Objective: To verify whether insulin promotes the differentiation of skeletal myoblasts into myocytes and whether wortmannin and U0126 inhibit the promoting effect with an attempt to explore the molecular mechanisms of inducing the differentiation of muscular cells in rats.
Methods: Primary skeletal myoblasts were separated and cultured from rats, and then were treated in DMEM containing various concentrations of insulin. The morphology of cells was monitored under a phase-contrast microscope. And the expression of myogenin was detected by immunocytochemistry and Western blotting. The change in the effect of insulin on the differentiation of myoblasts was observed after the intervention of a phosphatidylinositide 3-kinase (PI3K) inhibitor wortmannin and a specific MEK inhibitor U0126.
Results: Insulin markedly promoted myotube formation of myoblasts. Two days after insulin treatment, myotubes started to form; later, more and more myotubes appeared, and to the peak at 7 days. Insulin increased the expression of myogenin in a concentration-dependent manner. However, wortmannin and U0126 inhibited the effect of insulin on the differentiation of skeletal myoblasts in rats.
Conclusion: Wortmannin and U0126 can suppress the promoting effect of insulin on the differentiation of skeletal myoblasts into myocytes in rats and decrease the formation of myotubes and the expression of myogenin.