**3. Physiological functions of mitochondrial ROS in diverse cellular processes**

mROS generation is a ubiquitous phenomenon during life of eukaryotic cells [68]. mROS-induced oxidative stress is considered a main contributor to the aetiology of both normal senescence and severe pathologies. Under normal physiological conditions, mROS emission is accounted for ∼2% of the total O2 consumption, of which the decomposition is well-controlled [2]. Accumulation of mROS, which is an imbalance of neutralization, induces deleterious consequences such as neurodegenerative disease [69], cardiovascular disease [70] and cancers [71]. However, depending on the cellular environment, antioxidant machinery-regulated oxidative stress could initiate diverse cellular responses, involved in cell protection, initiating coordinated activation of mitochondrial fission and autophagy to carry out clearance of abnormal mitochondria and cells, which are to protect spreading the damage to the adjacent cells [72, 73]. H2O2 is the primary molecule of mROS utilized for intracellular signalling, which selectively reacts with cysteine residues in redox-sensitive proteins, altering activities or conformations of the proteins to regulate signal transduction [74–76]. Mechanistically, H2O2 oxidizes thiol groups (SH) on cysteine residues to form sulphenic acid (SOH), which react with GSH to become glutathionylated (GSSG), with neighbouring thiols to form a disulphide bond (S-S) or with amides to form a sulphenyl amide (S-N) [77, 78]. In this section, we introduce the physiological roles and regulations of mROS in diverse cellular processes such as proliferation, differentiation, autophagy, immunity and aging (**Figure 3**).

**Figure 3.**

*Physiological regulation by mitochondrial ROS. mROS contribute to the various physiological cellular processes, including proliferation, differentiation autophagy, immunity and aging.*

**17**

(SCs) [100, 101].

**3.2 Differentiation**

*Physiological Functions of Mitochondrial Reactive Oxygen Species*

Accumulation of mitochondria-derived ROS enables to prompt cell proliferation

inhibition and cellular senescence [79, 80]. However, the cells essentially utilize mROS for survival and growth via multiple mechanisms in diverse circumstances. mROS regulate cell proliferation during hypoxia. Under the hypoxic condition (a low O2 environment, generally 0.3–3% of O2), the cells raise transcriptional and non-transcriptional responses to increase O2 supply, simultaneously reducing O2 consumption. These adaptations to hypoxia are enhanced by mROS. The hypoxiainducible factors (HIFs) such as HIF1, HIF2 and HIF3 orchestrate the transcriptional response to the hypoxia, promoting erythropoietin (EPO) expression to increase erythropoiesis, vascular endothelial growth factor (VEGF) to promote blood vessel formation and glycolysis enzymes to retain ATP levels [81, 82]. HIFs are heterodimers consisting of two basic helix-loop-helix/PAS proteins: a stable β-subunit and one of three unstable labile α-subunits (HIF1α, HIF2α and HIF-3α) [83, 84]. Under normoxic conditions, prolyl hydroxylase domain protein 2 (PHD2)

leads to hydroxylation of HIFα at two proline residues, which target via Von

Hippel-Lindau (VHL) E3 ubiquitin ligase-dependent proteasomal degradation [85]. However, under the hypoxic condition, HIFα is stabilized, which is then dimerized with HIF-1β and binds HIF-response elements (HRE) to recruit gene transcription [86]. Moreover, mitochondrial DNA lacking ρ° cells are unable to stabilize HIFα proteins under hypoxic condition, which results from failure of mROS production by ETC deficiency. In contrast, MMP reconstitution restores mROS, which leads to HIFα and cell proliferation [87]. Chemical inhibition of mitochondrial ETC also attenuates mROS production in mitochondria-repleted cells, interrupting to stabilize HIFα under hypoxia [88]. Genetic loss of the complex III subunit Rieske iron-sulphur protein (RISP) or Cytc also inhibits mROS production and HIFα stabilization [89–91]. It is also indicated that mROS are requisite to activate HIFs by

mROS are also involved in vascular smooth muscle cell (VSMC) proliferation. Angiotensin II (AngII) is a peptide hormone basically involved in sodium and water homeostasis and vascular contraction, which is also recognized to influence cell growth and proliferation [93]. AngII exerts physiological effects by signalling via interacting with angiotensin type 1 receptors (AT1Rs) [94]. In VSMCs, AngII signalling is required to activate a multitude of mitogenic signalling cascades via crosstalk with growth factor receptors such as epidermal growth factor receptor (EGFR), platelet-derived growth factor receptors (PDGFR) and insulin receptor (IR). Intracellular signalling of VSMC proliferation is stimulated by AngII signallingtriggered mROS production and subsequently induced via mitogenic serine/threo-

