OVEREXPRESSION OF INSULIN-LIKE GROWTH FACTOR-II INDUCES ACCELERATED MYOBLAST DIFFERENTIATION

Previous studies have shown that exogenous insulin-like growth factors (IGFs) can stimulate the terminal differentiation of skeletal myoblas ts in culture and have established a correlation between the rate and the extent of ICF-II secretion by muscle cell lines and the rate of bi ochemical and morphological differentiation. To investigate the hypoth esis that autocrine secretion of IGF-II plays a critical role in stimu lating spontaneous myogenic differentiation in vitro, we have establis hed C2 muscle cell lines that stably express a mouse IGF-II cDNA under control of the strong, constitutively active Moloney sarcoma virus pr omoter, enabling us to study directly the effects of ICF-II overproduc tion. Similar to observations with other muscle cell lines, ICF-II ove rexpressing myoblasts proliferated normally in growth medium containin g 20% fetal serum, bur they underwent enhanced differentiation compare d with controls when incubated in low-serum differentiation medium. Ac celerated differentiation of IGF-II overexpressing C2 cells was preced ed by the rapid induction of myogenin mRNA and protein expression (wit hin 1 h, compared with 24-48 h in controls) and was accompanied by an enhanced proportion of the retinoblastoma protein in an underphosphryl ated and potentially active form, by a marked increase in activity of the muscle-specific enzyme, creatine phosphokinase, by extensive myotu be formation by 48 h, and by elevated secretion of IGF binding protein -5 when compared with controls. These results confirm a role for IGF-I I as an autocrine/paracrine differentiation factor for skeletal myobla sts, and they define a model cell system that will be useful in determ ining the biochemical mechanisms of ICF action in cellular differentia tion. (C) 1996 Wiley-Liss, Inc.