Heat shock protein 70 (Hsp70) is a highly conserved and ubiquitous protein that is reported to provide cytoprotection in various cell types and tissues. However, the importance of Hsp70 expression during skeletal muscle atrophy, when Hsp70 levels are significantly decreased, is not known. The current study aimed to determine whether plasmid-mediated overexpression of Hsp70, in the soleus muscle of rats, was sufficient to regulate specific atrophy signaling pathways and attenuate skeletal muscle disuse atrophy. We found that Hsp70 overexpression prevented disuse muscle fiber atrophy and inhibited the increased promoter activities of atrogin-1 and MuRF1. Importantly, the transcriptional activities of Foxo3a and NF-kappaB, which are implicated in the regulation of atrogin-1 and MuRF1, were abolished by Hsp70. These data suggest that Hsp70 may regulate key atrophy genes through inhibiting Foxo3a and NF-kappaB activities during disuse. Indeed, we show that specific inhibition of Foxo3a prevented the increases in both atrogin-1 and MuRF1 promoter activities during disuse. However, inhibition of NF-kappaB did not affect the activation of either promoter, suggesting its requirement for disuse atrophy is through its regulation of other atrophy genes. We conclude that overexpression of Hsp70 is sufficient to inhibit key atrophy signaling pathways and prevent skeletal muscle atrophy.

Powers SK, Kavazis AN, McClung JM. Oxidative stress and disuse muscle atrophy.

McClung JM, Judge AR, Powers SK, Yan Z. p38 MAPK links oxidative stress to autophagy-related gene expression in cachectic muscle wasting. Am J Physiol Cell Physiol 298: C542-C549, 2010. First published December 2, 2009; doi:10.1152/ajpcell.00192.2009.-Oxidative stress is a primary trigger of cachectic muscle wasting, but the signaling pathway(s) that links it to the muscle wasting processes remains to be defined. Here, we report that activation of p38 mitogenactivated protein kinase (MAPK) (phosphorylation) and increased oxidative stress (trans-4-hydroxy-2-nonenal protein modification) in skeletal muscle occur as early as 8 h after lipopolysaccharide (1 mg/kg) and 24 h after dexamethasone (25 mg/kg) injection (intraperitoneal) in mice, concurrent with upregulation of autophagy-related genes, Atg6, Atg7, and Atg12. Treating cultured C2C12 myotubes with oxidant hydrogen peroxide (4 h) resulted in increased p38 phosphorylation and reduced FoxO3 phosphorylation along with induced Atg7 mRNA expression without activation of NF-B or FoxO3a transcriptional activities. Furthermore, inhibition of p38␣/␤ by SB202190 blocked hydrogen peroxide-induced atrophy with diminished upregulation of Atg7 and atrogenes [muscle atrophy F-box protein (MAFbx/Atrogin-1), muscle ring finger protein 1 (MuRF-1), and Nedd4]. These findings provide direct evidence for p38␣/␤ MAPK in mediating oxidative stress-induced autophagy-related genes, suggesting that p38␣/␤ MAPK regulates both the ubiquitin-proteasome and the autophagylysosome systems in muscle wasting. skeletal muscle; atrophy; cachexia SKELETAL MUSCLE ATROPHY OCCURS as a consequence of numerous pathological conditions, including cachexia and disuse (reviewed in Refs. 35 and 41). Cachectic muscle wasting is a complex process that proceeds via a rapid decline in protein synthesis and a large, sustained increase in protein degradation. In concert with other pathological and functional changes, such as decreases in mitochondrial function, capillary supply, and contractile force production, cachectic muscle wasting poses a profound negative impact on physical performance and metabolism resulting in diminished quality of life and increased mortality. Understanding the molecular and signaling mechanisms underlying cachectic muscle wasting is of critical value to improving survival and quality of life in these patient populations.Many cellular signaling events contribute to skeletal muscle atrophy (20,27,37,39,43,45,50), and research in these areas has begun to explore the molecular signaling responsible for tipping the homeostatic balance of protein metabolism toward proteolysis and suggested the importance of two major proteolysis systems: the ubiquitin-proteasome system (UPS) and the autophagy-lysosome system (ALS) (37,43,50). In particular, activation of the forkhead box O (FoxO) class of transcription factors, which includes forkhead homolog 1 in rhabdomyosarcoma/ forkhead box O1 (FKHR/FoxO1), forkhead homolog in rhabdomyosarcoma like 1/forkhead box O3 (FKHR-L1/FoxO3), and AFX/FoxO4 (43)...

Rationale: Unloading the diaphragm via mechanical ventilation (MV) results in rapid diaphragmatic fiber atrophy. It is unknown whether the myonuclear domain (cytoplasmic myofiber volume/ myonucleus) of diaphragm myofibers is altered during MV. Objective: We tested the hypothesis that MV-induced diaphragmatic atrophy is associated with a loss of myonuclei via a caspase-3-mediated, apoptotic-like mechanism resulting in a constant myonuclear domain. Methods: To test this postulate, Sprague-Dawley rats were randomly assigned to a control group or to experimental groups exposed to 6 or 12 h of MV with or without administration of a caspase-3 inhibitor. Measurements and Main Results:After 12 h of MV, type I and type IIa diaphragm myofiber areas were decreased by 17 and 23%, respectively, and caspase-3 inhibition attenuated this decrease. Diaphragmatic myonuclear content decreased after 12 h of MV and resulted in the maintenance of a constant myonuclear domain in all fiber types. Both 6 and 12 h of MV resulted in caspase-3-dependent increases in apoptotic markers in the diaphragm (e.g., number of terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling positive nuclei and DNA fragmentation). Caspase-3-dependent increases in apoptotic markers occurred after 6 h of MV, before the onset of myofiber atrophy. Conclusions: Collectively, these data support the hypothesis that the myonuclear domain of diaphragm myofibers is maintained during prolonged MV and that caspase-3-mediated myonuclear apoptosis contributes to this process. Keywords: muscle atrophy; respiratory muscle; apoptosis; ventilatory weaning Mechanical ventilation (MV) is a clinical intervention for patients who are unable to maintain adequate alveolar ventilation. Recent evidence reveals that controlled MV results in a swift progression of diaphragmatic atrophy and weakness (1-6). It seems that this diaphragmatic atrophy and weakness contributes to difficulty in weaning patients from the ventilator (7). The mechanism(s) responsible for the rapid onset of diaphragmatic atrophy and weakness are not fully understood. Therefore, delineating these mechanisms is a prerequisite for the development of therapeutic strategies to circumvent weaning difficulties. Although mechanical ventilation-induced diaphragm inactivity results in fiber atrophy, it is unknown if prolonged mechanical ventilation is associated with alterations in myonuclear domain via apoptotic mechanisms. What This Study Adds to the FieldOur results reveal that inhibiting caspase-3 activation and myonuclear loss during mechanical ventilation attenuates diaphragmatic muscle atrophy.Mechanical ventilation-induced diaphragmatic atrophy and contractile dysfunction is characterized by oxidative stress and stress-related gene expression in myofibers that occurs within a matter of hours (7,8). In addition to myofibrillar protein loss, extracellular matrix expansion, and metabolic enzyme alterations (9-11), prolonged disuse of skeletal muscle results in the selective loss of myonuclei (12-16). Myonu...