Biogerontology DOI 10.1007/s10522-014-9546-8

REVIEW ARTICLE

Exercise improves mitochondrial and redox-regulated stress responses in the elderly: better late than never! James N. Cobley • Peter R. Moult • Jatin G. Burniston • James P. Morton Graeme L. Close

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Received: 15 September 2014 / Accepted: 5 December 2014 Ó Springer Science+Business Media Dordrecht 2014

Abstract Ageing is associated with several physiological declines to both the cardiovascular (e.g. reduced aerobic capacity) and musculoskeletal system (muscle function and mass). Ageing may also impair the adaptive response of skeletal muscle mitochondria and redox-regulated stress responses to an acute exercise bout, at least in mice and rodents. This is a functionally important phenomenon, since (1) aberrant mitochondrial and redox homeostasis are implicated in the pathophysiology of musculoskeletal ageing and (2) the response to repeated exercise bouts promotes exercise adaptations and some of these adaptations (e.g. improved aerobic capacity and exercise-induced mitochondrial remodelling) offset age-related physiological decline. Exercise-induced mitochondrial remodelling is mediated by upstream signalling events that converge on downstream transcriptional co-factors and factors that orchestrate a co-

J. N. Cobley Division of Sport and Exercise Sciences, Abertay University, Dundee DD1 1HG, UK P. R. Moult School of Science, Engineering and Technology, Abertay University, Dundee DD1 1HG, UK J. G. Burniston
J. P. Morton
G. L. Close (&) Research Institute for Sport and Exercise Science, Liverpool John Moores University, Tom Reilly Building, Byrom St Campus, Liverpool L3 3AF, UK e-mail: [email protected]

ordinated nuclear and mitochondrial transcriptional response associated with mitochondrial remodelling. Recent translational human investigations have demonstrated similar exercise-induced mitochondrial signalling responses in older compared with younger skeletal muscle, regardless of training status. This is consistent with data indicating normative mitochondrial remodelling responses to long-term exercise training in the elderly. Thus, human ageing is not accompanied by diminished mitochondrial plasticity to acute and chronic exercise stimuli, at least for the signalling pathways measured to date. Exerciseinduced increases in reactive oxygen and nitrogen species promote an acute redox-regulated stress response that manifests as increased heat shock protein and antioxidant enzyme content. In accordance with previous reports in rodents and mice, it appears that sedentary ageing is associated with a severely attenuated exercise-induced redox stress response that might be related to an absent redox signal. In this regard, regular exercise training affords some protection but does not completely override age-related defects. Despite some failed redox-regulated stress responses, it seems mitochondrial responses to exercise training are intact in skeletal muscle with age and this might underpin the protective effect of exercise training on age-related musculoskeletal decline. Whilst further investigation is required, recent data suggest that it is never too late to begin exercise training and that lifelong training provides protection against several age-related declines at both the

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molecular (e.g. reduced mitochondrial function) and whole-body level (e.g. aerobic capacity). Keywords Ageing
Exercise training
Skeletal muscle
Stress responses
Mitochondria
Reactive oxygen
Nitrogen species

Introduction Musculoskeletal ageing is characterised by striking deceases in skeletal muscle mass and function, a phenomenon termed sarcopenia (Deschenes 2004; Doherty 2003). This manifests as a decline in muscle cross sectional area, owing to an age-related reduction in both muscle fibre number and size (Doherty 2003; Faulkner et al. 2007; Lexell et al. 1988). There is also an age-related decline in several whole-body physiological variables, such as maximal oxygen uptake (VO2max), endurance performance, insulin sensitivity, muscle power and strength (Heath et al. 1981; Frontera et al. 1991; Deschenes 2004; Tanaka et al. 1997). Ultimately, musculoskeletal ageing often leaves elderly individuals unable to perform everyday tasks and increases their susceptibility to a plethora of agerelated maladies, notably fall events, cardiovascular disease, type II diabetes and cancer (Janssen et al. 2004). The significant socio-economic costs of musculoskeletal ageing, in an expanding elderly population, has emphasised the need to understand how therapeutic interventions, such as exercise training, exert beneficial effects.

Exercise training can attenuate age-related physiological decline Exercise training is a powerful strategy for offsetting age-related physiological decline (Booth et al. 2011). Exercise training can ameliorate the age-related decline in muscle mass, strength, power, endurance and aerobic capacity (Booth et al. 2011; Evans et al. 2005; Pahor et al. 2006; Melov et al. 2007; Menshikova et al. 2006; Pearson et al. 2002; Tanaka and Seals 2008). Of course, the nature of the exercise training undertaken will influence some parameters more than others, with resistance training being generally associated with improving muscle mass and endurance

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training improving endurance and aerobic capacity (Coffey and Hawley 2007). VO2max, a gold standard marker of aerobic capacity, is a useful tool for illustrating the protective effect of exercise training in the elderly. Cross-sectional studies have shown that lifelong endurance training is associated with preserved maximal aerobic capacity, resulting in a difference of *25–30 ml kg-1 min-1 in VO2max values between trained and untrained individuals between the ages of 25–65 years (Booth et al. 2011; Heath et al. 1981; Trappe et al. 1996). Beyond 65 years, VO2max values of *38 ml kg-1 min-1 have been documented in elite octogenarian athletes, this value is comparable to those observed in significantly younger (*20–30 years) untrained persons (Trappe et al. 2013). Longitudinal studies indicate that endurance training significantly improves VO2max in the elderly (Evans et al. 2005; Konopka et al. 2014). Although, physical activity levels are an important determinant of the magnitude of certain age-related physiological declines (Booth et al. 2011; Cobley et al. 2013) some age-related physiological decline is insidious and inevitable. For instance, despite vigorous exercise training masters athletes do not achieve the same performance times as their younger counterparts (Tanaka and Seals 2008). Indicating that there is an age-related deficit that even adherence to rigours exercise training regimes cannot override, at least at the whole-body level (Tanaka and Seals 2008). In attempting to explain this phenomenon at the tissue level, it could be that acute and chronic molecular responses to exercise are impaired in older skeletal muscle and that training does not override this deficit. The basic process of skeletal muscle adaptations is briefly reviewed before relevant molecular responses to acute and chronic exercise are considered in both murine/rodent and human skeletal muscle.

Skeletal muscle adaptation Skeletal muscle is a malleable tissue capable of considerable phenotypical adaptations to repeated exercise stimuli (Egan and Zierath 2013; Fluck 2006). The orchestration of the phenotypical response to repeated exercise stimuli (e.g. the increase in mitochondrial content with repeated endurance training) is extremely complex at the molecular level, requiring the synergistic interaction of multiple

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processes (e.g. signalling and translational processes) each of which is subject to intricate regulation. Current paradigms posit that phenotypical adaptations to exercise training are the culmination of transcriptional and translational responses to a series of acute bouts (Coffey and Hawley 2007; Perry et al. 2010). In this model, cell signalling processes orchestrate transcriptional responses (changes in mRNA abundance) to an acute exercise bout that with repeated bouts, yield differential protein content (Perry et al. 2010). In an ageing context, it could be that this response fails in skeletal muscle and that this could underpin reduced performance in even highly trained elderly individuals. One approach to determine the influence of age and exercise training on the exercise-induced stress response is to assay molecular markers of this response in younger and older, trained and untrained individuals. This review outlines recent developments in this area with particular emphasis on two important stress responses (1) mitochondrial responses associated with mitochondrial biogenesis and (2) redoxregulated stress responses associated with an acute cyto-protective response.

Mitochondrial ageing in skeletal muscle Mitochondria are responsible for producing the majority of cellular ATP (Peterson et al. 2012). Mitochondria are also involved in a plethora of processes beyond energy production, including apoptosis, calcium and redox regulation (Picard et al. 2013; Rizzuto et al. 2012). From an ageing perspective, mitochondrial abnormalities have been implicated in the pathogenesis of musculoskeletal ageing (Hiona and Leeuwenburgh 2008). It is beyond the scope of this review to explore this theory in depth (see Dai et al. 2014; Hiona and Leeuwenburgh 2008; Peterson et al. 2012). In brief, such theories were supported by greater mitochondrial genomic DNA mutation burden (Aiken et al. 2004; Bua et al. 2002, 2006; McKenzie et al. 2002), redox abnormalities (Dai et al. 2014), electron transport chain dysfunction as well as reduced mitochondrial content and function (Lanza and Nair 2010; Lanza et al. 2008; Short et al. 2005) in older compared with younger skeletal muscle. The mitochondrial isolation techniques utilised in some of these studies may, however, have exacerbated certain agerelated changes (e.g. altered calcium handling) since

this method disrupts mitochondrial integrity (Picard et al. 2010, 2011a, b). Further, many of these declines, particularly reduced mitochondrial content and function, appear related to reduced physical activity levels and not biological ageing per se (Cobley et al. 2012; Lanza et al. 2008). Several rodent investigations have, however, documented reduced mitochondrial responses to acute and chronic exercise stimuli in older rodent skeletal muscle (Betik et al. 2009; Debre et al. 2012; Koltai et al. 2012; Ljubicic and Hood 2009a, b; Ljubicic et al. 2009, 2010; Reznick et al. 2007). This phenomenon would suggest that ageing impairs the ability of skeletal muscle to properly adapt to acute and chronic exercise and hence some of the benefits of exercise training might be lost in later life. The process of exercise-induced mitochondrial biogenesis is briefly reviewed before relevant rodent and human evidence is presented.