Despite the detrimental effects, mROS function as signal transduction molecules in regulation of stem cells [96]. Depletion of ataxia telangiectasia mutated (ATM) kinase or forkhead box O (FOXO) transcriptional factors increases mROS levels, which impairs hematopoietic stem cell (HSC) proliferation [97–99]. Although the increased mROS level impairs the differentiation of HSCs, a decreased mROS level also has negative effects for self-renewal in neural and spermatogonia stem cells

mROS function as active signalling molecules for diverse cell differentiation. Stem cells (SCs, embryonic or adult) have potentials to self-renew for maintaining stem cell pool or differentiate to the multicellular organism and supply de novo

*DOI: http://dx.doi.org/10.5772/intechopen.88386*

**3.1 Proliferation**

non-hypoxic stimulus [92].

nine kinases, including ERK1/2 and p38MAPK [95].

## **3.1 Proliferation**

*Free Radical Medicine and Biology*

**processes**

aging (**Figure 3**).

and nicotinamide nucleotide transhydrogenase (NNT) [61]. Catalase catalyses the decomposition of hydrogen peroxide to water and oxygen, existing as a tetramer composed of four identical monomers, each of which contains a heme group at the active site. Catalase also requires NADPH as a reducing equivalent to

**3. Physiological functions of mitochondrial ROS in diverse cellular** 

mROS generation is a ubiquitous phenomenon during life of eukaryotic cells [68]. mROS-induced oxidative stress is considered a main contributor to the aetiology of both normal senescence and severe pathologies. Under normal physiological conditions, mROS emission is accounted for ∼2% of the total O2 consumption, of which the decomposition is well-controlled [2]. Accumulation of mROS, which is an imbalance of neutralization, induces deleterious consequences such as neurodegenerative disease [69], cardiovascular disease [70] and cancers [71]. However, depending on the cellular environment, antioxidant machinery-regulated oxidative stress could initiate diverse cellular responses, involved in cell protection, initiating coordinated activation of mitochondrial fission and autophagy to carry out clearance of abnormal mitochondria and cells, which are to protect spreading the damage to the adjacent cells [72, 73]. H2O2 is the primary molecule of mROS utilized for intracellular signalling, which selectively reacts with cysteine residues in redox-sensitive proteins, altering activities or conformations of the proteins to regulate signal transduction [74–76]. Mechanistically, H2O2 oxidizes thiol groups (SH) on cysteine residues to form sulphenic acid (SOH), which react with GSH to become glutathionylated (GSSG), with neighbouring thiols to form a disulphide bond (S-S) or with amides to form a sulphenyl amide (S-N) [77, 78]. In this section, we introduce the physiological roles and regulations of mROS in diverse cellular processes such as proliferation, differentiation, autophagy, immunity and

prevent oxidative inactivation of the enzyme [67] (**Figure 2**).

**16**

**Figure 3.**

*Physiological regulation by mitochondrial ROS. mROS contribute to the various physiological cellular processes,* 

*including proliferation, differentiation autophagy, immunity and aging.*

Accumulation of mitochondria-derived ROS enables to prompt cell proliferation inhibition and cellular senescence [79, 80]. However, the cells essentially utilize mROS for survival and growth via multiple mechanisms in diverse circumstances.

