Insulin Signaling and Caveolae: Role of TNF alpha in the development of insulin resistance in C2C12 myocytes

Abstract

Recent research has shown that TNF‐á affects insulin signaling by up‐regulating ganglioside‐GM3 production in adipocytes; however little is known about these effects in skeletal muscle. Caveolae are specialized plasma membrane microdomains thought to serve as signaling platforms for several receptors including the insulin receptor (IR) and glucose transporters (Glut‐1, ‐4), via interaction with caveolin proteins (Cav‐1, ‐3). We hypothesized that exposure of C2C12 cells to TNFá will result in deregulation of the insulin signaling cascade and alteration of the pattern of IR and Glut‐4 localization to the plasma membrane by promoting a shift from the caveolae to other membrane domains enriched in ganglioside‐GM3 expression. Here, C2C12 myocytes were differentiated into skeletal myotubes and treated with TNF‐á, insulin, and a TNF‐á antagonist. Treated cells were subjected to Western blot analysis with antibodies against IR, phosphorylated IR, Akt, GM3‐synthase, Glut‐4, Cav‐1, Cav‐3, and actin (loading control). Western blot demonstrated higher levels of AKT, and GM3‐S after treatment with TNF‐á. Immunofuorescence analysis demonstrated a disruption of the colocalization of IR and Glut‐4 to caveolae domains in cells treated with TNF‐á, which was reversed by treatment with the antagonist. These results identify TNF‐á and gangliosides as potential therapeutic targets during insulin resistance.