Exercise-induced mitochondrial biogenesis Mitochondria are constantly being remodelled through the processes of biogenesis, fusion, fission and mitophagy (Detmer and Chan 2007; Westermann 2010). Endurance training stimulates mitochondrial remodelling, culminating in an increase in mitochondrial content and function (Hood 2009, 2001). High intensity training (HIT), brief bouts of supra-maximal exercise, can also induce mitochondrial remodelling responses (Gibala et al. 2012). The exercise-induced increase in mitochondrial content is often termed mitochondrial biogenesis but because an increase in mitochondrial content reflects both mitochondrial degradation and synthesis (Miller and Hamilton 2012), the term mitochondrial remodelling is utilised herein. Mitochondrial remodelling facilitates exercise-induced improvements in VO2max and oxidative metabolism (Holloszy and Coyle 1984). The process of mitochondrial remodelling is exceedingly complex, requiring the co-ordination of nuclear and mitochondrial genomes to effect a change in the expression of over 1,000 genes, ultimately resulting in a change in the abundance of approximately 20 % of all cellular proteins (Lopez-Lluch et al. 2008; Scarpulla 2008; Safdar et al. 2011). The requirement to up-regulate genes from two sub-cellular compartments and then import their protein products into the mitochondrion

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makes mitochondrial remodelling a considerable cellular undertaking (Scarpulla 2008; Wenz 2013). The quest to unravel the molecular processes responsible for exercise-induced mitochondrial remodelling has placed much attention on PGC-1a (Gibala et al. 2012; Hood 2009). Current models of exercise-induced mitochondrial remodelling place PGC-1a at the nexus of a response that translates acute exercise signals (e.g. energy stress) into a coordinated nuclear and mitochondrial response, resulting in the up-regulation of mitochondrial proteins with repeated activation (Safdar et al. 2011). The response of PGC-1a to an acute exercise bout typifies this paradigm (see Fig. 1), with transient mRNA increases following acute sessions leading to increased PGC-1a protein content following exercise training (Baar et al. 2002; Burgomaster et al. 2008; Perry et al. 2010; Pilegaard et al. 2003; Russell et al. 2003). Analogously, the response of PGC-1a target genes follows this temporal pattern, with COXIV and b-HAD responding similarly (Perry et al. 2010). It is postulated that this increase in PGC-1a mRNA and protein

content accelerates the rate of exercise-induced mitochondrial remodelling (Perry et al. 2010). Activation of PGC-1a and its redistribution from the cytoplasm to both the nucleus (Little et al. 2010a, b; Wright et al. 2007) and mitochondria (Safdar et al. 2011) could underlie the increase in PGC-1a target genes following an acute bout of exercise. Activation of PGC-1a involves several post-translational modifications; in particular, AMPK and p38 MAPK mediated phosphorylation allied to sirtuin mediated deacetylation are important (Jager et al. 2007; Fernadnez-Marcos and Auwerx 2011; Rodgers et al. 2005) but not essential (Philp et al. 2011). We have presented a PGC-1a centric model of exercise-induced mitochondrial remodelling because several relevant ageing studies in this area are conceptually grounded in this model. This is likely because over-expression of PGC-1a in skeletal muscle is sufficient to prevent muscle wasting by reducing apoptosis, autophagy and proteasome degradation and also improves insulin sensitivity and preserves the neuromuscular junction (Sandri et al. 2006; Wenz et al. 2009), rendering PGC-1a an attractive therapeutic target. It is, however, emphasised that PGC-1a is dispensable for exercise-induced mitochondrial remodelling (Leick et al. 2008; Rowe et al. 2012) and that several other proteins (e.g. p53 Bartlett et al. 2014; Saleem and Hood 2013) and processes (e.g. epigenetic processes, such as histone modification [Konopka and Nair 2013]) contribute to mitochondrial remodelling, indeed seldom do biological processes rely on one or two proteins (Kholodenko et al. 2010; Timmons 2011).

Failed signalling responses associated with mitochondrial remodelling in aged rodents

Fig. 1 A schematic of exercise-induced PGC-1a mRNA expression a typical mitochondrial remodelling related stress response. Exercise stress activates signalling proteins upstream of PGC-1a, namely: p38 MAPK, SIRT1, AMPK. P38 MAPK and AMPK phosphorylate PGC-1a, priming PGC-1a for SIRT1 mediated deactylation. AMPK also phosphorylates histone deacteylase (HDAC) and cAMP response element binding protein (CREB) enabling these proteins to bind to the PGC-1a promoter and help activate myocyte enhancer factor 2 (MEF2). PGC-1a trans-locates to the nucleus and co-activates MEF-2 to up-regulate transcription of its own gene leading to an increase in PGC-1a mRNA post-exercise

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It is apparent that in adult human skeletal muscle acute endurance or HIT exercise stimulates a signalling response associated with exercise-induced mitochondrial remodelling (Egan and Zierath 2013; Gibala et al. 2012). Rodent models suggest that mitochondrial signalling responses associated with exercise-induced mitochondrial remodelling are defective in aged (*26 months) compared with adult (*6 months) rodent muscle (Betik et al. 2009; Debre et al. 2012; Koltai et al. 2012; Ljubicic and Hood 2009a, b; Ljubicic et al. 2009, 2010; Reznick et al. 2007 see Table 1). Reznick et al. (2007) reported blunted

Biogerontology Table 1 Summary of acute (one session) and short-term (B1 week) exercise studies documenting defective exercise adaptations in aged rodents Study

Species

Exercise

Muscle

Key findings

Reznick et al. (2007)

F344XBN rats

10 min treadmill run at 10 m/min per day for 4 days, then an incremental exhaustion run on the fifth day

Extensor digitorum longus (fast twitch)

Exercise was associated with an increase in AMPK (T172) phosphorylation in young but not aged rodents

In situ stimulation constant application of 1 Hz for 5 min

Red Gastrocnemius (slow twitch), white Gastrocnemius (fast twitch)

Diminished exercise-induced p38 MAPK phosphorylation in older compared with younger rats, in fast twitch but not slow twitch muscle

Ljubicic and Hood (2009a)

Ljubicic et al. (2009)

F344XBN rats

F344XBN rats

Electrical stimulation (10 Hz, 3 h/day for 7 days)

Anterior Tibialis (fast twitch)

Exercise was associated with an increase in acetyl-CoA carboxylase (S79) phosphorylation in young but not aged rodents

Diminished exercise-induced AMPK phosphorylation in older compared with younger rats, in fast twitch but not slow twitch muscle The magnitude of exercise-induced improvements in mitochondrial function was lower in older compared with young animals Short-term exercise training increased PGC-1a and SIRT1 protein expression in young and old skeletal muscle but the magnitude of this increase was lower in older skeletal muscle Expression of mitochondrial import proteins was blunted in older compared with younger animals following exercise

phosphorylation of AMPK, and its downstream substrate acetyl-CoA carboxylase, following short-term exercise and pharmacological interventions in aged compared with adult rodent fast-twitch muscle. These findings were extended by Ljubicic and Hood (2009a), who observed defective AMPK and p38 MAPK phosphorylation, a proxy marker of kinase activation, in aged fast twitch but not slow twitch rodent skeletal muscle following contractions. These findings are intriguing given the greater metabolic disturbances and hence exercise signal following contractions in fast twitch compared to slow twitch muscle (Ljubicic and Hood 2009a, b). They also highlight the perils of global statements since defective stress responses are not uniform across muscles (Ljubicic and Hood 2009a, b). Further, none of these studies united defective signalling with transcriptional and translational outcomes to an acute exercise bout. It is, therefore, difficult to properly appraise the functional impact of this phenomenon, particularly when it is considered

that a signalling response can be transduced even in the face of reduced kinase activation (Crozier et al. 2005). One way to appraise the functional impact of blunted acute mitochondrial remodelling responses is to assay proxy markers (e.g. PGC-1a protein content) of this process following short or long-term exercise training, since the acute response to repeated exercise bouts regulates certain exercise adaptations (Egan and Zierath 2013; Perry et al. 2010). Using this approach, Ljubicic et al. (2009) demonstrated that whilst aged rodent (36 months old, F344XBN strain) muscle upregulated PGC-1a, SIRT1 and COX activity in response to short-term electrical stimulation (7 days), the magnitude of these gains were less than the changes observed in adult (6 months old) skeletal muscle, culminating in a 50 % enhancement of endurance in young compared with aged animals post-training. Decrements in skeletal muscle adaptability could be rodent strain specific. In this regard,

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Koltai et al. (2012) demonstrated that chronic treadmill training in Wistar rats minimised, but did not completely reverse, blunted PGC-1a related responses in aged compared with young skeletal muscle. Indeed, one study reported that exercise training completely failed to increase PGC-1a protein levels in aged Wistar rats (Debre et al. 2012). It is possible that lifelong training could protect against impaired mitochondrial modelling responses in elderly rodents. When this possibility was addressed, Betik et al. (2009) found that continued endurance training during the transition from late middle (29 months) to old (34–36 months) age did not prevent the age-related decline in muscle oxidative capacity or PGC-1a protein levels in fisher 344 brown-norway rats. Collectively, these results suggest that signalling responses associated with mitochondrial remodelling are aberrant in aged rodent skeletal muscle (Betik et al. 2009; Debre et al. 2012; Koltai et al. 2012; Ljubicic et al. 2009, 2010).

