Lifelong dietary protein restriction accelerates skeletal muscle loss and reduces muscle fibre size by impairing proteostasis and mitochondrial homeostasis.

The early life environment significantly affects the development of age-related skeletal muscle disorders. However, the long-term effects of lactational protein restriction on skeletal muscle are still poorly defined. Our study revealed that male mice nursed by dams fed a low-protein diet during lactation exhibited skeletal muscle growth restriction. This was associated with a dysregulation in the expression levels of genes related to the ribosome, mitochondria and skeletal muscle development. We reported that lifelong protein restriction accelerated loss of type-IIa muscle fibres and reduced muscle fibre size by impairing mitochondrial homeostasis and proteostasis at 18 months of age. However, feeding a normal-protein diet following lactational protein restriction prevented accelerated fibre loss and fibre size reduction in later life. These findings provide novel insight into the mechanisms by which lactational protein restriction hinders skeletal muscle growth and includes evidence that lifelong dietary protein restriction accelerated skeletal muscle loss in later life.

Randomised controlled trial combining vitamin E-functionalised chocolate with physical exercise to reduce the risk of protein-energy malnutrition in predementia aged people: study protocol for Choko-Age.

OBJECTIVE: Protein-energy malnutrition and the subsequent muscle wasting (sarcopenia) are common ageing complications. It is knowing to be also associated with dementia. Our programme will test the cytoprotective functions of vitamin E combined with the cortisol-lowering effect of chocolate polyphenols (PP), in combination with muscle anabolic effect of adequate dietary protein intake and physical exercise to prevent the age-dependent decline of muscle mass and its key underpinning mechanisms including mitochondrial function, and nutrient metabolism in muscle in the elderly. METHODS AND ANALYSIS: In 2020, a 6-month double-blind randomised controlled trial in 75 predementia older people was launched to prevent muscle mass loss, in respond to the 'Joint Programming Initiative A healthy diet for a healthy life'. In the run-in phase, participants will be stabilised on a protein-rich diet (0.9-1.0 g protein/kg ideal body weight/day) and physical exercise programme (high-intensity interval training specifically developed for these subjects). Subsequently, they will be randomised into three groups (1:1:1). The study arms will have a similar isocaloric diet and follow a similar physical exercise programme. Control group (n=25) will maintain the baseline diet; intervention groups will consume either 30 g/day of dark chocolate containing 500 mg total PP (corresponding to 60 mg epicatechin) and 100 mg vitamin E (as RRR-alpha-tocopherol) (n=25); or the high polyphenol chocolate without additional vitamin E (n=25). Muscle mass will be the primary endpoint. Other outcomes are neurocognitive status and previously identified biomolecular indices of frailty in predementia patients. Muscle biopsies will be collected to assess myocyte contraction and mitochondrial metabolism. Blood and plasma samples will be analysed for laboratory endpoints including nutrition metabolism and omics. ETHICS AND DISSEMINATION: All the ethical and regulatory approvals have been obtained by the ethical committees of the Azienda Ospedaliera Universitaria Integrata of Verona with respect to scientific content and compliance with applicable research and human subjects' regulation. Given the broader interest of the society toward undernutrition in the elderly, we identify four main target audiences for our research activity: national and local health systems, both internal and external to the project; targeted population (the elderly); general public; and academia. These activities include scientific workshops, public health awareness campaigns, project dedicated website and publication is scientific peer-review journals. TRIAL REGISTRATION NUMBER: NCT05343611.

Older mice show decreased regeneration of neuromuscular junctions following lengthening contraction-induced injury.

Progressive muscle atrophy and loss of muscle strength associated with old age have been well documented. Although age-associated impairments in skeletal muscle regeneration following injury have been demonstrated, less is known about whether aging impacts the regenerative response of neuromuscular junctions (NMJ) following contraction-induced injury. Reduced ability of NMJs to regenerate could lead to increased numbers of denervated muscle fibers and therefore play a contributing role to age-related sarcopenia. To investigate the relationship between age and NMJ regeneration following injury, extensor digitorum longus (EDL) muscles of middle-aged (18-19 months) and old mice (27-28 months) were subjected to a protocol of lengthening contractions (LC) that resulted in an acute force deficit of ~55% as well as functional and histological evidence of a similar magnitude of injury 3 days post LCs that was not different between age groups. After 28 days, the architecture and innervation of the NMJs were evaluated. The numbers of fragmented endplates increased and of fully innervated NMJs decreased post-injury for the muscle of both middle-aged and old mice and for contralateral uninjured muscles of old compared with uninjured muscles of middle-aged controls. Thus, the diminished ability of the skeletal muscle of old mice to recover following injury may be due in part to an age-related decrease in the ability to regenerate NMJs in injured muscles. The impaired ability to regenerate NMJs may be a triggering factor for degenerative changes at the NMJ contributing to muscle fiber weakness and loss in old age.