mROS regulate cell proliferation during hypoxia. Under the hypoxic condition (a low O2 environment, generally 0.3–3% of O2), the cells raise transcriptional and non-transcriptional responses to increase O2 supply, simultaneously reducing O2 consumption. These adaptations to hypoxia are enhanced by mROS. The hypoxiainducible factors (HIFs) such as HIF1, HIF2 and HIF3 orchestrate the transcriptional response to the hypoxia, promoting erythropoietin (EPO) expression to increase erythropoiesis, vascular endothelial growth factor (VEGF) to promote blood vessel formation and glycolysis enzymes to retain ATP levels [81, 82]. HIFs are heterodimers consisting of two basic helix-loop-helix/PAS proteins: a stable β-subunit and one of three unstable labile α-subunits (HIF1α, HIF2α and HIF-3α) [83, 84]. Under normoxic conditions, prolyl hydroxylase domain protein 2 (PHD2) leads to hydroxylation of HIFα at two proline residues, which target via Von Hippel-Lindau (VHL) E3 ubiquitin ligase-dependent proteasomal degradation [85]. However, under the hypoxic condition, HIFα is stabilized, which is then dimerized with HIF-1β and binds HIF-response elements (HRE) to recruit gene transcription [86]. Moreover, mitochondrial DNA lacking ρ° cells are unable to stabilize HIFα proteins under hypoxic condition, which results from failure of mROS production by ETC deficiency. In contrast, MMP reconstitution restores mROS, which leads to HIFα and cell proliferation [87]. Chemical inhibition of mitochondrial ETC also attenuates mROS production in mitochondria-repleted cells, interrupting to stabilize HIFα under hypoxia [88]. Genetic loss of the complex III subunit Rieske iron-sulphur protein (RISP) or Cytc also inhibits mROS production and HIFα stabilization [89–91]. It is also indicated that mROS are requisite to activate HIFs by non-hypoxic stimulus [92].

mROS are also involved in vascular smooth muscle cell (VSMC) proliferation. Angiotensin II (AngII) is a peptide hormone basically involved in sodium and water homeostasis and vascular contraction, which is also recognized to influence cell growth and proliferation [93]. AngII exerts physiological effects by signalling via interacting with angiotensin type 1 receptors (AT1Rs) [94]. In VSMCs, AngII signalling is required to activate a multitude of mitogenic signalling cascades via crosstalk with growth factor receptors such as epidermal growth factor receptor (EGFR), platelet-derived growth factor receptors (PDGFR) and insulin receptor (IR). Intracellular signalling of VSMC proliferation is stimulated by AngII signallingtriggered mROS production and subsequently induced via mitogenic serine/threonine kinases, including ERK1/2 and p38MAPK [95].

Despite the detrimental effects, mROS function as signal transduction molecules in regulation of stem cells [96]. Depletion of ataxia telangiectasia mutated (ATM) kinase or forkhead box O (FOXO) transcriptional factors increases mROS levels, which impairs hematopoietic stem cell (HSC) proliferation [97–99]. Although the increased mROS level impairs the differentiation of HSCs, a decreased mROS level also has negative effects for self-renewal in neural and spermatogonia stem cells (SCs) [100, 101].

#### **3.2 Differentiation**

mROS function as active signalling molecules for diverse cell differentiation. Stem cells (SCs, embryonic or adult) have potentials to self-renew for maintaining stem cell pool or differentiate to the multicellular organism and supply de novo

functional cells to tissues throughout the life of the organism. During differentiation of SCs, the mitochondrial oxidative metabolism is highly stimulated, and thus cellular respiration and mROS production increase [102–105].

In SCs, generally, mitochondria exhibit immature mitochondrial networks and primitive cristae [106]. As bone marrow-derived human mesenchymal stem cells (MSCs) differentiate to osteoblasts, mitochondrial biosynthesis increases by PGC-1α activation [102]. Mitochondrial mass and oxygen consumption increase during differentiation of human embryonic stem cells (ESCs) [107] or pluripotent stem cells (PSCs) [108]. Knockdown of the complex III protein RISP or mitochondrial-targeted antioxidants inhibited differentiation of human MSCs to adipocytes, indicating that mROS are required for differentiation of MSCs [109]. Furthermore, during differentiation of human PSCs, uncoupling protein 2 (UCP2) expression is repressed, which is required for metabolic transition from glycolysis to mitochondrial glucose oxidation. Knockdown of UCP2 expression facilitates mROS accumulation, which stimulate the PSC differentiation to cardiomyocytes. Ectopic UCP2 expression impairs the differentiation with retardation of mROS accumulation and embryonic body formation [110].