Maintenance of normative mitochondrial remodelling related responses to acute and chronic exercise stimuli in elderly humans Rodent models permit an invasive and mechanistic analysis of signalling associated with mitochondrial remodelling but may not translate fully to the human situation. Whilst, there are several similarities between rodent and human models of ageing there are also some telling differences (Rennie et al. 2010), highlighting the need to translate rodent findings into the human situation. Recent investigations have attempted to do this (Bori et al. 2012; Cobley et al. 2012; Iversen et al. 2011) and delineate the influence of age and training status. Iversen et al. (2011) were the first to examine signalling associated with mitochondrial remodelling to an acute exercise bout in elderly trained and untrained volunteers. These authors observed increased p38 MAPK and AMPK phosphorylation post-exercise in both groups following a continuous exercise bout at 75 % VO2max performed to volitional exhaustion. The phosphorylation of these signalling proteins was associated with a concomitant increase in PGC-1a mRNA 2 h postexercise. These findings indicate that elderly untrained individuals are still able to mount a normal signalling response to acute exercise despite reduced

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mitochondrial content and function at baseline (Iversen et al. 2011). Whilst the Iversen et al. (2011) study represented an important advance, it did not enable the effect of age to be fully delineated since this requires the comparison of the responsiveness of both young and old groups. This was addressed by Cobley et al. (2012) who employed a four group design to determine the influence of age and training status on the transcriptional and translational response of molecules involved in mitochondrial remodelling following an acute exercise bout. These authors replicated and extended the findings of Iversen et al. (2011) reporting that signalling (p38 MAPK phosphorylation) and transcriptional responses (PGC-1a and COXIV mRNA) associated with mitochondrial remodelling were similar in older compared with younger individuals, irrespective of training status. It would appear, therefore, that there are no intrinsic age-related deficits in the ability to mount a mitochondrial stress response (at least for the molecules investigated to date) to an acute exercise bout; as indicated by similar up (signalling protein activation) and downstream (transcriptional responses) responses (Cobley et al. 2012; Iversen et al. 2011). The data of Bori et al. (2012) do, however, appear to challenge this notion. These authors observed a blunted transcriptional response, as manifested by reduced fission 1 and PGC-1a mRNA levels, in older compared to younger individuals following an acute bout of exercise, irrespective of training status. At first glance, this result is difficult to reconcile, however, these authors assayed gene expression 10–15 min post-exercise a time-point when the contractile signal (e.g. kinase phosphorylation) that precedes the transcriptional response is being transduced. A cogent argument for determining gene expression in the hours following exercise exists given PGC-1a mRNA is significantly up-regulated 3 h post-exercise in response to several exercise intensities (Nordsborg et al. 2010). Further, Mahoney et al. (2005) reported that 118 exercise-regulated gene transcripts were significantly altered 3 h post-exercise. The finding that young untrained subjects were able to increase PGC-1a mRNA at this time-point represents something of an anomaly and is not readily explainable (Bori et al. 2012), but could be related to rapid phosphorylation of p38 MAPK and AMPK during the exercise bout and hence more immediate mRNA

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responses. Nevertheless, several pertinent changes in gene expression could have been missed in this study due to the selection of a time-point that does not allow the transcriptional response to be completely resolved. In contrast to previous studies in rodents, it seems that signalling associated with mitochondrial remodelling is intact in older trained and untrained human skeletal muscle (Cobley et al. 2012; Iversen et al. 2011). It should be noted that human investigations have assayed the Vastus Lateralis a mixed fibre-type muscle (Godin et al. 2010) whereas rodent investigations documented defects in fast twitch muscle, so the lack of translation could be an artefact of the muscle sampled. Further measurements of mitochondrial protein synthesis and degradation alongside proxies of mitochondrial remodelling (e.g. translational responses) are required to fully elucidate the mitochondrial remodelling response to exercise (Konopka and Nair 2013; Miller and Hamilton 2012). Future researchers are encouraged to investigate mitochondrial protein dynamics in response to acute and chronic exercise interventions in elderly humans. Nevertheless, the notion that acute responses are intact in aged skeletal muscle, irrespective of training status, is consistent with similar molecular mitochondrial responses to exercise training in older compared with younger individuals (Ghosh et al. 2011; Konopka and Nair 2013; Konopka et al. 2014; Short et al. 2003). For instance, similar increases in molecular makers of mitochondrial remodelling (e.g. PGC-1a protein levels, electron transport chain activity and protein content) were observed in older compared to younger human skeletal muscle following 12 weeks of exercise training (Konopka et al. 2014). Overall, it appears that ageing does not impair molecular mitochondrial remodelling related responses to either acute or chronic exercise training. Further, it is consistent with the protective effect of regular exercise training on the age-related decline in molecular markers of mitochondrial content and function in older trained human skeletal muscle (Lanza et al. 2008; Joesph et al. 2012; Safdar et al. 2010b; Trappe et al. 2013). From a practical perspective, it is clear that (1) signalling responses associated with mitochondrial remodelling are intact in older untrained persons and (2) chronic training, particularly lifelong training, preserves proxies of mitochondrial function and content in later life. Exercise training should, therefore, be maintained across the lifespan and when discontinued should be

recommenced in later life to offset age-related physiological decline in mitochondrial content and function.

Redox homeostasis in skeletal muscle Reactive oxygen and nitrogen species (RONS) are highly reactive, short-lived species (half-life: milliseconds) that are perpetually produced by cells (Gutteridge and Halliwell 2010; Winterbourn 2008). Superoxide and nitric oxide are the two primary RONS generated in skeletal muscle (Sakellariou et al. 2014). In skeletal muscle, nitric oxide synthase (NOS) enzymes, mitochondria (predominately complexes 1 and 3 of the electron transport chain) and NADPH oxidases are the principle generators of the aforementioned parent radicals (Sakellariou et al. 2014). Metabolism of these two parent radicals yields several oxygen or nitrogen centred radical (e.g. peroxynitrite) and non-radical species (e.g. hydrogen peroxide), hence the term RONS (Halliwell and Gutteridge 2007). RONS are heterogeneous chemical entities having distinct reaction profiles, metabolism and functions (Murphy et al. 2011). As an example, the hydroxyl radical is extremely labile, reacting with the first biomolecule it encounters, whereas nitric oxide is deliberately manufactured by NOS isoforms, is relatively stable being unreactive with cellular biomolecules (save other RONS [e.g. superoxide, Carballal et al. 2014]) and is associated with several physiological functions (e.g. inhibition of platelet aggregation and cGMP signalling [Martinez-Ruiz et al. 2011]). RONS levels are regulated by an intricate network of exogenous (diet-derived vitamins [e.g. vitamin C]) and endogenous antioxidants, principally antioxidant enzymes and glutathione (Halliwell and Gutteridge 2007; Powers and Jackson 2008). Redox homeostasis describes the balance between the generation and removal of RONS. Altered redox homeostasis regulates several physiological (e.g. muscle contraction) and pathological processes (e.g. neurodegeneration). Indeed, aberrant redox homeostasis has been suggested to regulate the basic process of ageing (Harman 1956). It is beyond the scope of this review to outline free radical theories of ageing (see Dai et al. 2014; Jang and Van Remmen 2009; Salmon et al. 2010; Stuart et al. 2014). Save to say that the theory in its most simple manifestation (i.e. oxidative damage

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causes ageing) is likely incorrect and that the role of RONS in ageing is far more complex than initially thought (Jang and Van Remmen 2009; Salmon et al. 2010). Instead, the exercise-induced redox stress response with specific emphasis on human ageing is reviewed herein.

