Leucine increases mitochondrial metabolism and lipid content without altering insulin signaling in myotubes

Elevated circulating branched-chain amino acids (BCAA) such as leucine have been consistently correlated with increasing severity of insulin resistance across numerous populations. BCAA may promote insulin resistance through either mTOR-mediated suppression of insulin receptor substrate-1 or through the accumulation of toxic BCAA catabolites. Although the link between circulating BCAA and insulin resistance has been consistent, it has yet to be concluded if BCAA causally contribute to the development or worsening of insulin resistance. This work investigated the effect of leucine both with and without varying levels of insulin resistance on metabolism, metabolic gene expression, and insulin signaling. C2C12 myotubes were treated with and without varied concentrations of leucine up to 2 mM for 24 h both with and without varied levels of insulin resistance. Gene and protein expression were measured via qRT-PCR and Western blot, respectively. Mitochondrial metabolism was measured via O₂ consumption. Leucine at 2 mM increased oxidative metabolism as well as gene expression of mitochondrial biogenesis, which was associated with increased cellular lipid content. Despite increased lipid content of leucine-treated cells, neither acute nor chronic leucine treatment at 2 mM affected insulin signaling in insulin sensitive, mildly insulin resistant, or severely insulin resistant cells. Similarly, leucine at lower concentrations (0.25 mM, 0.5 mM, and 1 mM) did not alter insulin signaling either, regardless of insulin resistance. Leucine appears to improve myotube oxidative metabolism and related metabolic gene expression. And despite increased lipid content of leucine-treated cells, leucine does not appear to alter insulin sensitivity either acutely or chronically, regardless of level of insulin resistance.