Redox Control of Signalling Responses to Contractile Activity and Ageing in Skeletal Muscle.

Research over almost 40 years has established that reactive oxygen species are generated at different sites in skeletal muscle and that the generation of these species is increased by various forms of exercise. Initially, this was thought to be potentially deleterious to skeletal muscle and other tissues, but more recent data have identified key roles of these species in muscle adaptations to exercise. The aim of this review is to summarise our current understanding of these redox signalling roles of reactive oxygen species in mediating responses of muscle to contractile activity, with a particular focus on the effects of ageing on these processes. In addition, we provide evidence that disruption of the redox status of muscle mitochondria resulting from age-associated denervation of muscle fibres may be an important factor leading to an attenuation of some muscle responses to contractile activity, and we speculate on potential mechanisms involved.

Exercise stress leads to an acute loss of mitochondrial proteins and disruption of redox control in skeletal muscle of older subjects: An underlying decrease in resilience with aging?

Reactive oxygen species (ROS) are recognized as important signaling molecules in healthy skeletal muscle. Redox sensitive proteins can respond to intracellular changes in ROS by oxidation of reactive thiol groups on cysteine (Cys) residues. Exercise is known to induce the generation of superoxide and nitric oxide, resulting in the activation of several adaptive signaling pathways; however, it has been suggested that aging attenuates these redox-regulated adaptations to acute exercise. In the present study, we used redox proteomics to study the vastus lateralis muscles of Adult (n = 6 male, 6 female; 18-30 yrs) and Old (n = 6 male, 6 female; 64-79 yrs) adults. Participants completed a bout of high intensity cycling exercise consisting of five sets of 2-min intervals performed at 80% maximal aerobic power output (PPO), with 2 min recovery cycling at 40% PPO between sets. Muscle biopsies were collected prior to exercise, and immediately following the first, second, and fifth high intensity interval. Global proteomic analysis indicated differences in abundance of a number of individual proteins between skeletal muscles of Adult and Old subjects at rest with a significant exacerbation of these differences induced by the acute exercise. In particular, we observed an exercise-induced decrease in abundance of mitochondrial proteins in muscles from older subjects only. Redox proteome analysis revealed cysteines from five cytosolic proteins in older subjects with lower oxidation (i.e. greater reduction) than was seen in muscle from the young adults at rest. Redox homeostasis was well maintained in Adult subjects following exercise, but there was significant increase in oxidation of multiple mitochondrial and cytosolic protein cysteines in Old subjects. We also observed that oxidation of peroxiredoxin 3 occurred following exercise in both Adult and Old groups, supporting the possibility that this is a key effector protein for mitochondrial redox signaling. Thus, we show, for the first time that exercise reveals a lack of resilience in muscle of older human participants, that is apparent as a loss of mitochondrial proteins and oxidation of multiple protein cysteines that are not seen in younger subjects. The precise consequences of this redox disruption are unclear, but this likely play a role in the attenuation of multiple adaptations to exercise that are classically seen with aging. Such changes were only seen following the acute stress of exercise., highlighting the need to consider not only basal differences seen during aging but also the difference following physiological challenge.

On the mechanisms underlying attenuated redox responses to exercise in older individuals: A hypothesis.