The redox-regulated response of skeletal muscle to acute exercise Skeletal muscle contractions are associated with a transient increase in RONS generation (Close et al. 2005; Palomero et al. 2008; Pye et al. 2007; Sakellariou et al. 2013). Traditionally, it was believed that exerciseinduced increases in RONS production were purely deleterious (Jackson 2009). The realisation that reversible cysteine based modifications, including but not limited to: disulfide formation, S-glutathionation and S-nitrosylation, plays an important role in transducing beneficial adaptations revolutionised the field (Forman et al. 2014; Holmstrom and Finkel 2014; JanssenHeininger et al. 2008; Sies 2014; Winterbourn 2014a). It now appears that RONS are upstream signals implicated in the activation of redox-sensitive transcription factors (e.g. HSF-1, NF-KB and AP-1) that promote cyto-protective responses (Brooks et al. 2008; Brigelius-Flohe and Flohe 2011; Irrcher et al. 2009; Jackson 2009; Ji et al. 2006). This is associated with increased DNA binding of redox-sensitive transcription factors post-exercise (Ji et al. 2004, 2006; Vasilaki et al. 2006a, b). The acute cyto-protective response manifests as increased abundance and activity of antioxidant enzymes and heat shock proteins (HSPs), proteins that help protect against the potential cyto-toxic outcomes of RONS generation (Jackson and McArdle 2011; Morton et al. 2009). Increased antioxidant enzyme and HSP protein levels are often evident after a single bout of exercise (Cobley et al. 2014; Khassaf et al. 2001; McArdle et al. 2001; Morton et al. 2006).

Age-related aberrant redox homeostasis in skeletal muscle Ageing is associated with aberrant redox homeostasis (Dai et al. 2014; Jang and Van Remmen 2011; Stuart et al. 2014). Defining the nature of age-related redox perturbations is technically challenging, owing to

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difficulties associated with measuring RONS directly (Halliwell and Whiteman 2004). This has fostered a reliance on biochemical footprints of RONS generation, such as lipid peroxidation levels (Halliwell and Whiteman 2004). Several investigations report an elevation in global levels of oxidatively modified lipid, protein and DNA adducts in older skeletal muscle (Beltran Valls et al. 2014; Drew et al. 2003; Cobley et al. 2014; McDonagh et al. 2014; Radak et al. 2011; Safdar et al. 2010b). This body of indirect evidence indicates that ageing is associated with increased RONS generation. This interpretation is complicated by the fact that increased levels of oxidatively modified adducts could reflect differential repair of the modification (Murphy et al. 2011) or indeed dietary changes (Halliwell and Gutteridge 2007). Fluorescent probes can directly assay the generation of certain RONS (Kalyanaraman et al. 2012; Winterbourn 2014b). Recent application of this approach has revealed an increase in DCF fluorescence, a general redox indicator (Kalyanaraman et al. 2012), at rest in older but not younger isolated murine skeletal muscle fibres (Palomero et al. 2013). Other studies have utilised Amplex Red, a hydrogen peroxide probe, and reported that mitochondrial hydrogen peroxide generation is not increased in older permeabilised skeletal muscle fibres (Gousillou et al. 2014). In contrast, Bailey et al. (2010) documented increased intramuscular RONS generation in older human skeletal muscle using an ex vivo ESR spin-trapping technique, the potential toxicity of this method limits in vivo application in humans (Halliwell and Whiteman 2004). Overall, despite some inconsistencies that are probably related to the reactive species (e.g. muscle lipid-derived alkoxyl-alkyl radicals [Bailey et al. 2010]) vs mitochondrial hydrogen peroxide (e.g. Gousillou et al. 2014) assayed and methodological techniques, it appears that ageing is associated with redox disturbance.

Failed redox-regulated stress responses to acute exercise in older mice and rodents Clearly, analytical limitations have hindered attempts to unravel the nature of the redox perturbations that accompany ageing in human skeletal muscle. Despite this age-related redox disturbance has important implications for the exercise-induced redox stress

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response in mice and rodents (Jackson and McArdle 2011; Kayani et al. 2008). It appears that sedentary ageing, predominately in mice, recapitulates the exercise-induced redox stress response at rest (Jackson and McArdle 2011). This is manifested by constitutive AP-1, HSF-1 and NF-KB activation with concomitant antioxidant enzyme and HSP up-regulation in quiescent rodent and murine skeletal muscle (Vasilaki et al. 2002, 2003, 2006a, see Table 2). As a consequence of this elevated basal stress, muscles from old mice and rodents display a significantly attenuated ability to further increase RONS production and hence fail to up-regulate HSPs and antioxidant enzymes after an acute exercise challenge (Vasilaki et al. 2002, 2003, 2006a). This is consistent with the notion that resting values determine the magnitude of exercise-induced oxidative stress (Margaritelis et al. 2014). Aged rodent and murine skeletal muscle is unable to further upregulate HSPs, antioxidant enzyme activity and content following an acute exercise challenge, largely owing to increased baseline expression of these proteins (Kayani et al. 2008; Jackson and McArdle 2011; Vasilaki et al. 2002, 2003, 2006a, b). For instance, Vasilaki et al. (2003) reported that aged mice were unable to up-regulate HSP25 mRNA levels and protein content following exercise. Subsequent studies extended and replicated these findings to other HSPs and antioxidant enzymes, notably HSP70 cognate isoform (Vasilaki et al. 2002, 2006a). Additionally, the failure to up-regulate HSPs and antioxidant enzyme activity was due to an inability to completely activate redox-sensitive transcription factors postexercise (Vasilaki et al. 2006b). From a mechanistic perspective, these findings seem intrinsically connected to resting adaptations that obviate the need to further adapt to exercise stress; coupled to failed transmission of an exercise induced redox signal defined by reduced transcription factor DNA binding, likely owing to depressed exercise-induced RONS generation (Close et al. 2007; Palomero et al. 2013; Vasilaki et al. 2006b).

The redox regulated stress response to acute exercise in humans Based on findings from mice and rodents, it could be predicted that sedentary human ageing is associated with perturbed redox homeostasis that necessitates the

induction of a cyto-protective response in quiescent skeletal muscle (Jackson and McArdle 2011). Sedentary human ageing is, however, associated with increased protein carbonylation but not HSP (a-Bcrystallin, HSP27 and HSP70) up-regulation in nonsarcopenic and sarcopenic skeletal muscle (Beltran Valls et al. 2014). HSP72 (the inducible isoform) but not HSP27 is up-regulated with sedentary ageing in untrained elderly persons (Cobley et al. 2014). Further work examining a greater complement of HSPs is required before more definitive conclusions can be made. The effects of sedentary ageing on antioxidant enzyme content is variable, with levels of some being reduced (SOD2), unchanged (SOD1, GPx-1) or increased (PRX5, eNOS) with age in human skeletal muscle (Cobley et al. 2014; Nyberg et al. 2012, 2014; Safdar et al. 2010a, b). There is also discrepancy between reports with nNOS and CAT being upregulated in one study (Safdar et al. 2010b) but not others (Cobley et al. 2014; Nyberg et al. 2014), this is probably related to subject differences and the use of semi-quantitative measurement techniques (i.e. western blotting McGlory et al. 2014). In general, it appears that sedentary ageing alters antioxidant enzyme levels suggesting perturbed redox homeostasis, which is consistent with greater levels of oxidatively modified lipid, DNA and protein adducts in these individuals (Beltran Valls et al. 2014; Cobley et al. 2014; Radak et al. 2011; Safdar et al. 2010b). The effects of sedentary ageing on redox homeostasis are likely better resolved with measurements of protein function, such as enzymatic activity and post-translational modification (McDonagh et al. 2014). In support of this, SOD2 nitration (Safdar et al. 2010a), a modification that inactivates this enzyme (Demicheli et al. 2007), is increased with sedentary ageing, suggesting a reduced ability to metabolise superoxide and generate hydrogen peroxide for signalling. Exercise training appears to protect against the agerelated increase in oxidatively modified lipid, proteins and DNA (Cobley et al. 2014; Radak et al. 2011; Safdar et al. 2010b). In particular, 8-oxoguanine (a modified DNA adduct) levels are lower in trained compared with untrained elderly individuals, suggesting that exercise training might afford protection against age-related genomic stress (Cobley et al. 2013; Radak et al. 2011). However, this postulate is speculative at present and requires further research (Cobley et al. 2013). To the best of our knowledge, only

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Biogerontology Table 2 Summary of studies that have demonstrated defective up-regulation of HSPs and antioxidant enzymes in the skeletal muscle of aged animals following an acute exercise bout Study

Species

Exercise

Muscle

Key findings

Vasilaki et al. (2002)

Wistar Rats

15 min isometric contractions (0.5 s 60 V pulse every 5 s)

Gastrocnemius (predominately fast twitch)

No difference in basal HSP70 (72) and HSC70 protein content between adult and aged rodents

15 min isometric contractions (0.5 s 60 V pulse every 5 s)

Tibialis anterior (fast twitch)

15 min isometric contractions (0.5 s 60 V pulse every 5 s)

Gastrocnemius and tibialis anterior

Vasilaki et al. (2003)

Vasilaki et al. (2006a)

B6XSJL mice

B6XSJL mice

HSP70 increased in adult but not aged rodents postexercise HSC70 was not exercise responsive in either group HSP25 (27) mRNA increased post-exercise in adult but not aged mice HSP25 protein content increased post-exercise in adult but not aged mice These findings were not attributable to differential HSF1 binding Resting HSP25 protein content was not influence by age HSC70 was up-regulated in aged but not adult mice at rest SOD and catalase activity was up-regulated at rest in aged but not adult muscle Aged animals failed to up-regulate SOD1 and catalase activity and SOD1 protein content following exercise Despite elevated resting NF-KB and AP-1 DNA binding aged muscle was unable to further increase DNA binding of these transcription factors following exercise