mROS, at least within physiological concentrations, have critical roles in processes of myogenic differentiation and muscle regeneration [111]. mROS could promote mitochondrial biogenesis, which is an essential molecule in myogenic differentiation, via peroxisome proliferator-activated receptor gamma coactivator 1 (PGC1)-activated signalling pathway [112]. Myogenic cells are armed with antioxidant enzymes such as SODs, catalase, Gpxs, Prxs, γ-glutamylcysteine synthetase (γGCS) and heme oxygenase-1 (HO-1) [113–120]. These antioxidant enzymes could play as critical signalling molecules to maintain muscle homeostasis in company with primarily neutralizing excessive ROS [121]. mROS facilitate myoblast differentiation and hypertrophy via insulin growth factor 1 (IGF1) signalling pathway [111], which enhances phosphorylation of IGF1 receptor (IGF1R) [122]. Mitochondrial complex I-derived H2O2 acts as a signalling molecule to induce cardiac myogenic differentiation. Chemical inhibition of the complex I and treatment of mitochondrial-specific antioxidant exhibits reduction in mROS production and thus impairs the myoblast differentiation [123]. Moreover, mROS induced phosphatase and tensin homolog (PTEN) oxidative inactivation and thereby stimulated phosphoinositide 3-kinase (PI3K)-AKT signalling pathway to express myogenic genes during skeletal myoblast differentiation and muscle regeneration [124]. In differentiation of VSMCs, mROS production also elevates to activate p38 MAPK signalling pathway [125]. However, the complexity of mROS involvement still requires further investigation to elucidate the certain roles of oxidative stress in myogenic differentiation and muscle regeneration.

#### **3.3 Autophagy**

Autophagy is a conserved catabolic process that controls cellular degradation of unnecessary or dysfunctional cellular components in the lysosome [126]. Generally, the autophagy continuously occurs to recycle damaged proteins and organelles for cellular homeostasis under normal conditions [127]. The autophagy has at least three different types: (1) Macroautophagy (usually referred to as autophagy): cytosolic contents are delivered to the lysosome by autophagosomes. (2) Microautophagy: the contents are directly introduced into lysosomal membrane. (3) Chaperone-mediated autophagy: the target proteins contain a motif KFERQ, and then the chaperone (KFERQ )-protein complex binds lysosomeassociated membrane protein 2A (LAMP2A) receptors on the lysosomal membrane [128]. Autophagy induction results in recruitment of autophagy-related proteins

**19**

**3.4 Immunity**

*Physiological Functions of Mitochondrial Reactive Oxygen Species*

ling pathway in cardiac or skeletal muscle [137, 138].

which is a receptor for hypoxia-induced mitophagy [148].

adaptive immune responses [149, 150].

In immune system, it is well known that ROS contribute to directly eliminate pathogens via the oxidative burst mediated by NADPH oxidases (NOXs) that are plasma membrane-bound enzyme complexes in phagosomes. However, intracellular redox state intervened by mROS has emerged to be essential for innate and

mROS are crucial for Toll-like receptor (TLR) signalling pathways [19]. Activation of cell surface TLRs such as TLR1, TLR2 and TLR4 increases in mROS production via TNF receptor-associated factor 6 (TRAF) and evolutionary

(ATGs) to a punctate structure, phagophore assembly site (PAS), where proteins of the uncoordinated-51-like kinase 1 (ULK1) complex assemble to initiate autopha-

