Effects of aerobic exercise on cardiac function and gene expression of NADPH oxidases in diaphragm muscle of rats with aortic stenosis-induced heart failure.

We evaluated the influence of aerobic physical exercise (EX) on gene-encoding proteins associated with oxidative stress in diaphragm muscle of rats with aortic stenosis-induced heart failure (HF). Wistar male rats were divided into four groups: Control sedentary (C); Control exercise (C-Ex); Sedentary aortic stenosis (AS); Aortic stenosis exercise (AS-Ex). Exercised rats trained 5 times a week for 10 weeks on a treadmill. Statistical analysis was performed by ANOVA or Kruskal-Wallis test. In the final echocardiogram, animals with aortic stenosis subjected to exercise demonstrated improvement in systolic function compared to the sedentary aortic stenosis group. In diaphragm muscle, the activity of antioxidant enzymes, malondialdehyde malondialdehyde concentration, protein carbonylation, and protein expression of p65 and its inhibitor IκB did not differ between groups. Alterations in gene expression of sources that generate reactive species of oxygen were observed in AS-Ex group, which showed decreased mRNA abundance of NOX2 and NOX4 compared to the aortic stenosis group (p < 0.05). We concluded that aerobic exercise has a positive impact during heart failure, ameliorating systolic dysfunction and biomarkers of oxidative stress in diaphragm muscle of rats with aortic stenosis-induced heart failure.

Nox2 Inhibition Regulates Stress Response and Mitigates Skeletal Muscle Fiber Atrophy during Simulated Microgravity.

Insufficient stress response and elevated oxidative stress can contribute to skeletal muscle atrophy during mechanical unloading (e.g., spaceflight and bedrest). Perturbations in heat shock proteins (e.g., HSP70), antioxidant enzymes, and sarcolemmal neuronal nitric oxidase synthase (nNOS) have been linked to unloading-induced atrophy. We recently discovered that the sarcolemmal NADPH oxidase-2 complex (Nox2) is elevated during unloading, downstream of angiotensin II receptor 1, and concomitant with atrophy. Here, we hypothesized that peptidyl inhibition of Nox2 would attenuate disruption of HSP70, MnSOD, and sarcolemmal nNOS during unloading, and thus muscle fiber atrophy. F344 rats were divided into control (CON), hindlimb unloaded (HU), and hindlimb unloaded +7.5 mg/kg/day gp91ds-tat (HUG) groups. Unloading-induced elevation of the Nox2 subunit p67phox-positive staining was mitigated by gp91ds-tat. HSP70 protein abundance was significantly lower in HU muscles, but not HUG. MnSOD decreased with unloading; however, MnSOD was not rescued by gp91ds-tat. In contrast, Nox2 inhibition protected against unloading suppression of the antioxidant transcription factor Nrf2. nNOS bioactivity was reduced by HU, an effect abrogated by Nox2 inhibition. Unloading-induced soleus fiber atrophy was significantly attenuated by gp91ds-tat. These data establish a causal role for Nox2 in unloading-induced muscle atrophy, linked to preservation of HSP70, Nrf2, and sarcolemmal nNOS.

Effect of combined fish oil & Curcumin on murine skeletal muscle morphology and stress response proteins during mechanical unloading.

