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Atrogin1/MAFbx is an ubiquitin ligase that mediates muscle atrophy in a variety of catabolic states. We recently found that H2O2 stimulates atrogin1/MAFbx gene expression. Since the cytokine tumor necrosis factor-alpha (TNF-alpha) stimulates both reactive oxygen production and general activity of the ubiquitin conjugating pathway, we hypothesized that TNF-alpha would also increase atrogin1/MAFbx gene expression. As with H2O2, we found that TNF-alpha exposure up-regulates atrogin1/MAFbx mRNA within 2 h in C2C12 myotubes. Intraperitoneal injection of TNF-alpha increased atrogin1/MAFbx mRNA in skeletal muscle of adult mice within 4 h. Exposing myotubes to either TNF-alpha or H2O2 also produced general activation of the mitogen-activated protein kinases (MAPKs): p38, ERK1/2, and JNK. The increase in atrogin1/MAFbx gene expression induced by TNF-alpha was not altered significantly by ERK inhibitor PD98059 or the JNK inhibitor SP600125. In contrast, atrogin1/MAFbx up-regulation and the associated increase in ubiquitin conjugating activity were both blunted by p38 inhibitors, either SB203580 or curcumin. These data suggest that TNF-alpha acts via p38 to increase atrogin1/MAFbx gene expression in skeletal muscle.

Cancer cachexia is a severe skeletal muscle wasting syndrome directly responsible for 30% of all cancer deaths. Cancer exerts paracrine effects on skeletal muscle that trigger myosin loss and metabolic dysfunction. Clinical trials using cyclooxygenase 2 (COX2) inhibitors to treat cachexia had some success. Therefore, we hypothesized that tumor-induced COX2 expression contributes to myosin loss and mitochondrial dysfunction. We incubated C2C12 myotubes with control or Lewis lung carcinoma conditioned medium (LLCCM) for 48hr. We measured protein content by western blot, and oxygen consumption rate (OCR) of intact myotubes using a Seahorse XF24. Protein content is presented as means±SD in arbitrary units (n=6/group, t test). OCR is presented in pmoles O2/min (n=8/group, repeated measures ANOVA). LLCCM treatment decreased myosin content by 70%, from 7.2±2.7 to 2.1±1 AU (p

Exogenous ROS stimulate insulin-independent glucose uptake by skeletal muscle. AMP-activated protein kinase (AMPK) is a mediator of insulin-independent glucose transport. This study tested the hypo...