Cobley et al. (2014) have evaluated HSP abundance in older trained skeletal muscle, documenting greater HSP72 but not HSP27 protein abundance. Greater protein levels of HSP72 appeared to be age-related since this was also evident in the untrained elderly. Greater PRX5, nNOS, eNOS and SOD2 levels have been reported in older trained compared with untrained skeletal muscle (Cobley et al. 2014; Nyberg et al. 2014; Safdar et al. 2010b). Nitrated SOD2 levels are also lower in trained compared with untrained elderly individuals (Safdar et al. 2010b). The functionality of these changes are unknown, but we speculate that they protect against age-related redox disturbance. We also speculate that exercise training increases SOD2 and PRX5 levels, enzymes that metabolise superoxide (SOD2), hydrogen peroxide (PRX5) and peroxynitrite (PRX5), providing protection against elevated SOD2 nitration and protein nitration that is evident with sedentary human (Cobley et al. 2014; Safdar et al. 2010b, see Fig. 2) and indeed murine ageing (Pearson et al. 2014). Future research in this area should further explore the redox-related

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changes, with particular attention to RONS generation and redox-regulated transcription factor binding, that accompany exercise training in the elderly. To the best of our knowledge, only Cobley et al. (2014) have evaluated the exercise-induced redox stress response in young and old, trained and untrained individuals. These authors found that sedentary ageing altered the redox-regulated stress response, resulting in limited HSP72, SOD2 and CAT up-regulation and PRX5 down-regulation (Cobley et al. 2014). In contrast, exercise trained individuals were able to mount a normal HSP72 and CAT response to an acute exercise bout, suggesting exercise training preserves some redox-regulated stress responses (Cobley et al. 2014). Exercise training was, however, still associated with an abnormal SOD2 and PRX5 response, suggesting the existence of age-related defects that exercise training cannot override (Cobley et al. 2014). These findings broadly support previous work in mice and rodents documenting aberrant redox regulated stress responses (Jackson and McArdle 2011). Mechanistically, aberrant redox regulated

Biogerontology

Fig. 2 A theoretical model of the protective effect of exercise training on age-related protein nitration. In this model, greater SOD2 protein levels in older trained individuals mitigates against the proposed age-related increase in mitochondrial peroxynitrite production, as favoured by increased eNOS protein levels, resulting in lower 3-NT levels. 3-NT is a biomarker of protein nitration and proxy marker of peroxynitrite production. It is speculated that greater PRX5 function, as

indicated by greater peroxynitrite (ONOO) and lower 3-NT levels, in old trained (OT) compared with old untrained (OU) individuals contributes to this effect, since PRX5 is a peroxynitrite reductase. This model is based on the observations of Cobley et al. (2014) and Safdar et al. (2010a). O2- denotes superoxide; H2O2 denotes hydrogen peroxide and NO denotes nitric oxide

investigation. In support of this, Nyberg et al. (2014) recently demonstrated the exercise-induced increase in venous GSSG was absent in elderly individuals despite greater muscular NADPH oxidase expression, irrespective of training status. It is, however, clear that further work is necessary to validate our hypothesis that aberrant redox regulated stress responses to acute exercise are related to lower RONS generation during exercise in older compared with younger humans. Fig. 3 A theoretical model of the redox-regulated stress response to an acute exercise bout in young, old trained and old untrained individuals. In young individuals (irrespective of training status), exercise is accompanied by RONS generation, thiol oxidation and transcription factor binding resulting in HSP and AOX up-regulation. In old trained individuals, it is suggested that exercise is accompanied by reduced RONS generation, thiol oxidation and transcription factor binding resulting in only some HSP and AOX up-regulation. In old untrained individuals, it is suggested that exercise is accompanied by minimal RONS generation, thiol oxidation and transcription factor binding resulting in no significant HSP and AOX up-regulation. It is emphasised that several aspects of this model require further investigation in humans

stress responses might be related to blunted exerciseinduced RONS production, reduced thiol oxidation and redox-sensitive transcription factor DNA binding (see Fig. 3). This possibility warrants further

Conclusion Overall, molecular proxies associated with acute and chronic exercise-induced mitochondrial remodelling are similar in older compared to younger skeletal muscle, irrespective of training status. Exercise training is, therefore, a potent therapeutic strategy for offsetting age-related mitochondrial decline. Sedentary ageing is associated with redox imbalance, as evidenced by the accumulation of RONS-modified DNA, lipid and protein adducts, and a general failure of the redox-stress response to acute exercise, which may disrupt the adaptive response to acute exercise. In contrast, active ageing affords some protection against the accumulation of RONS-modified DNA, lipid and

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Biogerontology

protein adducts, and partially maintains redox stress responses following acute exercise. Further research is needed to better understand the functional impact of failed redox-regulated stress responses in the elderly and whether this is owing to aberrant exercise-induced RONS generation. In conclusion, whilst exercise training will never completely stop ageing it is, at present, our best weapon for offsetting age-related physiological decline; so perhaps it really is a case of ‘better late than never’. Acknowledgments Age UK are thanked for generous financial support. JNC and PRM would like to acknowledge financial support provided by the Carnegie Trust and Abertay University. Conflict of interest No conflicts of interest, financial or otherwise, are declared by the authors.

References Aiken J, Bau E, Cap Z, Lopez M, Wagangat J, McKiernan S (2004) Mitochondrial DNA deletion mutations and sarcopenia. Ann N Y Acad Sci 959:412–423 Baar K, Wende AR, Jones TE, Marison M, Nolte LA, Chen M et al (2002) Adaptations of skeletal muscle to exercise: rapid increase in transcriptional co-activator PGC-1. FASEB J 16:1879–1886 Bailey DM, McEneny J, Mathieu-Costello O, Henry RR, James PE, McCord JM et al (2010) Sedentary aging increases resting and exercise-induced intramuscular free radical formation. J Appl Physiol 109:449–456 Bartlett JD, Close GL, Drust B, Morton JP (2014) The emerging role of p53 in exercise metabolism. Sports Med 44:303–309 Betik AC, Thomas MM, Wright KJ, Riel CD, Hepple RT (2009) Exercise training from late middle age until senescence does not attenuate the declines in skeletal muscle aerobic function. Am J Physiol Regul Integr Comp Physiol 297:R744–R755 Booth FW, Laye MJ, Roberts MD (2011) Lifetime sedentary living accelerates some aspects of secondary aging. J Appl Physiol 111:1497–1504 Bori Z, Zhao Z, Kolati E, Fatouros IG, Jamurtas AZ, Dourousdos II et al (2012) The effects of aging, physical training, and a single bout of exercise on mitochondrial protein expression in human skeletal muscle. Exp Gerontol 47:417–424 Brigelius-Flohe R, Flohe L (2011) Basic principles and emerging concepts in the redox control of transcription factors. Antioxid Redox Signal 15:2335–2381 Brooks SV, Vasilaki A, Larkin LM, McArdle A, Jackson MJ (2008) Repeated bouts of aerobic exercise lead to reductions in free radical generation and nuclear kB activation. J Physiol 596:3979–3990