Skeletal muscle is a highly adaptable tissue capable of remodeling when dynamic stress is altered, including changes in mechanical loading and stretch. When muscle is subjected to an unloaded state (e.g., bedrest, immobilization, spaceflight) the resulting loss of muscle cross sectional area (CSA) impairs force production. In addition, muscle fiber-type shifts from slow to fast-twitch fibers. Unloading also results in a downregulation of heat shock proteins (e.g., HSP70) and anabolic signaling, which further exacerbate these morphological changes. Our lab recently showed reactive oxygen species (ROS) are causal in unloading-induced alterations in Akt and FoxO3a phosphorylation, muscle fiber atrophy, and fiber-type shift. Nutritional supplements such as fish oil and curcumin enhance anabolic signaling, glutathione levels, and heat shock proteins. We hypothesized that fish oil, rich in omega-3-fatty acids, combined with the polyphenol curcumin would enhance stress protective proteins and anabolic signaling in the rat soleus muscle, concomitant with synergistic protection of morphology. C57BL/6 mice were assigned to 3 groups (n = 6/group): ambulatory controls (CON), hindlimb unloading (HU), and hindlimb unloading with 5% fish oil, 1% curcumin in diet (FOC). FOC treatments began 10 days prior to HU and tissues were harvested following 7 days of HU. FOC mitigated the unloading induced decrease in CSA. FOC also enhanced abundance of HSP70 and anabolic signaling (Akt phosphorylation, p70S6K phosphorylation), while reducing Nox2, a source of oxidative stress. Therefore, we concluded that the combination of fish oil and curcumin prevents skeletal muscle atrophy due to a boost of heat shock proteins and anabolic signaling in an unloaded state.

Effect of Eukarion-134 on Akt-mTOR signalling in the rat soleus during 7 days of mechanical unloading.

NEW FINDINGS: What is the central question of this study? Translocation of nNOSµ initiates catabolic signalling via FoxO3a and skeletal muscle atrophy during mechanical unloading. Recent evidence suggests that unloading-induced muscle atrophy and FoxO3a activation are redox sensitive. Will a mimetic of superoxide dismutase and catalase (i.e. Eukarion-134) also mitigate suppression of the Akt-mTOR pathway? What is the main finding and its importance? Eukarion-134 rescued Akt-mTOR signalling and sarcolemmal nNOSµ, which were linked to protection against the unloading phenotype, muscle fibre atrophy and partial fibre-type shift from slow to fast twitch. The loss of nNOSµ from the sarcolemma appears crucial to Akt phosphorylation and is redox sensitive, although the mechanisms remain unresolved. ABSTRACT: Mechanical unloading stimulates rapid changes in skeletal muscle morphology, characterized by atrophy of muscle fibre cross-sectional area and a partial fibre-type shift from slow to fast twitch. Recent studies revealed that oxidative stress contributes to activation of forkhead box O3a (FoxO3a), proteolytic signalling and unloading-induced muscle atrophy via translocation of the µ-splice variant of neuronal nitric oxide synthase (nNOSµ) and activation of FoxO3a. There is limited understanding of the role of reactive oxygen species in the Akt-mammalian target of rapamycin (mTOR) pathway signalling during unloading. We hypothesized that Eukarion-134 (EUK-134), a mimetic of the antioxidant enzymes superoxide dismutase and catalase, would protect Akt-mTOR signalling in the unloaded rat soleus. Male Fischer 344 rats were separated into the following three study groups: ambulatory control (n = 11); 7 days of hindlimb unloading + saline injections (HU, n = 11); or 7 days of HU + EUK-134; (HU + EUK-134, n = 9). EUK-134 mitigated unloading-induced dephosphorylation of Akt, as well as FoxO3a, in the soleus. Phosphorylation of mTOR in the EUK-treated HU rats was not different from that in control animals. However, EUK-134 did not significantly rescue p70S6K phosphorylation. EUK-134 attenuated translocation of nNOSµ from the membrane to the cytosol, reduced nitration of tyrosine residues and suppressed upregulation of caveolin-3 and dysferlin. EUK-134 ameliorated HU-induced remodelling, atrophy of muscle fibres and the 12% increase in type II myosin heavy chain-positive fibres. Attenuation of the unloaded muscle phenotype was associated with decreased reactive oxygen species, as assessed by ethidium-positive nuclei. We conclude that oxidative stress affects Akt-mTOR signalling in unloaded skeletal muscle. Direct linkage of abrogation of nNOSµ translocation with Akt-mTOR signalling during unloading is the subject of future investigation.