In autophagy signalling, mitochondria are considered as main source of ROS [130]. mROS, especially as H2O2, are required for autophagy induction in response to nutrient starvation and rapamycin, tumour necrosis factor α (TNFα) and nerve growth factor (NGF) deprivation [131–134]. H2O2 modulates the cysteine protease Atg4, which cleaves c-terminus of Atg8 (or light chain 3, LC3), and thus enables the addition of phosphatidylethanolamine (PE) to Atg8. Subsequently the active Atg8 is conjugated on the autophagosomal membrane, leading to the autophagosome formation [131]. H2O2 also disrupts the MMP to inhibit Akt/mammalian target of rapamycin (mTOR) signalling pathway for autophagy initiation [135, 136]. Furthermore, elevated H2O2 induces autophagy via activation of p38 MAPK signal-

In physiological energy metabolism, mitochondrial ATP production by OXPHOS induces mROS generation, resulting in a certain degree of constitutive mitochondrial damage and submitochondrial particles. The damaged mitochondria cause ATP depletion and Cytc release, which eventually leads to activation of caspases and then onset of apoptosis [139, 140]. To prevent cell death, the dysfunctional mitochondria are thus sequestered from the mitochondrial network and eliminated by selective autophagy, mitophagy, to properly maintain mitochondrial quantity and quality [130]. Therefore, mitophagy limits further mROS generation, which promotes turnover of mitochondria and avoids accumulation of dysfunctional mitochondria. Mitophagy is mainly controlled by the PTEN-induced kinase 1 (PINK1)-Parkin pathway, which is stimulated upon the MMP depolarization. PINK1 is a Ser/Thr kinase that translocates on the outer mitochondrial membrane, which is stabilized by low MMP, thereby sensing mitochondrial depolarization [141–143]. Then, PINK1 recruits Parkin that ubiquitylates OMM-located proteins such as VDAC1, resulting in recruitment of autophagic machinery and the selective sequestration of ubiquitylated mitochondria within autophagosomes [130]. Furthermore, the mitochondrial proteins, BCL2/adenovirus E1B 19-kDainteracting protein 3 (Bnip3) and Bcl-2/adenovirus E1B 19-kDa-interacting protein 3 (Bnip3L/NIX), participate in mitophagy [144]. In response to oxidative stress after ischemia/reperfusion (I/R), Bnip3 is homodimerised, to be activated, resulting in induction of mitophagy [145]. NIX, an atypical BH3 protein, is required for mitophagy in erythrocyte development. It directly recognizes autophagosome-sited GABA receptor-associated protein (GABARAP) that is a functional homolog of LC3 and subsequently induces mitophagy [126, 146]. Bnip3 and NIX directly bind to the autophagy machinery components, differently to PINK1 or Parkin [147]. ULK1 also regulates mitophagy via translocation to mitochondria to phosphorylate FUN14 domain containing 1 (FUNDC1) protein, a mitochondrial outer membrane protein,

*DOI: http://dx.doi.org/10.5772/intechopen.88386*

gosome formation [129].

*Free Radical Medicine and Biology*

functional cells to tissues throughout the life of the organism. During differentiation of SCs, the mitochondrial oxidative metabolism is highly stimulated, and thus

In SCs, generally, mitochondria exhibit immature mitochondrial networks and primitive cristae [106]. As bone marrow-derived human mesenchymal stem cells (MSCs) differentiate to osteoblasts, mitochondrial biosynthesis increases by PGC-1α activation [102]. Mitochondrial mass and oxygen consumption increase during differentiation of human embryonic stem cells (ESCs) [107] or pluripotent stem cells (PSCs) [108]. Knockdown of the complex III protein RISP or mitochondrial-targeted antioxidants inhibited differentiation of human MSCs to adipocytes, indicating that mROS are required for differentiation of MSCs [109]. Furthermore, during differentiation of human PSCs, uncoupling protein 2 (UCP2) expression is repressed, which is required for metabolic transition from glycolysis to mitochondrial glucose oxidation. Knockdown of UCP2 expression facilitates mROS accumulation, which stimulate the PSC differentiation to cardiomyocytes. Ectopic UCP2 expression impairs the differentiation with retardation of mROS

