mucosal cells. Neutrophils are believed to be the main source of ROS/ RNS [69] and their production is catalyzed by NADPH oxidase on the cell membrane [59]. These highly reactive ROS (HOCl and˙OH) are used by the phagocyte to kill pathogenic bacteria. *H. pylori* infected gastric mucosal phagocytes produce greater amounts of ROS, which is believed to be the major cause of gastric mucosal damage.

#### *1.1.5 HIV and Hepatitis*

Oxidative Stress has always played a major pathogenic role in HIV and hepatitis infections. HIV causes decrements in glutathione (GSH), cystine, vitamin C and SOD levels, and increments in lipid peroxidation [70–73]. A decline in the antioxidant capacity represents a weakened immune system, thus requiring more antioxidants to maintain normal functionality [74]. Hepatitis, similar to HIV, also increases the lipid peroxidation (malondialdehyde (MDA) and 4-hydroxynonenal (HNE)) and activity of caspases, whereas reduces zinc [75–77] (**Table 1**).

**37**

**Table 1.**

*Modulations in Oxidative Stress of Erythrocytes during Bacterial and Viral Infections*

Tuberculosis- Increased Sepsis- Increased

Meningitis- Increased

Sepsis- Increased

Sepsis- Increased

Sepsis- Increased

Gastritis- Decreased

Tuberculosis- Decreased

Gastritis- Decreased

GST Paramyxovirus-Decreased

Meningitis- Decreased

Vit-E Covid-19- Decreased

*virus; DENV- Dengue virus; RSV- Respiratory syncytial virus; Covid-19-Coronavirus Disease 2019. TBARS- Thiobarbituric acid reactive substances; MDA- Malondialdehyde; 8-IP- 8-isoprostanes; 4-HNE-4-hydroxynonenal; PC- Protein Carbonyls; 8-OHdG- 8-Hydroxy-deoxyguanosine; NO- nitric oxide; NT-Nitrotyrosine; SOD- Superoxide dismutase; CAT- Catalase; GPx- Glutathione Peroxidase; GSH- Glutathione; GR- Glutathione Reductase, GST- Glutathione S-Transferase; UA- Uric acid; Vit-C- Vitamin C; Vit-E- Vitamin E;* 

*FRAP- ferric reducing antioxidant power; TRAP- Total-trapping radical antioxidant potential.*

GR Periodontitis- Decreased [36, 45]

β-Carotene Covid-19- Decreased [84] Bilurubin, UA, FRAP, TRAP Sepsis- Increased [79] *HBV- Hepatitis B virus; HCV- Hepatitis C virus; HIV- Human immunodeficiency virus; JEV- Japanese encephalitis* 

GSH Sepsis- Decreased HIV- Decreased

8-IP Periodontitis- Increased [26] 4-HNE Meningitis- Increased [80]

8-OHdG Periodontitis- Increased HCV- Increased [35]

**OS Markers Bacterial diseases Viral diseases References**

HBV- Increased HCV- Increased HIV- Increased JEV- Increased RSV- Increased

HBV- Decreased HIV- Decreased JEV- Increased DENV-Decreased RSV- Decreased

HBV- Decreased Paramyxovirus-Decreased DENV-Decreased RSV- Decreased

JEV- Increased

RSV- Decreased

Paramyxovirus-Decreased DENV-Decreased RSV- Decreased

Influenza virus-Decreased

Influenza virus-Decreased HIV- Decreased Covid-19- Decreased

Influenza virus-Decreased

[26] [78] [79]

[26] [80] [81]

[31] [79]

[81]

[82] [81]

[80] [82]

[36, 45] [83] [81]

[36, 45] [78] [81]

[36, 45] [83] [81]

[82] [81]

[81]

[79] [80] [81] [84]

[81] [84]

RSV- Increased [78]

*DOI: http://dx.doi.org/10.5772/intechopen.98236*

TBARS Periodontitis- Increased

MDA Periodontitis- Increased

PC Periodontitis- Increased

NO Tuberculosis- Decreased

NT Meningitis- Increased

SOD Periodontitis- Decreased

CAT Periodontitis- Decreased

GPx Periodontitis- Decreased

Vit-C Sepsis- Increased

*Oxidative stress (OS) in bacterial and viral diseases.*


*Modulations in Oxidative Stress of Erythrocytes during Bacterial and Viral Infections DOI: http://dx.doi.org/10.5772/intechopen.98236*