Responding appropriately to exercise is essential to maintenance of skeletal muscle mass and function at all ages and particularly during aging. Here, a hypothesis is presented that a key component of the inability of skeletal muscle to respond effectively to exercise in aging is a denervation-induced failure of muscle redox signalling. This novel hypothesis proposes that an initial increase in oxidation in muscle mitochondria leads to a paradoxical increase in the reductive state of specific cysteines of signalling proteins in the muscle cytosol that suppresses their ability to respond to normal oxidising redox signals during exercise. The following are presented for consideration:Transient loss of integrity of peripheral motor neurons occurs repeatedly throughout life and is normally rapidly repaired by reinnervation, but this repair process becomes less efficient with aging. Each transient loss of neuromuscular integrity leads to a rapid, large increase in mitochondrial peroxide production in the denervated muscle fibers and in neighbouring muscle fibers. This peroxide may initially act to stimulate axonal sprouting and regeneration, but also stimulates retrograde mitonuclear communication to increase expression of a range of cytoprotective proteins in an attempt to protect the fiber and neighbouring tissues against oxidative damage. The increased peroxide within mitochondria does not lead to an increased cytosolic peroxide, but the increases in adaptive cytoprotective proteins include some located to the muscle cytosol which modify the local cytosol redox environment to induce a more reductive state in key cysteines of specific signalling proteins. Key adaptations of skeletal muscle to exercise involve transient peroxiredoxin oxidation as effectors of redox signalling in the cytosol. This requires sensitive oxidation of key cysteine residues. In aging, the chronic change to a more reductive cytosolic environment prevents the transient oxidation of peroxiredoxin 2 and hence prevents essential adaptations to exercise, thus contributing to loss of muscle mass and function. Experimental approaches suitable for testing the hypothesis are also outlined.

Neuron-specific deletion of CuZnSOD leads to an advanced sarcopenic phenotype in older mice.

Age-associated loss of muscle mass and function (sarcopenia) has a profound effect on the quality of life in the elderly. Our previous studies show that CuZnSOD deletion in mice (Sod1-/- mice) recapitulates sarcopenia phenotypes, including elevated oxidative stress and accelerated muscle atrophy, weakness, and disruption of neuromuscular junctions (NMJs). To determine whether deletion of Sod1 initiated in neurons in adult mice is sufficient to induce muscle atrophy, we treated young (2- to 4-month-old) Sod1flox/SlickHCre mice with tamoxifen to generate i-mn-Sod1KO mice. CuZnSOD protein was 40-50% lower in neuronal tissue in i-mn-Sod1KO mice. Motor neuron number in ventral spinal cord was reduced 28% at 10 months and more than 50% in 18- to 22-month-old i-mn-Sod1KO mice. By 24 months, 22% of NMJs in i-mn-Sod1KO mice displayed a complete lack of innervation and deficits in specific force that are partially reversed by direct muscle stimulation, supporting the loss of NMJ structure and function. Muscle mass was significantly reduced by 16 months of age and further decreased at 24 months of age. Overall, our findings show that neuronal-specific deletion of CuZnSOD is sufficient to cause motor neuron loss in young mice, but that NMJ disruption, muscle atrophy, and weakness are not evident until past middle age. These results suggest that loss of innervation is critical but may not be sufficient until the muscle reaches a threshold beyond which it cannot compensate for neuronal loss or rescue additional fibers past the maximum size of the motor unit.

2-Cys peroxiredoxin oxidation in response to hydrogen peroxide and contractile activity in skeletal muscle: A novel insight into exercise-induced redox signalling?

Skeletal muscle generates superoxide during contractions which is rapidly converted to H2O2. This molecule has been proposed to activate signalling pathways and transcription factors that regulate key adaptive responses to exercise but the concentration of H2O2 required to oxidise and activate key signalling proteins in vitro is much higher than the intracellular concentration in muscle fibers following exercise. We hypothesised that Peroxiredoxins (Prx), which reacts with H2O2 at the low intracellular concentrations found in muscle, would be rapidly oxidised in contracting muscle and hence potentially transmit oxidising equivalents to downstream signalling proteins as a method for their oxidation and activation. The aim of this study was to characterise the effects of muscle contractile activity on the oxidation of Prx1, 2 and 3 and determine if these were affected by aging. Prx1, 2 and 3 were all rapidly and reversibly oxidised following treatment with low micromolar concentrations of H2O2 in C2C12 myotubes and also in isolated mature flexor digitalis brevis fibers from adult mice following a protocol of repeated isometric contractions. Significant oxidation of Prx2 was seen within 1 min (i.e. after 12 contractions), whereas significant oxidation was seen after 2 min for Prx1 and 3. In muscle fibers from old mice, Prx2 oxidation was significantly attenuated following contractile activity. Thus we show for the first time that Prx are rapidly and reversibly oxidised in response to contractile activity in skeletal muscle and hypothesise that these proteins act as effectors of muscle redox signalling pathways which are key to adaptations to exercise that are attenuated during aging.