123

Bua EA, McKeirnan SH, Wanagat J, McKenzie D, Aiken J (2002) Mitochondrial abnormalities are more frequent in muscles undergoing sarcopenia. J Appl Physiol 92:2617–2624 Bua EA, Johnson J, Herbst A, Delong B, McKenzie D, Salmat S, Aiken JM (2006) Mitochondrial DNA mutations accumulate to detrimental levels in aged human muscle fibres. Am J Hum Genet 79:469–480 Burgomaster KA, Howarth KR, Phillips SM, Rakobowchuk M, MacDonald MJ, McGee SL et al (2008) Similar metabolic adaptations during exercise after low volume sprint interval and traditional endurance training in humans. J Physiol 586:151–160 Carballal S, Bartesaghi S, Radi R (2014) Kinetic and mechanistic considerations to assess the biological fate of peroxynitrite. Biochim Biophys Acta 1840:768–780 Close GL, Ashton T, McArdle A, Jackson MJ (2005) Microdialysis studies of extracellular reactive oxygen species production in skeletal muscle: factors influencing the reduction of cytochrome c and hydroxylation of salicylate. Free Radic Biol Med 39:1460–1467 Close GL, Kayani AC, Ashton T, McArdle A, Jackson MJ (2007) Release of superoxide from skeletal muscle of adult and old mice: an experimental test of the reductive hotspot hypothesis. Aging Cell 6:189–195 Cobley JN, Bartlett JD, Kayani AC, Murray SW, Louhelainen J, Donovan T et al (2012) PGC-1a transcriptional response and mitochondrial adaptation to acute exercise is maintained in skeletal muscle of sedentary elderly males. Biogerontology 13:621–631 Cobley JN, Sakellariou GK, Waldron S, Murray G, Burniston JG, Morton JP et al (2013) Life-long training attenuates residual genotoxic stress in the elderly. Longev Healthspan 2:11 Cobley JN, Sakellariou GK, Owens DJ, Murray S, Waldron S, Gregson W et al (2014) Lifelong training preserves some redox-regulated adaptive responses following an acute exercise stimulus in aged human skeletal muscle. Free Radic Biol Med 70:23–32 Coffey VG, Hawley JA (2007) The molecular basis of training adaptation. Sports Med 37:737–763 Crozier SJ, Kimball SR, Emmert SW, Anthon JC, Jefferson LS (2005) Oral leucine administration stimulates protein Synthesis in rat skeletal muscle. J Nutr 135:376–382 Dai D, Chia YA, Marcinek DJ, Szeto HH, Rabinovtich PS (2014) Mitochondrial oxidative stress in aging and healthspan. Longev Healthspan 3:6 Debre F, Gomez-Cabrera MC, Nasciemnto AL, Sanchis-Gomar F, Martinez-Bello VE, Tresguerres JAF et al (2012) Age associated low mitochondrial biogenesis may be explained by a lack of response of PGC-1a to exercise training. Age 34:669–679 Demicheli V, Quijano C, Alvarez B, Radi R (2007) Inactivation and nitration of human superoxide dismutase (SOD) by fluxes of nitric oxide and superoxide. Free Radic Biol Med 42:1359–1368 Deschenes MR (2004) Effects of aging on muscle fibre type and size. Sport Med 34:809–824 Detmer SA, Chan DC (2007) Functions and dysfunctions of mitochondrial dynamics. Nat Rev Mol Cell Biol 8:870–879

Biogerontology Doherty TJ (2003) Invited review: aging and sarcopenia. J Appl Physiol 111:1717–1727 Drew B, Phaneuf S, Driks A, Selman C, Gredilla R, Lezza A et al (2003) Effects of aging and caloric restriction on mitochondrial energy production in Gastrocnemius muscle and heart. Am J Physiol Regul Integr Comp Physiol 284:R474–R480 Egan B, Zierath JR (2013) Exercise metabolism and the molecular regulation of skeletal muscle adaptation. Cell Metab 17:162–184 Evans EM, Racette SB, Petersen LR, Villareal DT, Greiwe JS, Holloszy JO (2005) Aerobic power and insulin action improve in response to endurance exercise training in healthy 77–78 year olds. J Appl Physiol 98:40–45 Faulkner JM, Larkin LM, Claflin DR, Brooks SV (2007) Agerelated changes in the structure and function of skeletal muscles. Proc Aust Physiol Soc 38:69–75 Fernadnez-Marcos PJ, Auwerx J (2011) Regulation of PGC-1a, a nodal regulator of mitochondrial biogenesis. Am J Clin Nutr 93:884S–890S Fluck M (2006) Functional, structural and molecular plasticity of mammalian skeletal muscle in response to exercise stimuli. J Exp Biol 209:2239–2248 Forman HJ, Davies KJA, Ursini F (2014) How do nutritional antioxidants really work: nucleophilic tone and para-hormesis versus free radical scavenging in vivo. Free Radic Biol Med 66:24–35 Frontera WR, Hughes VA, Lutz KJ, Evans WJ (1991) A crosssectional study of muscle strength and mass in 45–78 year old men and women. J Appl Physiol 71:644–650 Ghosh S, Lertwattanarak R, Lefort N, Molina-Carrion M, JoyaGaleana J, Bowen BP et al (2011) Reduction in reactive oxygen species production by mitochondria from elderly subjects with normal and impaired glucose tolerance. Diabetes 60:2051–2060 Gibala M, Little JP, MacDonald MJ, Hawley JA (2012) Physiological adaptations to low-volume, high-intensity interval training in health and disease. J Physiol 590:1077–1084 Godin R, Ascah A, Daussin FN (2010) Intensity-dependent activation of intracellular signaling pathways in skeletal muscle: role of fibre type recruitment during exercise. J Physiol 588:4073–4074 Gousillou G, Sgarioto N, Kapchinksy S, Purves-Smith F, Norris B, Pion CH et al (2014) Increased sensitivity to mitochondrial permeability transition and myonuclear translocation of endonuclease G in atrophied muscle of physically active older humans. FASEB J 28:1621–1633 Gutteridge JMC, Halliwell B (2010) Antioxidants: molecules, medicines, and myths. Biochem Biophys Res Commun 393:561–564 Halliwell B, Gutteridge JMG (2007) Free Radicals in Biology & Medicine, 4th edn. Oxford University Press, Oxford Halliwell B, Whiteman M (2004) Measuring reactive species and oxidative damage in vivo and in cell culture: how should you do it and what do the results mean? Br J Pharmacol 142:231–255 Harman D (1956) Aging: a theory based on free radical and radiation chemistry. J Gerontol 11:298–300 Heath GW, Hagberg JM, Ehsani AA, Holloszy JO (1981) A physiological comparison of young and older endurance athletes. J Appl Physiol 51:634–640

Hiona A, Leeuwenburgh C (2008) The role of mitochondrial DNA mutations in aging and sarcopenia: implications for the mitochondrial vicious cycle theory of aging. Exp Gerontol 43:23–33 Holloszy JO, Coyle EF (1984) Adaptations of skeletal muscle to endurance exercise and their metabolic consequences. J Appl Physiol 56:831–838 Holmstrom KM, Finkel T (2014) Cellular mechanisms and physiological consequences of redox-dependent signalling. Nat Rev Mol Cell Biol 15:411–421 Hood DA (2001) Contractile activity-induced mitochondrial biogenesis in skeletal muscle. J Appl Physiol 90: 1137–1157 Hood DA (2009) Mechanisms of exercise-induced mitochondrial biogenesis in skeletal muscle. Appl Physiol Nutr Metab 34:465–472 Irrcher R, Ljubicic V, Hood DA (2009) Interactions between ROS and AMP kinase activity in the regulation of PGC-1a transcription in skeletal muscle cells. Am J Physiol Cell Physiol 296:C116–C123 Iversen N, Krsutrup P, Rasmussen HN, Rasmussen UF, Saltin B, Pilegaard H (2011) Mitochondrial biogenesis and angiogenesis in muscle of the elderly. Exp Gerontol 46:670–678 Jackson MJ (2009) Redox regulation of adaptive responses of skeletal muscle to contractile activity. Free Radic Biol Med 47:1267–1275 Jackson MJ, McArdle A (2011) Age-related changes in skeletal muscle reactive oxygen species generation and adaptive responses to reactive oxygen species. J Physiol 589:2139–2145 Jager S, Handschin C, St-Pierre J, Spiegelman BM (2007) AMPactivated protein kinase (AMPK) action in skeletal muscle via direct phosphorylation of PGC-1a. Proc Natl Acad Sci USA 104:12017–12022 Jang YC, Van Remmen H (2009) The mitochondrial theory of aging: insight from transgenic and knockout mouse models. Exp Gerontol 44:256–260 Jang YC, Van Remmen H (2011) Age-associated alterations of the neuromuscular junction. Exp Gerontol 46:193–198 Janssen I, Shepard P, Katzmarzyk PT, Roubenoff R (2004) The healthcare costs of sarcopenia in the United States. J Am Geriatr Soc 52:80–85 Janssen-Heininger YM, Mossman BT, Heintz NH, Forman HJ, Kalyanaraman B, Finkel T et al (2008) Redox-based regulation of signal transduction: principles: pitfalls, and promises. Free Radic Biol Med 45:1–17 Ji LL, Gomez-Cabrera MC, Steinhafel N, Vina J (2004) Acute exercise activates nuclear factor (NF)-KB signalling pathway in rat skeletal muscle. FASEB J 18:1499–1506 Ji LL, Gomez-Cabrera MC, Vina J (2006) Exercise and hormesis: activation of cellular antioxidant signalling pathway. Ann NY Acad Sci 1067:425–435 Joesph AM, Adhihttey PJ, Buford TW, Wohlgemuth SE, Lees HA, Nguen LM et al (2012) The impact of aging on mitochondrial function and biogenesis pathways in skeletal muscle of sedentary high- and low functioning elderly individuals. Aging Cell 11:801–809 Kalyanaraman B, Usmar V, Davies KJA, Dennery PA, Forman RJ, Grisham MB et al (2012) Measuring reactive oxygen and nitrogen species with fluorescent probes: challenges and limitations. Free Radic Biol Med 52:1–6

