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Journal of Sport and Health Science 2 (2013) 67e74

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Review

Mitochondrial redox metabolism in aging: Effect of exercise interventions

Hai Bo a,b, Ning Jiang a, Li Li Ji a,c, Yong Zhang a,*

aTianjin Key Laboratory of Exercise Physiology and Sports Medicine, Department of Health & Exercise Science, Tianjin University of Sport,

Tianjin 300381, ChinabDepartment of Military Training Medicines, Logistics University of Chinese People’s Armed Police Force, Tianjin 300162, China

c Laboratory of Physiological Hygiene and Exercise Science, School of Kinesiology, University of Minnesota, Minneapolis, MN 55455, USA

Received 12 December 2012; revised 20 January 2013; accepted 22 February 2013

Abstract

Mitochondrial redox metabolism has long been recognized as being central to the effects of aging and the development of age-relatedpathologies in the major oxidative organs. Consistent evidence has shown that exercise is able to retard the onset and impede the progression ofaging by modifying mitochondrial oxidanteantioxidant homeostasis. Here we provide a broad overview of the research evidence showing therelationship between mitochondrial redox metabolism, aging and exercise. We address part aspects of mitochondrial reactive oxygen species(ROS) metabolism, from superoxide production to ROS detoxification, especially antioxidant enzymes and uncoupling protein. Furthermore, wedescribe mitochondrial remodeling response to aging and exercise, which is accompanied by bioenergetics and redox regulation. In addition,potential mechanisms for redox signaling involved in mitochondrial remodeling and redox metabolism regulation are also reviewed.Copyright � 2013, Shanghai University of Sport. Production and hosting by Elsevier B.V. All rights reserved.

Keywords: Aging; Exercise; Mitochondrial remodeling; PGC-1a; Reactive oxygen species

1. Introduction

The aging process results in a gradual and progressivestructural and functional deterioration of biomolecules that isassociated with many pathological conditions, includingcancer, neurodegenerative diseases, sarcopenia, and liverdysfunction. In the 1950s, Harman1 first proposed the “freeradical theory of aging”. This theory contended that freeradicals, produced from normal metabolism, lead to

* Corresponding author.

E-mail address: yzhang@tjus.edu.cn (Y. Zhang)

Peer review under responsibility of Shanghai University of Sport

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irreversible cell damage and an overall functional declineduring aging. In the 1970s, he extended the theory to implicatemitochondrial production of reactive oxygen species (ROS,such as O2

�� and H2O2) as the main cause for age-relatedmacromolecular damage.2

Epidemiological studies clearly demonstrate that endur-ance exercise reduces the risk of chronic diseases and extendslife expectancy.3 Consistent evidence has shown that exerciseis able to retard the onset and impede the progression ofaging by modifying mitochondrial oxidanteantioxidanthomeostasis.4 However, there are still unanswered basicscientific questions in the implication of exercise in modu-lating aging. Especially, how increased ROS productionduring physical exercise could be involved in exercise-induced health-promoting benefits. In this review, weconsider the evidence that specific aspects of mitochondrialredox metabolism affect aging. We then discuss redox-sensitive signal transduction pathways associated with exer-cise in modulating aging.

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68 H. Bo et al.

2. Mitochondrial ROS production and aging

In the mitochondrial respiratory chain, under conditions inwhich complexes I and III are highly reduced, a significantproportion of electrons are diverted from them directly tomolecular oxygen, thus forming the radical superoxide (O2

��).In addition, it recently has been shown that p66shc, a proteinpresent in the intermembrane space, can also catalyze O2

��

production via electron transfer from reduced cytochrome c tomolecular oxygen.5 Although O2

�� is not membrane perme-able, it can be spontaneously or catalytically dismutated tohydrogen peroxide (H2O2), which can efficiently pass throughbiomembranes and therefore acts not only at its site offormation but also elsewhere.

