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Tumor necrosis factor‐α (TNF) effects on mitochondrial metabolism in C2C12 myotubes

TNF appears to function as an endocrine stimulus causing muscle dysfunction during chronic inflammatory conditions. TNF‐induced reactive oxygen species (ROS) in skeletal muscle act on myofibrillar proteins to diminish force, yet their source remains elusive. Our data intimate that TNF signals via the TNF receptor 1 subtype to activate neutral sphingomyelinase 3, increasing ceramide and stimulating ROS production. In murine skeletal muscle cells (C2C12s), mitochondrial‐specific ROS inhibitor, SS31, ablates TNF‐induced increase in dichlorfluorescein‐diacetate fluorescence (P<.002). We hypothesize that TNF alters mitochondrial electron transport chain function. Control experiments using an automated extracellular flux analyzer have shown oxygen consumption rate (OCR) in untreated C2C12s to be stable (352±7pmoles/min) over 90 minutes. In initial experiments testing uncoupled, state 3, and state 4 respiration, it appears that TNF may lower maximal respiration (p<.07) and increase extracellular acidification rate (p<.06). When exposed to inhibitors for complexes IV and V, control and TNF OCR were nearly identical, which suggests TNF‐induced proton leak originates upstream of Complex IV. These data suggest TNF alters mitochondrial metabolism, which may contribute to increased ROS production. Supported by NIH grant 5 AR055974–04 (MBR) and NIH/NINDS P30 NS051220 (PGS).