Hydrogen peroxide as a signal for skeletal muscle adaptations to exercise: What do concentrations tell us about potential mechanisms?

Hydrogen peroxide appears to be the key reactive oxygen species involved in redox signalling, but comparisons of the low concentrations of hydrogen peroxide that are calculated to exist within cells with those previously shown to activate common signalling events in vitro indicate that direct oxidation of key thiol groups on "redox-sensitive" signalling proteins is unlikely to occur. A number of potential mechanisms have been proposed to explain how cells overcome this block to hydrogen peroxide-stimulated redox signalling and these will be discussed in the context of the redox-stimulation of specific adaptations of skeletal muscle to contractile activity and exercise. It is argued that current data implicate a role for currently unidentified effector molecules (likely to be highly reactive peroxidases) in propagation of the redox signal from sites of hydrogen peroxide generation to common adaptive signalling pathways.

Mechanistic models to guide redox investigations and interventions in musculoskeletal ageing.

Age is the greatest risk factor for the major chronic musculoskeletal disorders, osteoarthritis, osteoporosis and age-related loss of skeletal muscle mass and function (sarcopenia). Dramatic advances in understanding of the fundamental mechanisms underlying the ageing process are being exploited to understand the causes of these age-related disorders and identify approaches to prevent or treat these disorders. This review will focus on one of these fundamental mechanisms, redox regulation, and the role of redox changes in age-related loss of skeletal muscle mass and function (sarcopenia). Key to understanding the role of such pathways has been the development and study of experimental models of musculoskeletal ageing that are designed to examine the effect of modification of ROS regulatory enzymes. These have primarily involved genetic deletion of regulatory enzymes for ROS in mice. Many of the models studied show increased oxidative damage in tissues, but no clear relationship with skeletal muscle aging has been seen The exception to this has been mice with disruption of the superoxide dismutases and, in particular, deletion of Cu,ZnSOD (SOD1) localised in the cytosol and mitochondrial intermembrane space. Studies of tissue specific models lacking SOD1 have highlighted the potential role that disrupted redox pathways can play in muscle loss and weakness and have demonstrated the need to study both motor neurons and muscle to understand age-related loss of skeletal muscle. The complex interplay that has been identified between changes in redox homeostasis in the motor neuron and skeletal muscle and their role in premature loss of muscle mass and function illustrates the utility of modifiable models to establish key pathways that may contribute to age-related changes and identify potential logical approaches to intervention.

Redox responses in skeletal muscle following denervation.

Previous studies have shown a significant increase in the mitochondrial generation of hydrogen peroxide (H2O2) and other peroxides in recently denervated muscle fibers. The mechanisms for generation of these peroxides and how the muscle responds to these peroxides are not fully established. The aim of this work was to determine the effect of denervation on the muscle content of proteins that may contribute to mitochondrial peroxide release and the muscle responses to this generation. Denervation of the tibialis anterior (TA) and extensor digitorum longus (EDL) muscles in mice was achieved by surgical removal of a small section of the peroneal nerve prior to its entry into the muscle. An increase in mitochondrial peroxide generation has been observed from 7 days and sustained up to 21 days following denervation in the TA muscle fibers. This increased peroxide generation was reduced by incubation of skinned fibers with inhibitors of monoamine oxidases, NADPH oxidases or phospholipase A2 enzymes and the muscle content of these enzymes together with peroxiredoxin 6 were increased following denervation. Denervated muscle also showed significant adaptations in the content of several enzymes involved in the protection of cells against oxidative damage. Morphological analyses indicated a progressive significant loss of muscle mass in the TA muscle from 7 days up to 21 days following denervation due to fiber atrophy but without fiber loss. These results support the possibility that, at least initially, the increase in peroxide production may stimulate adaptations in an attempt to protect the muscle fibers, but that these processes are insufficient and the increased peroxide generation over the longer term may activate degenerative and atrophic processes in the denervated muscle fibers.

Accelerated sarcopenia in Cu/Zn superoxide dismutase knockout mice.