At increased abundance, ROS are able to damage cellularcomponents such as proteins, nucleic acids, and lipids by theabstraction of electrons. The damaging potential of mito-chondrial ROS is the kernel of the “mitochondrial free radicaltheory of aging”, which suggests that mitochondrial ROS leadto an accumulation of molecular damage over time and, afterpassing certain thresholds, to cellular degeneration and death.In general, ROS production is found to be increased in cellsand is associated with aging. Furthermore, genetic studies inworm, fly, and mouse have linked enhanced stress resistanceor reduced free radical production with increased lifespan.

Because mitochondria are the major producer of ROS inmammalian cells, the close proximity to ROS places mito-chondrial prone to oxidative damage. During aging, theincrease in ROS production is associated with increased levelsof 8-oxoguanine (8-oxoG) from mitochondrial DNA (mtDNA)oxidation, increased nitration of complex II, and alteredcarbonylation of complex I, complex V, and isocitrate dehy-drogenase. Oxidative damage induced impaired function ofrespiratory chain will in turn trigger further accumulation ofROS, which results in a vicious cycle leading to energydepletion and ultimately cell death.

3. Mitochondrial antioxidant enzymes in aging

To avoid the potentially damaging effects of ROS, cells usea variety of systems to keep ROS within tolerable limits. Thisis especially important in mitochondria because the enzymesthat drive ATP production are rich in thiol residues that can beirreversibly oxidized. Superoxide generated by mitochondriais quickly dismutated to H2O2 by either manganese (Mn) orcopper/zinc (Cu/Zn)-dependent superoxide dismutase (SOD),which are located in the matrix and intermembrane space,respectively. Although H2O2 is the main ROS used insignaling, it is degraded by a variety of enzymes and lowmolecular weight molecules to prevent oxidative stress. Thechief H2O2-quenching systems in mitochondria include theGSH/glutathione peroxidase (GPx) system, peroxiredoxins,thioredoxins, and glutaredoxins. Catalase (CAT) also degradesH2O2 but only when H2O2 reaches high concentrations.

MnSOD is an intramitochondrial free radical scavengingenzyme that is the first line of defense against O2

�� produced.Over-expression of MnSOD in Drosophila melanogaster was

shown to have beneficial effects, extending lifespan.6

Furthermore, the tissue-specific over-expression of MnSODin motor neurons was sufficient to mediate the same effect.Conversely, complete ablation of this enzyme in mice causessevere oxidative damage to mitochondria and early postnataldeath associated with dilated cardiomyopathy and neuro-degeneration.7 MnSOD activity, both in rats and men, arereported to increase significantly with aging and to respondto age-related oxidative stress in the mitochondria by up-regulation. However, MnSOD protein and mRNA levels wasunchanged or decreased with age in most tissues. In addition,binding of transcription factor nuclear factor-kB (NF-kB) andtranscription factor activator protein-1 (AP-1) to correspond-ing oligonucleotide sequences present in the MnSOD genewere decreased in aged cells.8 Both NF-kB and AP-1 bindingsites are present in the promoter of the mammalian MnSODgene, and oxidative stress has been shown to activate theirbinding. A decrease in the binding of these nuclear factorsdespite increased ROS generation observed in aged cellssuggests that molecular signaling of MnSOD gene expressionis retarded due to aging. The observed increase in MnSODactivity in some tissues appears to be due to a posttranslationalmodification (activation) of the enzyme molecules duringaging.

CAT is widely distributed in the cell, with the majority ofactivity occurring in the mitochondria and peroxisomes, whereit functions to catalyze the decomposition of H2O2 to waterand oxygen. Transgenic mice that overexpress mitochondrialCAT have increased median and maximum lifespans, attenu-ated age-related changes including decline of diastolic func-tion, myocardial performance as well as ventricular fibrosis.9

CAT largely determines the functional antioxidant capacityof mitochondria and is the enzyme that is most affected inaging.10 In rats and mice cellular CAT levels drop with age,which is accompanied by an increase in oxidative stress.Chronically reducing CAT activity causes cells to displaya cascade of accelerated aging reactions.11 Sirtuin1 (Sirt1) hasbeen shown to affect CAT expression and be a determinant ofcell apoptosis by regulating cellular ROS levels. NAD-dependent deacetylase Sirt1 maintains cell survival by regu-lating CAT expression and by preventing the depletion of ROSrequired for cell survival. In contrast, excess ROS upregulatesSirt1, which activates CAT leading to rescuing apoptosis.12

