Defects in neuromuscular innervation contribute significantly to the age-related decline in muscle mass and function (sarcopenia). Our previous studies demonstrated that denervation induces muscle mitochondrial hydroperoxide production (H 2 o 2 and lipid hydroperoxides (LOOHs)). Here we define the relative contribution of mitochondrial electron transport chain (ETC) derived H 2 o 2 versus cytosolic phospholipase A 2 (cPLA 2) derived LOOHs in neurogenic muscle atrophy. We show that denervation increases muscle cPLA 2 protein content, activity, and metabolites downstream of cPLA 2 including LOOHs. Increased scavenging of mitochondrial H 2 o 2 does not protect against denervation atrophy, suggesting ETC generated H 2 o 2 is not a critical player. In contrast, inhibition of cPLA 2 in vivo mitigates LOOH production and muscle atrophy and maintains individual muscle fiber size while decreasing oxidative damage. Overall, we show that loss of innervation in several muscle atrophy models including aging induces generation of LOOHs produced by arachidonic acid metabolism in the cPLA 2 pathway contributing to loss of muscle mass. The pathological age-related loss of skeletal muscle mass and function (sarcopenia) contributes to decreased quality of life and independence and increases the risk of injury and chronic disease 1,2. Preventing or reducing the effects of sarcopenia could increase quality of life, reduce risk of comorbidity development, and save billions of dollars in healthcare costs annually. Designing therapeutic interventions will remain difficult until the complex pathways and processes underlying sarcopenia are identified 3. Studies supporting a loss of innervation with age and a link between denervation and decreased muscle fiber size suggest that sarcopenia is a form of neurogenic atrophy 4-6. In addition to the loss of muscle mass, contractile force also declines with age 7. Loss of muscle strength occurs more rapidly than can be explained by the loss of muscle mass alone, and evidence suggests that a reduction in muscle quality and neuromuscular junction signaling also play a role in the functional decline of aging muscle 8. Our previous studies using mice lacking the cytosolic superoxide (O 2 −) scavenger CuZn superoxide dismutase (CuZn SOD, Sod1 −/−) demonstrate that elevated oxidative stress leads to an accelerated sarcopenia in the Sod1 −/− mice that is characterized by denervation, muscle hydroperoxide production, and muscle atrophy 9-11. Returning expression of CuZnSOD specifically to motor neurons of the Sod1 −/− mice prevented denervation, muscle hydroperoxide production, and atrophy, supporting a link between loss of innervation, hydroperoxides, and muscle atrophy 12. We have also shown that loss of innervation to skeletal muscle directly induces basal hydroperoxide production from isolated mitochondria including both hydrogen peroxide (H 2 O 2) and lipid hydroperoxides (LOOHs) 11. The magnitude of this

Background Cancer is associated with muscle atrophy (cancer cachexia) that is linked to up to 40% of cancer-related deaths. Oxidative stress is a critical player in the induction and progression of age-related loss of muscle mass and weakness (sarcopenia); however, the role of oxidative stress in cancer cachexia has not been defined. The purpose of this study was to examine if elevated oxidative stress exacerbates cancer cachexia. Methods Cu/Zn superoxide dismutase knockout (Sod1KO) mice were used as an established mouse model of elevated oxidative stress. Cancer cachexia was induced by injection of one million Lewis lung carcinoma (LLC) cells or phosphate-buffered saline (saline) into the hind flank of female wild-type mice or Sod1KO mice at approximately 4 months of age. The tumour developed for 3 weeks. Muscle mass, contractile function, neuromuscular junction (NMJ) fragmentation, metabolic proteins, mitochondrial function, and motor neuron function were measured in wild-type and Sod1KO saline and tumour-bearing mice. Data were analysed by two-way ANOVA with Tukey-Kramer post hoc test when significant F ratios were determined and α was set at 0.05. Unless otherwise noted, results in abstract are mean ±SEM. Results Muscle mass and cross-sectional area were significantly reduced, in tumour-bearing mice. Metabolic enzymes were dysregulated in Sod1KO mice and cancer exacerbated this phenotype. NMJ fragmentation was exacerbated in tumour-bearing Sod1KO mice. Myofibrillar protein degradation increased in tumour-bearing wild-type mice (wild-type saline, 0.00847 ± 0.00205; wildtype LLC, 0.0211 ± 0.00184) and tumour-bearing Sod1KO mice (Sod1KO saline, 0.0180 ± 0.00118; Sod1KO LLC, 0.0490 ± 0.00132). Muscle mitochondrial oxygen consumption was reduced in tumour-bearing mice compared with saline-injected wild-type mice. Mitochondrial protein degradation increased in tumour-bearing wild-type mice (wild-type saline, 0.0204 ± 0.00159; wild-type LLC, 0.167 ± 0.00157) and tumour-bearing Sod1KO mice (Sod1KO saline, 0.0231 ± 0.00108; Sod1 KO LLC, 0.0645 ± 0.000631). Sciatic nerve conduction velocity was decreased in tumour-bearing wild-type mice (wild-type saline, 38.2 ± 0.861; wild-type LLC, 28.8 ± 0.772). Three out of eleven of the tumour-bearing Sod1KO mice did not survive the 3-week period following tumour implantation. Conclusions Oxidative stress does not exacerbate cancer-induced muscle loss; however, cancer cachexia may accelerate NMJ disruption.