*HBV- Hepatitis B virus; HCV- Hepatitis C virus; HIV- Human immunodeficiency virus; JEV- Japanese encephalitis virus; DENV- Dengue virus; RSV- Respiratory syncytial virus; Covid-19-Coronavirus Disease 2019. TBARS- Thiobarbituric acid reactive substances; MDA- Malondialdehyde; 8-IP- 8-isoprostanes; 4-HNE-4-hydroxynonenal; PC- Protein Carbonyls; 8-OHdG- 8-Hydroxy-deoxyguanosine; NO- nitric oxide; NT-Nitrotyrosine; SOD- Superoxide dismutase; CAT- Catalase; GPx- Glutathione Peroxidase; GSH- Glutathione; GR- Glutathione Reductase, GST- Glutathione S-Transferase; UA- Uric acid; Vit-C- Vitamin C; Vit-E- Vitamin E; FRAP- ferric reducing antioxidant power; TRAP- Total-trapping radical antioxidant potential.*

#### **Table 1.**

*Oxidative stress (OS) in bacterial and viral diseases.*

*Erythrocyte - A Peripheral Biomarker for Infection and Inflammation*

*1.1.3 Pneumococcal meningitis-* streptococcus pneumoniae

*1.1.4 Gastritis/gastric cancer-* Helicobacter pylori

disintegration of bacterial cell membrane, DNA damage, deactivation of metabolic enzymes or proteins [53–56]. After invasion into the host, mycobacteria induce NADPH oxidase 2 (NOX2) expression to generate superoxide radicals

**.−**), which are then converted to more toxic hydrogen peroxide (H2O2) by superoxide dismutase (SOD) and subsequently reduced to water and molecular oxygen by catalase [57, 58]. NADPH oxidase 2 (NOX2) is the key enzyme responsible for the cellular ROS production by using superoxide radicals (O2-) as precursor molecule [59]. Alterations in regulatory components of NOX2 results in generation of phagocytic oxidative stress and phagocytic burst to eliminate

However, pathogenic mycobacteria can inhibit oxidative stress mechanisms by modulation of cell signaling mechanisms, up-regulation of antioxidant enzymes

Pneumococcal meningitis is a life-threatening disease characterized by acute

can damage neurons and glial cells by lipid peroxidation

infection affecting the pia mater, arachnoid, and subarachnoid spaces [63]. *Streptococcus pneumoniae* crosses the blood–brain barrier (BBB) and disrupts the intraepithelial tight junctions. Host polymorphonuclear leukocytes produce nitric oxide, superoxide radicals, and hydrogen peroxide in response to bacterial infection. O2 − ∙ and NO∙ can lead to the formation of peroxynitrite (ONOO), a strong

and cell membrane destabilization, resulting in DNA disintegration and subsequent poly (ADP-ribose) polymerase (PARP) activation. Elevated 4-HNE and MDA levels are found in bacterial meningitis patients [67]. Thus, ROS/ RNS can be considered key players of immune activation, blood–brain barrier disruption, vascular failure, neuronal injury, and cochlear damage during pneumococcal

*Helicobacter pylori* is the causative pathogen for human gastritis or gastric cancer, which is characterized with inflammation and ulceration of the stomach and duodenum. Gastric cancer arises from oxidative stress and environmental toxins, which increase DNA mutation rates [68]. The possible sources of ROS/ RNS in *H. pylori* infected stomach, include neutrophils, vascular endothelial cells and gastric mucosal cells. Neutrophils are believed to be the main source of ROS/ RNS [69] and their production is catalyzed by NADPH oxidase on the cell membrane [59]. These highly reactive ROS (HOCl and˙OH) are used by the phagocyte to kill pathogenic bacteria. *H. pylori* infected gastric mucosal phagocytes produce greater amounts of ROS, which is believed to be the major cause of gastric muco-

Oxidative Stress has always played a major pathogenic role in HIV and hepatitis infections. HIV causes decrements in glutathione (GSH), cystine, vitamin C and SOD levels, and increments in lipid peroxidation [70–73]. A decline in the antioxidant capacity represents a weakened immune system, thus requiring more antioxidants to maintain normal functionality [74]. Hepatitis, similar to HIV, also increases the lipid peroxidation (malondialdehyde (MDA) and 4-hydroxynonenal (HNE))

and activity of caspases, whereas reduces zinc [75–77] (**Table 1**).

**36**

(O2

enclosed pathogen [58].

oxidant [64–66]. ONOO<sup>−</sup>

meningitis.

sal damage.

*1.1.5 HIV and Hepatitis*

and redox buffering systems [59–62].