4. Exercise interventions affecting mitochondrialantioxidant enzymes

There is an abundance of literature reporting muscle anti-oxidant adaptation to chronic exercise training and severalreviews have also addressed this topic thoroughly. Amongantioxidant enzymes in skeletal muscle, MnSOD activity hasconsistently been shown to increase with exercise training inan intensity-dependent manner. GPx activity has also shown toincrease after endurance training by most authors. GSH playsa critical role in muscle antioxidant defense during exercise.Several studies have demonstrated that GSH content and therate-limiting enzyme for GSH synthesis, a-glutamylcysteine

Mitochondrial redox metabolism in aging and exercise 69

synthetase (GCS), can be induced by endurance training in ratskeletal muscle. However, training effect on CAT activity hasbeen inconsistent and controversial.13

As mentioned previously several oxidative stress-sensitivesignaling pathways are operational in mammalian systems andplay an important role in maintaining cellular oxi-danteantioxidant balance. One of the most important involvesthe NF-kB. Several antioxidant enzymes contain NF-kBbinding sites in their gene promoter region, such as MnSOD,inducible nitric oxide synthetase, and GCS. Therefore, theycan be potential targets for exercise-activated upregulation viathe NF-kB signaling pathway. Ji et al.14 reported that an acutebout of treadmill running activated NF-kB binding in ratskeletal muscle 2 h after exercise. Furthermore, the NF-kBsignaling pathway was activated in a redox-sensitive mannerduring muscular contraction.14 Consistently, Gomez-Cabreraet al.15 have demonstrated that marathon running inducesactivation of the p50 subunit of the NF-kB complex inlymphocytes. This effect was prevented by treatment withallopurinol.

It is well documented that a single episode of muscularcontraction can activate the MAPK pathway in human skeletalmuscle. As members of MAPK family, extracellular signal-regulated kinase (ERK) and p38 MAPK activity can lead tothe sequential phosphorylation of a series of proteins, resultingin increased expression AP-1, which is an important DNA-binding site on many genes responsive to oxidative stress.16 Itwas also found that ROS generated during exercise activateMAPKs, which in turn activate NF-kB, resulting in anincreased expression of important enzymes associated withcell antioxidant defense (MnSOD and GPx). Prevention ofROS formation by inhibition of XO abolishes these effects.17

This result showed the role of cross-talk in the MAPK andNF-kB pathway in exercise-activated antioxidant signaling.

5. Uncoupling proteins, mitochondrial ROS production,and aging

It is well established that there is a strong positive corre-lation between mitochondrial membrane potential (Dj) andROS production. It has been reported that, at high Dj, evena small increase in membrane potential gives rise to a largestimulation of H2O2 production. Similarly, only a smalldecrease in Dj (10 mV), which is termed “mild uncoupling”,is able to inhibit H2O2 production by 70%. Hence, the milduncoupling of mitochondrial oxidative phosphorylation mayrepresent the first line of defense against oxidative stress.Brand18 first proposed the “uncoupling to survive” hypothesis,which states that increased uncoupling leads to greater oxygenconsumption and reduces the proton motive force by reducingDj which subsequently reduces ROS generation. Furtherevidence supports this hypothesis as a number of studies showhigher mitochondrial respiration and greater mitochondrialuncoupling increases lifespan in a diverse range of organisms.