mROS, at least within physiological concentrations, have critical roles in processes of myogenic differentiation and muscle regeneration [111]. mROS could promote mitochondrial biogenesis, which is an essential molecule in myogenic differentiation, via peroxisome proliferator-activated receptor gamma coactivator 1 (PGC1)-activated signalling pathway [112]. Myogenic cells are armed with antioxidant enzymes such as SODs, catalase, Gpxs, Prxs, γ-glutamylcysteine synthetase (γGCS) and heme oxygenase-1 (HO-1) [113–120]. These antioxidant enzymes could play as critical signalling molecules to maintain muscle homeostasis in company with primarily neutralizing excessive ROS [121]. mROS facilitate myoblast differentiation and hypertrophy via insulin growth factor 1 (IGF1) signalling pathway [111], which enhances phosphorylation of IGF1 receptor (IGF1R) [122]. Mitochondrial complex I-derived H2O2 acts as a signalling molecule to induce cardiac myogenic differentiation. Chemical inhibition of the complex I and treatment of mitochondrial-specific antioxidant exhibits reduction in mROS production and thus impairs the myoblast differentiation [123]. Moreover, mROS induced phosphatase and tensin homolog (PTEN) oxidative inactivation and thereby stimulated phosphoinositide 3-kinase (PI3K)-AKT signalling pathway to express myogenic genes during skeletal myoblast differentiation and muscle regeneration [124]. In differentiation of VSMCs, mROS production also elevates to activate p38 MAPK signalling pathway [125]. However, the complexity of mROS involvement still requires further investigation to elucidate the certain roles of oxidative stress in

Autophagy is a conserved catabolic process that controls cellular degradation of unnecessary or dysfunctional cellular components in the lysosome [126]. Generally, the autophagy continuously occurs to recycle damaged proteins and organelles for cellular homeostasis under normal conditions [127]. The autophagy has at least three different types: (1) Macroautophagy (usually referred to as autophagy): cytosolic contents are delivered to the lysosome by autophagosomes. (2) Microautophagy: the contents are directly introduced into lysosomal membrane. (3) Chaperone-mediated autophagy: the target proteins contain a motif KFERQ, and then the chaperone (KFERQ )-protein complex binds lysosomeassociated membrane protein 2A (LAMP2A) receptors on the lysosomal membrane [128]. Autophagy induction results in recruitment of autophagy-related proteins

cellular respiration and mROS production increase [102–105].

accumulation and embryonic body formation [110].

myogenic differentiation and muscle regeneration.

**18**

**3.3 Autophagy**

(ATGs) to a punctate structure, phagophore assembly site (PAS), where proteins of the uncoordinated-51-like kinase 1 (ULK1) complex assemble to initiate autophagosome formation [129].

In autophagy signalling, mitochondria are considered as main source of ROS [130]. mROS, especially as H2O2, are required for autophagy induction in response to nutrient starvation and rapamycin, tumour necrosis factor α (TNFα) and nerve growth factor (NGF) deprivation [131–134]. H2O2 modulates the cysteine protease Atg4, which cleaves c-terminus of Atg8 (or light chain 3, LC3), and thus enables the addition of phosphatidylethanolamine (PE) to Atg8. Subsequently the active Atg8 is conjugated on the autophagosomal membrane, leading to the autophagosome formation [131]. H2O2 also disrupts the MMP to inhibit Akt/mammalian target of rapamycin (mTOR) signalling pathway for autophagy initiation [135, 136]. Furthermore, elevated H2O2 induces autophagy via activation of p38 MAPK signalling pathway in cardiac or skeletal muscle [137, 138].