Age‐related muscle atrophy and weakness, or sarcopenia, are significant contributors to compromised health and quality of life in the elderly. While the mechanisms driving this pathology are not fully defined, reactive oxygen species, neuromuscular junction (NMJ) disruption, and loss of innervation are important risk factors. The goal of this study is to determine the impact of mitochondrial hydrogen peroxide on neurogenic atrophy and contractile dysfunction. Mice with muscle‐specific overexpression of the mitochondrial H2O2 scavenger peroxiredoxin3 (mPRDX3) were crossed to Sod1KO mice, an established mouse model of sarcopenia, to determine whether reduced mitochondrial H2O2 can prevent or delay the redox‐dependent sarcopenia. Basal rates of H2O2 generation were elevated in isolated muscle mitochondria from Sod1KO, but normalized by mPRDX3 overexpression. The mPRDX3 overexpression prevented the declines in maximum mitochondrial oxygen consumption rate and calcium retention capacity in Sod1KO. Muscle atrophy in Sod1KO was mitigated by ~20% by mPRDX3 overexpression, which was associated with an increase in myofiber cross‐sectional area. With direct muscle stimulation, maximum isometric specific force was reduced by ~20% in Sod1KO mice, and mPRDX3 overexpression preserved specific force at wild‐type levels. The force deficit with nerve stimulation was exacerbated in Sod1KO compared to direct muscle stimulation, suggesting NMJ disruption in Sod1KO. Notably, this defect was not resolved by overexpression of mPRDX3. Our findings demonstrate that muscle‐specific PRDX3 overexpression reduces mitochondrial H2O2 generation, improves mitochondrial function, and mitigates loss of muscle quantity and quality, despite persisting NMJ impairment in a murine model of redox‐dependent sarcopenia.

Emerging evidence suggests that patients with Alzheimer’s disease (AD) may show accelerated sarcopenia phenotypes. To investigate whether pathological changes associated with neuronal death and cognitive dysfunction also occur in peripheral motor neurons and muscle as a function of age, we used the triple transgenic mouse model of AD (3xTgAD mice) that carries transgenes for mutant forms of APP, Tau, and presenilin proteins that are associated with AD pathology. We measured changes in motor neurons and skeletal muscle function and metabolism in young (2 to 4 month) female control and 3xTgAD mice and in older (18–20 month) control and 3xTgAD female mice. In older 3xTgAD mice, we observed a number of sarcopenia-related phenotypes, including significantly fragmented and denervated neuromuscular junctions (NMJs) associated with a 17% reduction in sciatic nerve induced vs. direct muscle stimulation induced contractile force production, and a 30% decrease in gastrocnemius muscle mass. On the contrary, none of these outcomes were found in young 3xTgAD mice. We also measured an accumulation of amyloid-β (Aβ) in both skeletal muscle and neuronal tissue in old 3xTgAD mice that may potentially contribute to muscle atrophy and NMJ disruption in the older 3xTgAD mice. Furthermore, the TGF-β mediated atrophy signaling pathway is activated in old 3xTgAD mice and is a potential contributing factor in the muscle atrophy that occurs in this group. Perhaps surprisingly, mitochondrial oxygen consumption and reactive oxygen species (ROS) production are not elevated in skeletal muscle from old 3xTgAD mice. Together, these results provide new insights into the effect of AD pathological mechanisms on peripheral changes in skeletal muscle.

Mitochondrial dysfunction, reactive oxygen species (ROS) and oxidative damage have been implicated to play a causative role in age-related skeletal muscle atrophy and weakness (i.e. sarcopenia). Mice lacking the superoxide scavenger CuZnSOD (Sod1−/−) exhibit high levels of oxygen-derived radicals and oxidative damage, associated with neuronal and muscular phenotypes consistent with sarcopenia. We used magnetic resonance imaging (MRI) technology combined with immunospin-trapping (IST) to measure in vivo free radical levels in skeletal muscle from wildtype, Sod1−/− and SynTgSod1−/− mice, a mouse model generated using targeted expression of the human Sod1 transgene specifically in neuronal tissues to determine the impact of motor neuron degeneration in muscle atrophy. By combining the spin trap DMPO (5,5-dimethyl-1-pyrroline N-oxide) and molecular MRI (mMRI), we monitored the level of free radicals in mouse hindlimb muscle. The level of membrane-bound macromolecular radicals in the quadriceps muscle was elevated by ~3-fold in Sod1−/− mice, but normalized to wildtype levels in SynTgSod1−/− rescue mice. Skeletal muscle mass was reduced by ~25–30% in Sod1−/− mice, but fully reversed in muscle from SynTgSod1−/− mice. Using perfusion MRI we also measured the dynamics of blood flow within mouse hindlimb. Relative muscle blood flow in Sod1−/− is decreased to ~50% of wildtype and remained low in the SynTgSod1−/− mice. Our findings are significant in that we have shown for the first time that in vivo free radical production in skeletal muscle is directly correlated to muscle atrophy in an experimental model of oxidative stress. Neuron-specific expression of CuZnSOD reverses the in vivo free radical production in skeletal muscle in the Sod1−/− mouse model and prevents muscle atrophy. These results further support the feasibility of using in vivo assessments of redox status in the progression of a pathological process such as sarcopenia. This approach can also be valuable for evaluating responses to pharmacologic interventions.