Uncoupling proteins (UCPs), a heterogeneous family ofproteins located in the mitochondrial inner membrane, appearto be a logical candidate to achieve mild uncoupling. UCP2

and UCP3 are w73% homologous to each other and w58%homologous to the thermogenic protein, UCP1. UCP3 ishighly selectively expressed in skeletal muscle and brown fat,whereas UCP2 is ubiquitously expressed. It has been criticallyexamined that UCP2/UCP3 become true uncoupling proteinswhen stimulated by ROS or 4-hydroxynonenal, and thusbecome capable of diminishing oxidative damage by a feed-back mechanism activating mild uncoupling.19 Evidence forthis includes observations that mice deficient in UCP3 havehigher levels of membrane lipid peroxidative damage.20

Numerous studies have demonstrated that UCP2 over-expression diminishes ROS production in a large number oftissues examined, including the heart, pancreas, adipose tissue,muscle, immune system, spleen, and steatotic liver.21

Andrews and Horvath22 reported that UCP2 gene deletiondramatically reduces maximum lifespan in mice, and trans-genic UCP2 overexpression can partially rescue the earlyneonatal lethality of MnSOD deficiency. Similar observationshave been made with UCP3. Mitochondria from UCP2knockout mice produce more ROS and contain more oxidativedamage in comparison to wild-type.23 In addition, over-expression of UCP3 has been found to lower the increasedROS formation associated with aging.24 However, simulta-neous overexpression of UCP2 and UCP3 has minimal effecton longevity.19 This could be related to the requirement ofthese proteins for effector molecules. Maximal activity ofUCPs requires stimulation with fatty acids, including endproducts of fatty acid peroxidation. UCPs activities are alsoinhibited by adenine nucleotides. Thus, increasing UCPexpression without modifying levels of stimulators andinhibitors of their activities may not affect membrane potentialunder normal conditions.

It has been repeatedly demonstrated that aging is associatedwith an increase in oxidative stress, and therefore an increasein uncoupling may be expected with aging. However, therewas a dramatic reduction of UCP3 content associated withdecreased state 4 respiration of skeletal muscle mitochondriafrom old rats.25 In addition, it was found that muscle with themost active fiber type (type I) and higher resting O2 uptake hadmild uncoupling in adults but stable mitochondrial functioninto old age. This protection of mitochondrial functionsuggests that mild uncoupling is a mechanism that reduces theROS production, ameliorates mitochondrial damage, andassists fiber survival with age. In contrast, muscle with greatercontent of the least active fiber type (type II) and lower restingO2 uptake was well coupled in adults yet showed defect inmitochondrial coupling and depletion of ATP with ageconsistent with ROS damage. Thus, mild uncoupling mayexplain the paradox of greater longevity in the most activefibers by providing a protective mechanism that impacts thepace of cellular aging.26

6. Mitochondrial uncoupling in response to exercise

Several studies have shown that UCP3 expression wasincreased in response to an acute bout of exercise orcontractile activity in mammalian skeletal muscle.27 We

70 H. Bo et al.

investigated UCP2 and UCP3 expression along with mito-chondrial respiratory function and ROS generation in rat heartand skeletal muscle during and after an acute bout of pro-longed exercise up to 2.5 h. The data demonstrate that UCP2/UCP3 expression can be rapidly upregulated during prolongedexercise, and a coordinated regulation to reduce ROS gener-ation and to preserve mitochondrial respiratory function.28,29

Noticeably, MnSOD expression and enzyme activity was notup-regulated until after exercise under this experimentalcondition. Thus, our data suggest that during an acute bout ofdemanding exercise, the primary strategy of mitochondria toreduce oxidative stress is decreasing the production of O2

�� byoverexpressing UCP2/UCP3, instead of enhancing theremoval of O2

�� by overexpressing MnSOD.Our study also demonstrated that endurance training

attenuated acute stress-induced UCP2/UCP3 expression andactivity in rat. Meanwhile, endurance training significantlyelevated MnSOD activity and protein level.28,30 Similarly,Noland et al.31 reported that acute exercise increases skeletalmuscle UCP3 expression in untrained but not trained humans.These findings raised some interesting questions regarding therelationship among mitochondrial respiratory uncoupling,ROS generation, antioxidant defense, and ATP synthesis. Anameliorated exercise response of UCP2/UCP3 after trainingcould be viewed as a compensatory mechanism to avoidexcessive sacrifice of oxidative phosphorylation efficiency andheat production. McLeod et al.32 reported that in mitochondriafrom SOD2 knockout mice, ATP production was markedlyreduced due to a higher O2

�� concentration which activatedUCP2 excessively. Taken together, exercise training adaptationof MnSOD may enhance mitochondrial tolerance to ROSproduction and hence a smaller UCP2/UCP3 activation duringintensive exercise, thus protecting the efficiency of oxidativephosphorylation.