In physiological energy metabolism, mitochondrial ATP production by OXPHOS induces mROS generation, resulting in a certain degree of constitutive mitochondrial damage and submitochondrial particles. The damaged mitochondria cause ATP depletion and Cytc release, which eventually leads to activation of caspases and then onset of apoptosis [139, 140]. To prevent cell death, the dysfunctional mitochondria are thus sequestered from the mitochondrial network and eliminated by selective autophagy, mitophagy, to properly maintain mitochondrial quantity and quality [130]. Therefore, mitophagy limits further mROS generation, which promotes turnover of mitochondria and avoids accumulation of dysfunctional mitochondria. Mitophagy is mainly controlled by the PTEN-induced kinase 1 (PINK1)-Parkin pathway, which is stimulated upon the MMP depolarization. PINK1 is a Ser/Thr kinase that translocates on the outer mitochondrial membrane, which is stabilized by low MMP, thereby sensing mitochondrial depolarization [141–143]. Then, PINK1 recruits Parkin that ubiquitylates OMM-located proteins such as VDAC1, resulting in recruitment of autophagic machinery and the selective sequestration of ubiquitylated mitochondria within autophagosomes [130]. Furthermore, the mitochondrial proteins, BCL2/adenovirus E1B 19-kDainteracting protein 3 (Bnip3) and Bcl-2/adenovirus E1B 19-kDa-interacting protein 3 (Bnip3L/NIX), participate in mitophagy [144]. In response to oxidative stress after ischemia/reperfusion (I/R), Bnip3 is homodimerised, to be activated, resulting in induction of mitophagy [145]. NIX, an atypical BH3 protein, is required for mitophagy in erythrocyte development. It directly recognizes autophagosome-sited GABA receptor-associated protein (GABARAP) that is a functional homolog of LC3 and subsequently induces mitophagy [126, 146]. Bnip3 and NIX directly bind to the autophagy machinery components, differently to PINK1 or Parkin [147]. ULK1 also regulates mitophagy via translocation to mitochondria to phosphorylate FUN14 domain containing 1 (FUNDC1) protein, a mitochondrial outer membrane protein, which is a receptor for hypoxia-induced mitophagy [148].

#### **3.4 Immunity**

In immune system, it is well known that ROS contribute to directly eliminate pathogens via the oxidative burst mediated by NADPH oxidases (NOXs) that are plasma membrane-bound enzyme complexes in phagosomes. However, intracellular redox state intervened by mROS has emerged to be essential for innate and adaptive immune responses [149, 150].

mROS are crucial for Toll-like receptor (TLR) signalling pathways [19]. Activation of cell surface TLRs such as TLR1, TLR2 and TLR4 increases in mROS production via TNF receptor-associated factor 6 (TRAF) and evolutionary

conserved signalling intermediate in Toll pathways (ECSIT) signalling pathway [151]. The TRAF6 or ECSIT depletion promotes reduction of mROS generation in macrophages and thus impairment of bacterial clearance [151]. Lipopolysaccharide (LPS)-induced pro-inflammatory cytokines such as TNFα and IL-6 are controlled by mROS generation [152]. Innate immune response enhancement in patients with TNF receptor-associated periodic syndrome (TRAPS) that is an autoinflammatory disorder is affected by missense mutations in the type 1 TNF receptor (TNFR1), which might be attributable to mitochondrial ROS generation [152].

mROS control pattern recognition receptors (PRRs) such as nuclear oligomerization domain (NOD)-like receptors (NLRs). NLRs form multisubunit protein complexes termed inflammasomes that activate caspase-1 resulting in proteolytic cleavage and pro-inflammatory cytokine IL-1β maturation [153, 154]. Pathogenassociated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs) such as lipopolysaccharide (LPS), asbestos, ATP and uric acid activate NLR family pyrin domain containing 3 (NLRP3) inflammasome via mROS generation [155, 156]. Pharmacologic or genetic inhibition of autophagy elevates mROS concentration, which heightens inflammasome activation [157, 158]. Increase of mROS persuades lysosomal membrane permeabilization, which is required for NLRP3 activation [159]. Activation of NLRP3 inflammasome results in mitochondrial damage, interrupting mitophagic signalling [160]. Notably, calcium influx contributes to mitochondrial damage, which might increase mROS production and mtDNA release to amplify NLRP3 inflammasome activation [161, 162]. However, it remains to be further delineated how PAMPs and DAMPs increase mROS to properly activate NLRP3 inflammasome.

