Insulin-like Growth Factor-II, Phosphatidylinositol 3-Kinase, Nuclear Factor-κB and Inducible Nitric-oxide Synthase Define a Common Myogenic Signaling Pathway

Insulin-like growth factors (IGFs) are potent inducers of skeletal muscle differentiation and phosphatidylinositol (PI) 3-kinase activity is essential for this process. Here we show that IGF-II induces nuclear factor-κB (NF-κB) and nitric-oxide synthase (NOS) activities downstream from PI 3-kinase and that these events are critical for myogenesis. Differentiation of rat L6E9 myoblasts with IGF-II transiently induced NF-κB DNA binding activity, inducible nitric-oxide synthase (iNOS) expression, and nitric oxide (NO) production. IGF-II-induced iNOS expression and NO production were blocked by NF-κB inhibition. Both NF-κB and NOS activities were essential for IGF-II-induced terminal differentiation (myotube formation and expression of skeletal muscle proteins: myosin heavy chain, GLUT 4, and caveolin 3), which was totally blocked by NF-κB or NOS inhibitors in rat and human myoblasts. Moreover, the NOS substrate l -Arg induced myogenesis in the absence of IGFs in both rat and human myoblasts, and this effect was blocked by NOS inhibition. Regarding the mechanisms involved in IGF-II activation of NF-κB, PI 3-kinase inhibition prevented NF-κB activation, iNOS expression, and NO production. Moreover, IGF-II induced, through a PI 3-kinase-dependent pathway, a decrease in IκB-α protein content that correlated with a decrease in the amount of IκB-α associated with p65 NF-κB.