7. Mitochondrial remodeling is impaired in aging

Mitochondria are highly mobile and remarkably plasticorganelles that continuously undergo organellar turnover andmorphology remodeling. Evidence is accumulating thatmitochondrial dynamic remodeling are sensitive to variousphysiological and pathological stimuli, and mitochondrialremodeling are no epiphenomena, but central to cell functionand survival.

Mitochondria are able to change their architecture fromindividual structures to complex tubular networks throughfusion and fission. It has been established that this dynamicstructure is regulated by proteins controlling fission, such asmitochondrial fission 1 protein (Fis1) and dynamin-relatedprotein 1 (Drp1), and fusion, such as mitofusin homologsMfn1/2 and Dynamin-like 120 kDa protein OPA1. Geneticdefects in the proteins involved in mitochondrial fusion andfission lead to severely altered mitochondrial shape, loss ofmtDNA integrity, increased oxidative stress and apoptotic celldeath.33 In healthy cells, mitochondrial fusion providesa synchronized internal cable for translocating metabolites andintramitochondrial mixing during biogenesis, whereas

mitochondrial division facilitates the equal distribution ofmitochondria into daughter cells during mitosis and allows theselective degradation of damaged mitochondria throughautophagy. However, it has become increasingly clear thatthese protective mechanisms are markedly impaired in agingand that faulty mitochondrial dynamics might be involved inthe aging process. Crane et al.34 reported that the expression ofMfn2 and Drp1 genes is reduced in the skeletal muscle ofaging individuals. Furthermore, Using RNA interference toreduce Fis1 in mammalian cells, it was found that mitochon-dria become enlarged and flattened and these morphologicalchanges are correlated with increased a-galactosidase activity,a marker of cell senescence, and decreased mitochondrialmembrane potential, increased ROS generation, and DNAdamage.35

The constant renewal of mitochondria is crucial for main-taining healthy mitochondria with age. Accurate organellarturnover requires the coordination of two key cellularprocesses: mitochondrial biogenesis and selective degradation(autophagy). The capacity for mitochondrial biogenesisdiminishes with age and this is an important parameter in themitochondrial dysfunction associated with aging.36 Ifbiogenesis is affected, it is reasonable to expect that mito-chondrial turnover must be slower and the accumulation ofmodified lipids, proteins and DNA must also increase, furtheraggravating the situation resulting from the deficient activityof aged mitochondria. The precise reason for the decrease inthe rate of mitochondrial biogenesis during aging is currentlyunknown. However, it seems that both extra- and intracellularregulatory factors of mitochondrial biogenesis are implicated.In fact, the pleiotropic signaling of mitochondria, H2O2 andNO diffusion to the cytosol is modified in aged animals andcontributes to the decreased mitochondrial biogenesis in oldanimals.37

Autophagy appears to decline with age, and several keyplayers in the autophagic pathway (e.g., ATG5 and ATG7)show decreased expression in the brains of aging individuals.Stimulation of autophagy can increase the healthy lifespan inmultiple model organisms, including mice and primates.38

Autophagy can mitigate inflammatory reactions throughseveral mechanisms. Recently, it was found that autophagycan inhibit NLRP3 activation by removing permeabilized orROS producing mitochondria.39 Because pathological aging isaccompanied by chronic inflammation, these anti-inflamma-tory effects of autophagy may mediate additional healthbenefits. Mitochondrial dynamics are also important for properorganellar turnover in that they affect mitochondrial degra-dation pathways. Fis1 expression was found to be reduced inabnormal mitochondria, whereas overexpression of thisprotein blocked the senescence-related phenotype and main-tained cells in a proliferating state. The link between fissionproteins and mitochondrial turnover has also been illustratedin neuronal cells, in which Fis1 was shown to activate auto-phagy and the selective degradation of depolarizedmitochondria.40

