Aims: Clinical and experimental data suggest that early insulin therapy could reduce lipotoxicity in subjects and animal models with type 2 diabetes mellitus. However, the underlying mechanisms need to be clarified. Sterol regulatory element-binding protein 1c (SREBP-1c), which is negatively regulated by AMP-activated protein kinase (AMPK), plays a critical role in lipotoxicity and insulin resistance in skeletal muscle cells. Here, we investigated the effect and molecular mechanism of insulin intervention on the AMPK/SREBP-1c pathway in skeletal muscle cells with chronic exposure to palmitic acid (PA).
Methods: Male C57BL/6 mice were fed with a high-fat diet for 12 weeks and were then treated with insulin, AMPK inhibitor, or metformin. L6 myotubes incubated with palmitic acid (PA) were treated with insulin or metformin. Dominant-negative AMPKα2 (DN-AMPKα2) lentivirus, AMPKα2 siRNA, or Rho-kinase 1 (ROCK1) siRNA were transfected into PA-treated L6 myotubes.
Results: We found that the ability of PA to stimulate SREBP-1c and inhibit AMPK was reversed by insulin in L6 cells. Moreover, DN-AMPKα2 lentivirus and AMPKα2 siRNA were transfected into PA-treated L6 myotubes, and the decrease in SREBP-1c expression caused by insulin was blocked by AMPK inhibition independent of the phosphatidylinositol-4,5-biphosphate-3-kinase (PI3K)/AKT pathway. The serine/threonine kinase Rho-kinase (ROCK) 1, a downstream effector of the small G protein RhoA, was activated by PA. Interestingly, knockdown of ROCK1 by siRNA blocked the downregulation of AMPK phosphorylation under PA-treated L6 myotubes, which indicated that ROCK1 mediated the effect of insulin action on AMPK.
Conclusions: Our study indicated that insulin reduced lipotoxicity via ROCK1 and then improved AMPK/SREBP-1c signaling in skeletal muscle under PA-induced insulin resistance.
Keywords: AMPK; Insulin; Rho-kinase 1; SREBP-1c; Skeletal muscle.