Mice lacking Cu/Zn-superoxide dismutase (Sod1-/- or Sod1KO mice) show high levels of oxidative stress/damage and a 30% decrease in lifespan. The Sod1KO mice also show many phenotypes of accelerated aging with the loss of muscle mass and function being one of the most prominent aging phenotypes. Using various genetic models targeting the expression of Cu/Zn-superoxide dismutase to specific tissues, we evaluated the role of motor neurons and skeletal muscle in the accelerated loss of muscle mass and function in Sod1KO mice. Our data are consistent with the sarcopenia in Sod1KO mice arising through a two-hit mechanism involving both motor neurons and skeletal muscle. Sarcopenia is initiated in motor neurons leading to a disruption of neuromuscular junctions that results in mitochondrial dysfunction and increased generation of reactive oxygen species (ROS) in skeletal muscle. The mitochondrial ROS generated in muscle feedback on the neuromuscular junctions propagating more disruption of neuromuscular junctions and more ROS production by muscle resulting in a vicious cycle that eventually leads to disaggregation of neuromuscular junctions, denervation, and loss of muscle fibers.

Advanced glycation end products-related modulation of cathepsin L and NF-κB signalling effectors in retinal pigment epithelium lead to augmented response to TNFα.

The retinal pigment epithelium (RPE) plays a central role in neuroretinal homoeostasis throughout life. Altered proteolysis and inflammatory processes involving RPE contribute to the pathophysiology of age-related macular degeneration (AMD), but the link between these remains elusive. We report for the first time the effect of advanced glycation end products (AGE)-known to accumulate on the ageing RPE's underlying Bruch's membrane in situ-on both key lysosomal cathepsins and NF-κB signalling in RPE. Cathepsin L activity and NF-κB effector levels decreased significantly following 2-week AGE exposure. Chemical cathepsin L inhibition also decreased total p65 protein levels, indicating that AGE-related change of NF-κB effectors in RPE cells may be modulated by cathepsin L. However, upon TNFα stimulation, AGE-exposed cells had significantly higher ratio of phospho-p65(Ser536)/total p65 compared to non-AGEd controls, with an even higher fold increase than in the presence of cathepsin L inhibition alone. Increased proportion of active p65 indicates an AGE-related activation of NF-κB signalling in a higher proportion of cells and/or an enhanced response to TNFα. Thus, NF-κB signalling modulation in the AGEd environment, partially regulated via cathepsin L, is employed by RPE cells as a protective (para-inflammatory) mechanism but renders them more responsive to pro-inflammatory stimuli.

Aberrant redox signalling and stress response in age-related muscle decline: Role in inter- and intra-cellular signalling.

Age-associated frailty is predominantly due to loss of muscle mass and function. The loss of muscle mass is also associated with a greater loss of muscle strength, suggesting that the remaining muscle fibres are weaker than those of adults. The mechanisms by which muscle is lost with age are unclear, but in this review we aim to pull together various strands of evidence to explain how muscle contractions support proteostasis in non-muscle tissues, particularly focussed on the production and potential transfer of Heat Shock Proteins (HSPs) and how this may fail during ageing, Furthermore we will identify logical approaches, based on this hypothesis, by which muscle loss in ageing may be reduced. Skeletal muscle generates superoxide and nitric oxide at rest and this generation is increased by contractile activity. In adults, this increased generation of reactive oxygen and nitrogen species (RONS) activate redox-sensitive transcription factors such as nuclear factor κB (NFκB), activator protein-1 (AP1) and heat shock factor 1 (HSF1), resulting in increases in cytoprotective proteins such as the superoxide dismutases, catalase and heat shock proteins that prevent oxidative damage to tissues and facilitate remodelling and proteostasis in both an intra- and inter-cellular manner. During ageing, the ability of skeletal muscle from aged organisms to respond to an increase in ROS generation by increased expression of cytoprotective proteins through activation of redox-sensitive transcription factors is severely attenuated. This age-related lack of physiological adaptations to the ROS induced by contractile activity appears to contribute to a loss of ROS homeostasis, increased oxidative damage and age-related dysfunction in skeletal muscle and potentially other tissues.

Towards a toolkit for the assessment and monitoring of musculoskeletal ageing.