In summary, aging compromises the remodeling of mito-chondria by disrupting the homeostatic regulation of

Mitochondrial redox metabolism in aging and exercise 71

mitochondrial fusion and fission pathways. The presence offragmented mitochondria owing to a decline in fusion and/oran increase in fission events can compromise mtDNA integrity,mitochondrial structural and functional complementation, andmitochondrial biogenesis, all of which can lead to mitochon-drial dysfunction. Conversely, the formation of enlargedmitochondria as a result of decreased fission and/or increasedfusion events can diminish mitochondrial turnover byimpairing autophagy and biogenesis, leading to the accumu-lation of damaged mitochondria in aged cells. In both cases,the abnormal mitochondria are unable to fulfill their life-sustaining roles. Therefore, age associated alterations inmitochondrial fusion and fission dynamics might play a caus-ative role in mitochondrial remodeling impairment andincrease susceptibility to cell death in response to varioustypes of stress during progressive aging.

8. PGC-1a, mitochondrial remodeling, and aging

Despite the complexity of the various signaling pathwaysthat regulate mitochondrial remodeling, they all seem to shareproliferator-activated receptor-a coactivator 1a (PGC-1a)family of transcription factors. PGC-1a appears to act asa master regulator of energy metabolism and mitochondrialremodeling by coordinating the activity of multiple tran-scription factors. PGC-1a strongly co-activates NRF-1, whichis an intermediate transcription factor to stimulate thesynthesis of mitochondrial transcription factor A (Tfam).Tfam can be considered the most important mammaliantranscription factor for mtDNA because it stimulates mito-chondrial DNA transcription and replication. Recently, Aqui-lano et al.41 showed that PGC-1a and Sirt1 not only colocalizewith the nuclear, but also interact with Tfam in the mito-chondria to form multiprotein complexes. In addition, PGC-1ahas been shown to be involved in mitochondrial networkremodeling through controlled fusion and fission. H2O2-stim-ulated Mfn1/2 expression was dramatically attenuated inPGC-1a knockout C2C12 muscle cells,42 whereas PGC-1aover-expression markedly enhances Mfn2 mRNA and proteinlevels in cultured muscle cells.43 PGC-1a also stimulates thetranscriptional activity of the human Mitofusin 2 (Mfn2) genepromoter, which requires the integrity of the nuclear hormoneestrogen-related receptor ERRa-binding element.44 Finally,PGC-1a controls most of the enzymes and proteins affectingoxidanteantioxidant balance such as UCP2/UCP3, MnSOD,and GPx expression.45

Kang et al.46 reported that aged (24 months) rats hadsignificantly lower gene expression in the PGC-1a signalingpathways, shown by decreased mRNA and protein contents forPGC-1a, Tfam and cytochrome c compared to young(4 months) rats. Furthermore, phosphorylation of the upstreamenzymes AMP kinase and p38 MAPK, as well as cAMPresponse element nuclear binding, was down-regulated. Der-bre et al.47 studied mitochondrial biogenesis and PGC-1aexpression in old rats and compared them with PGC-1aknockout mice. They found a striking similarity between theresponse to exercise training in PGC-1a knockout mice and in

old rats. It was shown that an age-associated lack of expres-sion of PGC-1a in response to not only training but also tocold exposure or triiodothyronine. Recently, Farhoud et al.48

reported that chronic oxidative stress suppresses the geneexpression of PGC-1a through activation of the ubiquitin-proteasome system, thereby inhibiting its downstream mito-chondrial biogenesis and metabolic gene expression programs.In addition, chronic inhibition of NF-kB via the pharmaco-logical agent pyrolidine dithiocarbamate (PDTC) was found toincrease PGC-1a level suggesting inactivity might down-regulate PGC-1a due to elevated NF-kB activity.49 It could bepresumed that aging related inflammation and subsequentactivation of NF-kB could negatively influence the expressionof PGC-1a and its effect on mitochondrial biogenesis. Theseresults showed that loss of mitochondrial biogenesis duringaging may be due to a lack of induction of PGC-1a.