In adaptive immune responses, T cells are functionally crucial in response to the pathogens [150, 156]. In infectious condition, naïve T cells promptly proliferate and differentiate into effector T cells [163]. The activation of T cells requires increase in glycolysis and mitochondrial metabolism for synthesis of macromolecules in process of the proliferation and differentiation [156, 164, 165]. Elevated mROS concentration contributes to the T-cell activation; treatment of antioxidants inhibits cellular proliferation and interleukin-2 (IL-2) production [166]. Similarly, antioxidant administration to mice exhibits their reduced immunity after infection of the virus, suggesting that mROS are indispensable for the T-cell functions in vivo [167, 168]. The T-cell receptor (TCR) stimulation induces mROS production from complex I, which leads to activation of NF-κB and AP1 signalling, and in turn facilitates IL-2 and IL-4 productions that are imperative drivers in T-cell activation [169, 170].

#### **3.5 Aging**

Aging is a process that is concomitant with the accumulation of cellular damage over the time of all living organisms. In the 1950s, Denham Harman suggested the 'free radical theory of aging' as a molecular explanation for aging [171], in which free radicals, as byproducts of energy metabolism, develop cumulative cellular damage resulting in loss of organismal ability over time. The theory has been revised that the mitochondria-derived free radicals are causative of aging [172]. Mitochondrial dysfunction and consequent excessive ROS production result in inevitable cellular damage and subsequent cell death [173]. Oxidative damage to genomes, proteins and lipids has been associated with mitochondrial dysfunction and ultimately cellular senescence or death [174]. Consistently, overexpression of antioxidant enzymes reduces ROS production and subsequently protects DNA, which is interconnected to a prolonged life span in *Drosophila melanogaster* [175, 176].

Despite numerous evidences underpinning the detrimental roles of mROS in aging, the discoveries are questioning a direct correlation between oxidative stress

**21**

*Physiological Functions of Mitochondrial Reactive Oxygen Species*

responses that are beneficial to the organism and extend life span.

and the lifespan. A mitochondrial enzyme, doublecortin-like kinase 1 (MCLK1), reduction induces mitochondrial dysfunction that displays the regression of electron transport in mitochondrial respiratory chain and decline of TCA cycle activity [177]. In *Drosophila melanogaster*, mROS levels elevate along with age, but do not intervene with life span [178]. Furthermore, moderate ROS levels have been associated with an extension of longevity in *Drosophila melanogaster* and in young mice [179–181]. Therefore, physiologically controlled mROS might activate adaptive

Mitochondria are primary energy producers to generate ATPs via oxidative phosphorylation. For a long time, mROS have been considered as byproducts of biological energy metabolism during the ATP generation or by cellular redox system imbalance, which are highly aggressive and detrimental to the neighbouring cells and tissues. However, the roles of mROS have been extensively substantiated to understand normal physiology and pathology over the past decades. Mitochondriaderived H2O2 have been unequivocally recognized as essential molecules in a range

In this chapter, we have provided a brief discussion of current understanding of physiological roles of mROS by which mitochondria indeed contribute to the implementation of cellular proliferation, differentiation autophagy, innate and adaptive immunity and aging. In understanding the mechanisms regulating mitochondrial physiology and homeostasis, mROS production might provide a significant potential for the development of novel therapeutic strategy for the treatment of a wide

This work was supported by the National Research Foundation of Korea (NRF) grant funded by the Korean government (MEST) (NRF-2013R1A1A2061214, NRF-

2018R1D1A1B07048909 and NRF-2018R1A6A1A03025124).

The authors declare no conflict of interest.

*DOI: http://dx.doi.org/10.5772/intechopen.88386*

**4. Conclusion**

of physiological processes in cells.

range of human pathologies.

**Acknowledgements**

**Conflict of interest**

*Physiological Functions of Mitochondrial Reactive Oxygen Species DOI: http://dx.doi.org/10.5772/intechopen.88386*

and the lifespan. A mitochondrial enzyme, doublecortin-like kinase 1 (MCLK1), reduction induces mitochondrial dysfunction that displays the regression of electron transport in mitochondrial respiratory chain and decline of TCA cycle activity [177]. In *Drosophila melanogaster*, mROS levels elevate along with age, but do not intervene with life span [178]. Furthermore, moderate ROS levels have been associated with an extension of longevity in *Drosophila melanogaster* and in young mice [179–181]. Therefore, physiologically controlled mROS might activate adaptive responses that are beneficial to the organism and extend life span.