123

Biogerontology Kayani AC, Morton JP, McArdle A (2008) The exerciseinduced stress response in skeletal muscle: failure during aging. Appl Physiol Nutr Metab 33:1033–1041 Khassaf M, Child RB, McArdle A, Brodie DA, Esanu C, Jackson MJ (2001) Time course of adaptive responses of human skeletal muscle to oxidative stress induced by nondamaging exercise. J Appl Physiol 90:1031–1036 Kholodenko B, Hancock JF, Kolch W (2010) Cell signalling ballet in space and time. Nat Rev Mol Cell Biol 11: 414–426 Koltai E, Hart N, Taylor AW, Goto S, Nyo JK, Davies KJ et al (2012) Age-associated declines in mitochondrial biogenesis are minimised by exercise training. Am J Physiol Regul Integr Comp Physiol 303:R127–R134 Konopka AR, Nair SK (2013) Mitochondrial and skeletal muscle health with advancing age. Mol Cell Endocrinol 379:19–29 Konopka AR, Suer MK, Wolff CA, Harber MP (2014) Markers of human skeletal muscle mitochondrial biogenesis and quality control. Effects of age and aerobic exercise training. J Gerontol A Biol Sci Med Sci 69:371–378 Lanza IR, Nair KS (2010) Regulation of skeletal muscle mitochondrial function: genes to proteins. Acta Physiol 199: 529–547 Lanza IR, Short DK, Short KR, Raghavakaimal S, Basu R, Joyner MJ et al (2008) Endurance exercise as a countermeasure for aging. Diabetes 57:2933–2942 Leick L, Wojtaszewski JFP, Johansen ST, Kiilerich K, Gomes G, Hellsten Y et al (2008) PGC-1a is not mandatory for exercise- and training-induced adaptive gene responses in mouse skeletal muscle. Am J Physiol Endocrinol Metab 294:E463–E474 Lexell J, Taylor CC, Sjostrum M (1988) What is the cause of ageing atrophy? Total number, size and proportion of different fibre types studied in whole vastus lateralis muscle from 15 to 83 year old men. J Neurol Sci 84:275–294 Little JP, Safdar A, Cermak N, Tarnopolsky MA, Gibala MJ (2010a) Acute endurance exercise increases the nuclear abundance of PGC-1a in trained human skeletal muscle. Am J Physiol Regul Integr Comp Physiol 298:R912–R917 Little JP, Safdar A, Wilkin GP, Tranopolsky MA, Gibala MJ (2010b) A practical model of low-volume high-intensity interval training induces mitochondrial biogenesis in human skeletal muscle: potential mechanisms. J Physiol 588:1011–1022 Ljubicic V, Hood DA (2009a) Diminished contraction-induced intracellular signalling towards mitochondrial biogenesis in aged skeletal muscle. Aging Cell 8:394–404 Ljubicic V, Hood DA (2009b) Specific attenuation of protein kinase phosphorylation in muscle with a high mitochondrial content. Am J Physiol Endocrinol Metab 297:E749– E758 Ljubicic V, Joesph A, Adhihetty PJ, Huang JH, Saleem A, Uguccioni G et al (2009) Molecular basis for an attenuated mitochondrial plasticity in aged skeletal muscle. Aging 9:818–830 Ljubicic V, Joseph AM, Saleem A, Uguccioni G, Collu-Marchese M, Lai RY et al (2010) Transcriptional and posttranscriptional regulation of mitochondrial biogenesis in skeletal muscle: effects of exercise and aging. Biochim Biophys Acta 1800:223–234

123

Lopez-Lluch G, Irusta PM, Navas P, de Cabo R (2008) Mitochondrial biogenesis and healthy aging. Exp Gerontol 43:813–819 Mahoney DJ, Praise G, Melov S, Safdar A, Tarnopolsky MA (2005) Analysis of global mRNA expression from human skeletal muscle during recovery from endurance exercise. FASEB J 19:1498–1500 Margaritelis NV, Kyparos A, Paschalis V, Theodorou AA, Panayiotou G, Zafeiridis A et al (2014) Reductive stress after exercise: the issue of redox individuality. Redox Biol 2:520–528 Martinez-Ruiz A, Cadenas S, Lamas S (2011) Nitric oxide signalling: classical, less classical and nonclassical mechanisms. Free Radic Biol Med 51:17–29 McArdle A, Patwell D, Vasilaki A, Griffiths RD, Jackson MJ (2001) Contractile-activity induced oxidative stress: cellular origin and adaptive responses. Am J Physiol Cell Physiol 280:C621–C627 McDonagh B, Sakellariou GK, Smith N, Brownridge P, Jackson MJ (2014) Differential cysteine labelling and global labelfree proteomics reveals an altered metabolic state in skeletal muscle aging. J Prot Res. doi:10.1021/pr5006394 McGlory C, White A, Treins C, Drust B, Close GL, MacLaren DPM et al (2014) Application of the [gamma-P-32] ATP kinase assay to study anabolic signaling in human skeletal muscle. J Appl Physiol 116:504–513 McKenzie D, Bua E, McKiernan S, Cao Z, Wanagat J, Aiken JM (2002) Mitochondrial DNA deletion mutations: a causal role in sarcopenia. Euro J Biochem 269:2010–2015 Melov A, Tarnopolsky MA, Beckman K, Felkey K, Hubbard A (2007) Resistance exercise reverses aging in human skeletal muscle. PLoS ONE 2:e465 Menshikova EV, Ritov VB, Fairfull L, Ferrel RE, Kelley DE, Goodpaster BH (2006) Effects of exercise on mitochondrial content and function in aging human muscle. J Gerontol 61:534–540 Miller BF, Hamilton KL (2012) A perspective on the determination of mitochondrial biogenesis. Am J Physiol Endocrinol Metab 302:E496–E499 Morton JP, MacLaren DPM, Cable NT, Bongers T, Griffiths RD, Campbell IT et al (2006) Time-course and differential responses of the major heat shock protein families in human skeletal muscle following acute non-damaging treadmill exercise. J Appl Physiol 101:176–182 Morton JP, Kayani AC, McArdle A, Drust B (2009) The exercise-induced stress response of skeletal muscle with specific emphasis on humans. Sports Med 39:643–662 Murphy MP, Holmgren A, Larsson N, Halliwell B, Chang CJ, Kalyanaraman B et al (2011) Unravelling the biological roles of reactive oxygen species. Cell Metab 13:361–366 Nordsborg NB, Lundby C, Leick L, Pilegaard H (2010) Relative workload determine exercise-induced increases in PGC1alpaha mRNA. Med Sci Sports Exerc 42:1477–1484 Nyberg M, Blackwell JR, Damsgaard R, Jones AM, Hellsten Y, Mortensen SP (2012) Lifelong physical activity prevents an age-related reduction in arterial and skeletal muscle nitric oxide bioavailability in humans. J Physiol 590:5361–5370 Nyberg M, Mortensen SP, Cabo H, Gomez-Cabrera MC, Vina J, Hellensten Y (2014) Role of sedentary aging and lifelong physical activity on exchange of glutathione across

Biogerontology exercising human skeletal muscle. Free Radic Biol Med 73:166–173 Pahor M, Blair SN, Espeland M, Fielding R, Gill TM, Guralink JM et al (2006) Effects of a physical activity intervention on measures of physical activity intervention on measures of physical performance: results of the lifestyle interventions and independence for Elders Pilot (LIFE-P) study. J Gerontol A Biol Sci Med Sci 61:1157–1165 Palomero J, Pye D, Kabayo T, Spiller DG, Jackson MJ (2008) In situ detection and measurement of intracellular reactive oxygen species in single isolated mature skeletal muscle fibres by real-time fluorescence microscopy. Antioxid Redox Signal 10:1463–1474 Palomero J, Vasilaki A, Pye D, McArdle A, Jackson MJ (2013) Aging increases the oxidation of dichlorohydrofluorescein in single isolated skeletal muscle fibres at rest, but not during contractions. Am J Physiol Regul Integr Comp Physiol 305:R351–R358 Pearson SJ, Young A, Macaluso A, Devito G, Nimmo MA, Cobbold M et al (2002) Muscle function in elite master weightlifters. Med Sci Sports Exerc 34:1199–1206 Pearson T, McArdle A, Jackson MJ (2014) Nitric oxide availability is increased in contracting skeletal muscle from aged mice, but does not differentially decrease muscle superoxide. Free Radic Biol Med. doi:10.1016/j.freeradbiomed. 2014.10.505i Perry CGR, Lally J, Holloway GP, Heigenhauser GJF, Bonen A, Spriet LL (2010) Repeated transient mRNA bursts precede increases in transcriptional and mitochondrial proteins during training in human skeletal muscle. J Physiol 588:4795–4810 Peterson CM, Johannsen DL, Ravussin E (2012) Skeletal muscle mitochondria and aging: a review. J Aging Res 2012:194821 Philp A, Chen A, Lan D, Meyer GA, Murphy A, Knapp AE et al (2011) Sirtuin 1 (SIRT1) deactylase activity is not required for mitochondrial biogenesis or peroxisome proliferator activate receptor-c co-activator-1a (PGC-1a) deactylation following exercise. J Biol Chem 286:30561–30570 Picard M, Ritchie D, Wright KJ, Romestaing C, Thomas MM, Rowan SL et al (2010) Mitochondrial functional impairment with aging is exaggerated in isolated mitochondria compared to permealised fibres. Aging Cell 6:1032–1046 Picard M, Taivassalo T, Gouspillou G, Hepple RT (2011a) Mitochondria: isolation, structure and function. J Physiol 589:4421–4431 Picard M, Ritchie D, Thomas MM, Wright KJ, Hepple RT (2011b) Alterations in intrinsic mitochondrial function with aging are fibre type-specific and do not explain differential atrophy between muscles. Aging Cell 10:1047–1055 Picard M, Shirihai OS, Gentil BJ, Burelle Y (2013) Mitochondrial morphology transitions and functions: implications for retrograde signalling. Am J Physiol Regul Integr Comp Physiol 304:R393–R406 Pilegaard H, Saltin B, Neufer DP (2003) Exercise induces transient transcriptional activation of the PGC-1a gene in human skeletal muscle. J Physiol 546:851–858 Powers SK, Jackson MJ (2008) Exercise-induced oxidative stress: cellular mechanisms and impact on muscle force production. Physiol Rev 88:1243–1276