9. Mitochondrial remodeling, ROS, and exercise

The term Hormesis has been adopted to explain how a mildoxidative stress associated with exercise can result in favorableadaptations that protect the body against more severe stressesand disorders derived from physical stress or other etiologicalorigin. The typical reaction to ROS can be described by a bell-shaped curve: low concentrations have a stimulating effect(signaling, receptor stimulation, and enzymatic stimulation),while a massive level of ROS inhibits enzyme activity andcauses apoptosis or necrosis. Recent evidence suggests thatsuppression of ROS production fails to extend lifespan inworms and may even decrease lifespan in humans, presumablydue to the reduction of the ROS signaling, which seems to beimportant for different cellular processes.50,51 It was suggestedthat exercise could increase the formation of ROS to a levelthat may induce significant, but tolerable, damage that can inturn induce beneficial adaptations. Short-term ROS productionis apparently important in prevention of aging by induction ofa process named mitohormesis and redox signaling.52 Thisprocess seems to be particularly important for the health effectof exercise.

Recently, we studied the effect of aging and exercise onmitochondrial remodeling in skeletal muscle because it containsmainly post-mitotic cells and because of the importance of thisorgan in aging (Fig. 1). Mfn1 and Drp1 protein were unchangedat 15 months. Mfn1 was decreased 39% at 20 months vs.5 months ( p < 0.05), whereas Drp1 showedw70% ( p < 0.01)increase at 20 months vs. 5 months. Compared with controlgroup, 12-week treadmill training resulted in an increase inMfn1 in 15 months ( p < 0.01), whereas in a small decrease in20 months ( p< 0.05) (Fig. 1A). In contrast to Mfn1, comparedwith control group, training resulted significantly increased inDrp1 protein both at 15 and 20 months vs. 5 months ( p < 0.01and p< 0.05, respectively) (Fig. 1B). Furthermore, cytochromec oxidase subunit IV (COXIV), the marker of mitochondrialbiogenesis, showed a 30% increase at 20 months vs. 5 months( p < 0.05). Compared with control group, training increasedCOXIV content by w30% at 5 and 15 months old rats( p < 0.05), and by w20% at 20 months old rats ( p < 0.05)

Fig. 1. Western blot analysis of Mfn1 (A), Drp1 (B), COXIV (C), and mitochondrial ATP synthase activity (D) in rat skeletal muscle of 12-week treadmill trained

5-, 15-, and 20-month-old rats and controls. yp < 0.05, yyp < 0.01: trained group compared with control group at various ages; *p < 0.05, **p < 0.01: in control

group, compared with 5 months; #p < 0.05, ##p < 0.01: in trained group, compared with 5 months.

72 H. Bo et al.

(Fig. 1C).We alsomeasured ATP synthase activity as indicationof mitochondrial energy production. ATP synthase activity wasunchanged at 15 and 20 months vs. 5 months, whereas trainingincreased its activity at various ages, 5-, 15-, and 20-month-old,compared with control group ( p< 0.01, p< 0.05, and p< 0.05,respectively) (Fig. 1D). Taken together, our data suggest thatregular exercise training stimulates mitochondrial biogenesis,a rejuvenation of the mitochondrial network via fission andfusion, and an improved efficient of mitochondrial energytransfer.All of these properties, working in conjunctionwith oneanother, would improve the overall functionality of mitochon-dria in aged cells (Zhang et al., unpublished data).