The complexities and heterogeneity of the ageing process have slowed the development of consensus on appropriate biomarkers of healthy ageing. The MRC-Arthritis Research UK Centre for Integrated research into Musculoskeletal Ageing (CIMA) is a collaboration between researchers and clinicians at the Universities of Liverpool, Sheffield and Newcastle. One of CIMA's objectives is to 'Identify and share optimal techniques and approaches to monitor age-related changes in all musculoskeletal tissues, and to provide an integrated assessment of musculoskeletal function', i.e. to develop a toolkit for assessing musculoskeletal ageing. This toolkit is envisaged as an instrument that can be used to characterise and quantify musculoskeletal function during 'normal' ageing, lend itself to use in large-scale, internationally important cohorts, and provide a set of biomarker outcome measures for epidemiological and intervention studies designed to enhance healthy musculoskeletal ageing. Such potential biomarkers include: biochemical measurements in biofluids or tissue samples, in vivo measurements of body composition, imaging of structural and physical properties, and functional tests. The CIMA Toolkit Working Group assessed candidate biomarkers of musculoskeletal ageing under these four headings, detailed their biological bases, strengths and limitations, and made practical recommendations for their use. In addition, the CIMA Toolkit Working Group identified gaps in the evidence base and suggested priorities for further research on biomarkers of musculoskeletal ageing.

Developing a toolkit for the assessment and monitoring of musculoskeletal ageing.

The complexities and heterogeneity of the ageing process have slowed the development of consensus on appropriate biomarkers of healthy ageing. The Medical Research Council-Arthritis Research UK Centre for Integrated research into Musculoskeletal Ageing (CIMA) is a collaboration between researchers and clinicians at the Universities of Liverpool, Sheffield and Newcastle. One of CIMA's objectives is to 'Identify and share optimal techniques and approaches to monitor age-related changes in all musculoskeletal tissues, and to provide an integrated assessment of musculoskeletal function'-in other words to develop a toolkit for assessing musculoskeletal ageing. This toolkit is envisaged as an instrument that can be used to characterise and quantify musculoskeletal function during 'normal' ageing, lend itself to use in large-scale, internationally important cohorts, and provide a set of biomarker outcome measures for epidemiological and intervention studies designed to enhance healthy musculoskeletal ageing. Such potential biomarkers include: biochemical measurements in biofluids or tissue samples, in vivo measurements of body composition, imaging of structural and physical properties, and functional tests. This review assesses candidate biomarkers of musculoskeletal ageing under these four headings, details their biological bases, strengths and limitations, and makes practical recommendations for their use. In addition, we identify gaps in the evidence base and priorities for further research on biomarkers of musculoskeletal ageing.

Redox responses are preserved across muscle fibres with differential susceptibility to aging.

Age-related loss of muscle mass and function is associated with increased frailty and loss of independence. The mechanisms underlying the susceptibility of different muscle types to age-related atrophy are not fully understood. Reactive oxygen species (ROS) are recognised as important signalling molecules in healthy muscle and redox sensitive proteins can respond to intracellular changes in ROS concentrations modifying reactive thiol groups on Cysteine (Cys) residues. Conserved Cys residues tend to occur in functionally important locations and can have a direct impact on protein function through modifications at the active site or determining protein conformation. The aim of this work was to determine age-related changes in the redox proteome of two metabolically distinct murine skeletal muscles, the quadriceps a predominantly glycolytic muscle and the soleus which contains a higher proportion of mitochondria. To examine the effects of aging on the global proteome and the oxidation state of individual redox sensitive Cys residues, we employed a label free proteomics approach including a differential labelling of reduced and reversibly oxidised Cys residues. Our results indicate the proteomic response to aging is dependent on muscle type but redox changes that occur primarily in metabolic and cytoskeletal proteins are generally preserved between metabolically distinct tissues. BIOLOGICAL SIGNIFICANCE: Skeletal muscle containing fast twitch glycolytic fibres are more susceptible to age related atrophy compared to muscles with higher proportions of oxidative slow twitch fibres. Contracting skeletal muscle generates reactive oxygen species that are required for correct signalling and adaptation to exercise and it is also known that the intracellular redox environment changes with age. To identify potential mechanisms for the distinct response to age, this article combines a global proteomic approach and a differential labelling of reduced and reversibly oxidised Cysteine residues in two metabolically distinct skeletal muscles, quadriceps and soleus, from adult and old mice. Our results indicate that the global proteomic changes with age in skeletal muscles are dependent on fibre type. However, redox specific changes are preserved across muscle types and accompanied with a reduction in the number of redox sensitive Cysteine residues.