Pye D, Palomero J, Kabayo T, Jackson MJ (2007) Real-time measurement of nitric oxide in single mature mouse skeletal muscle fibres during contractions. J Physiol 581:309–318 Radak Z, Bori Z, Kolati E, Fatourous GI, Jamaturas AZ, Douroudos II et al (2011) Age-dependent changes in 8-oxoguanine-DNA glyocloase activity are modulated by the adaptive response to physical exercise in human muscle. Free Radic Biol Med 51:417–423 Rennie MJ, Selby A, Atherton P, Smith K, Glover EL, Phillips SM (2010) Facts noise and wishful thinking: muscle protein turnover in aging and human disuse atrophy. Scand J Med Sci Sport 20:5–9 Reznick RM, Zong H, Li J, Morino K, Moore IK, Yu HJ et al (2007) Aging-associated reductions in AMP-activated protein kinase activity and mitochondrial biogenesis. Cell Metab 5:151–156 Rizzuto R, De Stefani D, Raffaello A, Mammucari C (2012) Mitochondria as sensors and regulators of calcium signalling. Nat Rev Mol Cell Biol 11:872–884 Rodgers JT, Lerin C, Haas W, Gygy SP, Spiegelman BM, Puigserver P (2005) Nutrient control of glucose homeostasis through a complex of PGC-1a and SIRT1. Nature 434:113–118 Rowe GC, El-Khoury R, Patten IS, Rustin P, Arany Z (2012) PGC-1a is dispensable for exercise-induced mitochondrial biogenesis in skeletal muscle. PLoS ONE 7:e41817 Russell AP, Feilchenfeldt J, Schreiber S, Praz M, Crettenand A, Gobelet C et al (2003) Endurance training in humans leads to fiber type-specific increases in levels of peroxisome proliferator-activated receptor-c coactivator-1a in skeletal muscle. Diabetes 52:2874–28812 Safdar A, DeBeer J, Tarnopolsky MA (2010a) Dysfunctional Nrf2-Keap1 redox signalling in skeletal muscle of the sedentary old. Free Radic Biol Med 49:1487–1493 Safdar A, Hamadeh MJ, Kaczor JJ, Raha S, deBeer J, Tranopolsky MA (2010b) Aberrant mitochondrial homeostasis in the skeletal muscle of sedentary adults. PLoS ONE 5:e10788 Safdar A, Little JP, Stokl AJ, Hettinga BP, Akhtar M, Tarnopolsky MA (2011) Exercise increases mitochondrial PGC1a content and promotes nuclear-mitochondrial cross-talk to coordinate mitochondrial biogenesis. J Biol Chem 286:10605–10617 Sakellariou GK, Vasilaki A, Palomero J, Kayani A, Zibrik L, McArdle A et al (2013) Studies of mitochondrial and nonmitochondrial sources implicate nicotinamide adenine dinucleotide phosphate oxidase (s) in the increase skeletal muscle superoxide generation that occurs following contractile activity. Antiox Redox Signal 18:603–621 Sakellariou GK, Jackson MJ, Vasilaki A (2014) Redefining the major contributors to superoxide production in contracting skeletal muscle. Role of NAD(P)H oxidases. Free Radic Res 48:12–29 Saleem A, Hood DA (2013) Acute exercise induces tumour suppressor protein p53 translocation to the mitochondria and promotes a p53-Tfam-mitochondrial DNA complex in skeletal muscle. J Physiol 591:3625–3636 Salmon AB, Richardson A, Perez VI (2010) Update on the oxidative stress theory of aging: does oxidative stress play a role in aging or healthy aging? Free Radic Biol Med 48:642–655

123

Biogerontology Sandri M, Lin J, Handschin C, Yang W, Arany ZP, Lecker SH et al (2006) PGC-1alpha protects form atrophy by suppressing FOXO3 action and atrophy-specific gene transcription. Proc Natl Acad Sci USA 103:16260–16265 Scarpulla RC (2008) Transcriptional paradigms in mammalian mitochondrial biogenesis and function. Physiol Rev 88: 611–638 Short KR, Vittone JL, Brigelow ML, Proctor DN, Rizza RA, Coenen-Schimke JM et al (2003) Impact of aerobic exercise training on age-related changes in insulin sensitivity and muscle oxidative capacity. Diabetes 52:1888–1896 Short KR, Bigelow ML, Kahl J, Singh R, Coenen-Schimke J, Raghavakaimal S, Nair KS (2005) Decline in skeletal muscle mitochondrial function with aging in humans. Proc Natl Acad Sci USA 102:5618–5623 Sies H (2014) Role of metabolic H2O2 generation: redox signalling and oxidative stress. J Biol Chem. doi:10.1074/jbc. R113.544635 Stuart JA, Maddalena LA, Merilovich M, Robb EL (2014) A midlife crisis for the mitochondrial free radical theory of aging. Longev Healthspan. 3:4 Tanaka H, Seals DR (2008) Endurance exercise and performance in masters athletes: age-associated changes and underlying physiological mechanisms. J Physiol 586:55–63 Tanaka H, DeSouza CA, Jones PP, Stevenson ET, Davy KP, Seals DR (1997) Greater decline in maximal aerobic capacity with age in physically active vs sedentary healthy women. J Appl Physiol 83:1947–1953 Timmons JA (2011) Variability in training-induced skeletal muscle adaptation. J Appl Physiol 110:846–853 Trappe SW, Costill DL, Vukovich MD, Jones J, Melham T (1996) Aging among elite distance runners: a 22 year longitudinal study. J Appl Physiol 80:285–290 Trappe S, Hayes E, Galpin A, Kaminsky L, Jemiolo B, Fink W et al (2013) New records in aerobic power among octogenarian lifelong endurance athletes. J Appl Physiol 114:3–10 Valls MRB, Wilkinson DJ, Narici M, Smith K, Phillips BE et al (2014) Protein carbonylation and heat shock proteins in

123

human skeletal muscle: relationships to age and sarcopenia. J Gerontol A Biol Sci Med Sci. doi:10.1093/gerona/ glu007 Vasilaki A, Jackson MJ, McArdle A (2002) Attenuated HSP70 response in skeletal muscle of aged rats following contractile activity. Muscle Nerve 25:902–905 Vasilaki A, Iwanejko I, McArdle F, Broome CS, Jackson MJ, McArdle A (2003) Skeletal muscle of aged male mice fail to adapt following contractile activity. Biochem Soc Trans 31:455–456 Vasilaki A, McArdle F, Iwanejko LM, McArdle A (2006a) Adaptive response of mouse skeletal muscle to contractile activity: the effect of age. Mech Ageing Dev 127:830–839 Vasilaki A, Mansouri A, Van Remmen H, Van Der Meulen JH, Larkin L, Richardson AG et al (2006b) Free radical generation by skeletal muscle of adult and old mice: effect of contractile activity. Aging Cell 5:109–117 Wenz T (2013) Regulation of mitochondrial biogenesis and PGC-1a under cellular stress. Mitochondrion 13:134–142 Wenz T, Rosse SG, Rotundo RL, Spiegelman BM, Moraes CT (2009) Increased muscle PGC-1a expression protects from sarcopenia and metabolic disease. Proc Natl Acad Sci USA 106:20405–20410 Westermann B (2010) Mitochondrial fusion and fission in cell life and death. Nat Rev Mol Cell Biol 11:872–884 Winterbourn CC (2008) Reconciling the chemistry and biology of reactive oxygen species. Nat Chem Biol 4:278–286 Winterbourn CC (2014a) Are free radicals involved in thiolbased redox signalling? Free Radic Biol Med. doi:10.1016/ j.freeradbiolmed.2014.08.017i Winterbourn CC (2014b) The challenges of using fluorescent probes to detect and quantify specific reactive oxygen species in living cells. Biochim Biophys Acta 1840:730– 738 Wright DC, Han DH, Garcia-Roves PM, Geiger PC, Jones TE, Holloszy JO (2007) Exercise-induced mitochondrial biogenesis begins before the increase in muscle PGC-1a expression. J Biol Chem 282:194–199