Gomez-Cabrera et al.53 demonstrated a reduction in mito-chondrial biogenesis and protein quality control factorscompared to placebo control, following supplementation ofvitamin C during a 6-week endurance protocol in rats. It isclearly shown that exercise induced mitochondrial massremodeling is a redox-sensitive process. Recently, we foundthat exercise induced mitochondrial ROS may contribute tothe rapidly alteration in mitochondrial fusion/fission proteinexpression.54 Koopman et al.55 reported that vitamin Esupplementation reduced ROS production and restored aber-rant mitochondrial morphology in fibroblasts with mitochon-drial complex I deficiency, suggesting that ROS is involved incontrolling mitochondrial shape in these cells. Using a non-apoptotic concentration of H2O2 in long-term treatment,

originally long and interconnected mitochondrial tubules weretransiently shortened and weakly fragmented.56 In addition,low doses H2O2 in a transient exposure induce mitochondriahyperfuse and form a highly interconnected network in cells.57

Based on the above evidence it can be concluded that ROSinduce remodeling of the mitochondrial network depending onthe concentration of ROS and duration of the treatment. ROSis also an important signal for induction of autophagy. Star-vation-induced autophagy can be suppressed by antioxidantssuppressing the well-known prosurvival function of starvation-induced autophagy.58 Recently, it is demonstrated that exerciseis a potent inducer of autophagy, and that acute and chronicexercise enhances glucose metabolism in mice capable ofinducing autophagy but not in autophagy-deficient mice.59

Several studies have shown that an acute bout of enduranceexercise and stimulatedmuscle contraction can upregulate PGC-1a and activate mitochondrial protein synthesis and prolifera-tion.60 Furthermore, repeated exercise bouts (exercise training)could result in accumulation of PGC-1a, NRF-1, and Tfamprotein levels.61 These observations were thought that PGC-1aplays an important role in mediatingmitochondrial adaptation toexercise, such as elevated respiratory activity, increasedexpression of Krebs cycle enzymes, promoted endogenousantioxidant defense capacity, enhanced fatty acid oxidation, andmitochondrial morphological changes. Garnier et al.62 demon-strated in healthy humans that VO2peak was dependent on

Mitochondrial redox metabolism in aging and exercise 73

coordinated expression of PGC-1a, which was lineally corre-lated to Mfn2 and Drp1 levels in human leg muscle. In C2C12skeletal muscle cells, increasing the production of ROS byincubation with 300 mM H2O2, results in increased PGC-1apromoter activity and PGC-1a mRNA expression requiringthe activation of AMPK.63 Ristow et al.64 demonstratedexercise-induced oxidative stress elevated PGC-1a andPGC-1b expression and ameliorated insulin resistance, whichwere precluded by antioxidants supplementation.

10. Conclusion

A vast body of data has accumulated linking mitochondrialredox metabolism to the aging process. Therefore, the accu-mulation of oxidative damage can be decreased either bylowering the generation of ROS (caloric restriction) or byregular exposure to a small amount of ROS (such as mildexercise) that could result in slight oxidative damage, whichthen leads to up-regulation of antioxidant systems andamelioration of mitochondrial remodeling. Signal transductionby coordinated action of PGC-1, NF-kB, and MAPK (p38)may be potential regulators in these events. These productsinclude (1) antioxidant enzymes (e.g., MnSOD, GPx, GCS);(2) molecules controlling metabolic status and thus ROSproduction, (e.g., UCPs, enzymes in fatty acid and glucosemetabolism); (3) transcription factors for mitochondrialbiogenesis (e.g., PGC-1a, NRF-1, Tfam), and (4) a wide rangeof proteins that could influence mitochondrial dynamicremodeling, such as mitochondrial fusion/fission and auto-phagy proteins. Exercise training is a useful and inexpensiveintervention for preventing aging and degenerative disease, inwhich mitochondrial redox metabolism plays a key role.

Acknowledgments

Thisworkwas supported by research grants from theNationalNatural Science Foundation of China (No. 31110103919,31200894, 31000523, 30771048, 30470837, 31071040, and30270638), Tianjin Municipal Sci-tech-innovation Base Project(No. 10SYSYJC28400), Tianjin Science and TechnologyPlanning Project (No. 12JCQNJC07900), and General Admin-istration of Sport of China Basic Project (No. 10B058).

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