Skeletal muscle cell-derived insulin-like growth factor (IGF) binding proteins inhibit IGF-I-induced myogenesis in rat L6E9 cells

Abstract

The insulin-like growth factors (IGFs) stimulate the differentiation of skeletal muscle cells. IGF binding proteins (IGFBPs), which are expressed by skeletal muscle cells, may enhance or inhibit IGF actions. To explore the role of skeletal muscle-derived IGFBPs in IGF-induced myogenesis, we compared the differentiation-inducing effects of IGF-I and des(1-3)IGF-I in rat L6E9 skeletal myoblasts. Des(1-3)IGF-I is a naturally occurring IGF-I analog with markedly reduced affinity for IGFBPs but with an affinity for the IGF-I receptor that is comparable to that for native IGF-I. We find that rat L6E9 cells produce principally IGFBP-4 and BP-6, with a minor component of IGFBP-5. Both IGFBP-4 and BP-6 accumulate during differentiation and increase further in response to IGF-I or des(1-3)IGF-I treatment. We find that an IGF-I analog with reduced affinity for IGFBPs is significantly more potent than native IGF-I in stimulating myogenesis (as assessed by myogenin messenger RNA abundance and muscle creatine kinase activity), indicating that IGFBPs expressed by skeletal muscle cells inhibit differentiation induced by IGF-I. In view of the relative abundance of IGFBP-4, its relatively high affinity for IGF-I and the low affinity of IGFBP-6 for IGF-I, it is likely that the inhibitory effect of rat skeletal muscle-derived IGFBPs on IGF-I-induced myogenesis is mediated principally by IGFBP-4.

Characterisation of the IGF system in a primary adult human skeletal muscle cell model, and comparison of the effects of insulin and IGF-I on protein metabolism

“…6. Surprisingly, this included large amounts of intact IGFBP-3, which has not been shown to be produced by any of the other skeletal muscle cell lines examined to date (McCusker & Clemmons 1988, Tollefsen et al 1989, Florini et al 1995, Silverman et al 1995. As the anti-IGFBP-3 antibody does not cross-react with bovine IGFBP-3 in FCS (see lane 6 of the IGFBP-3 immunoblot, Fig.…”

Functional inactivation of the IGF-I receptor delays differentiation of skeletal muscle cells

“…Samples were boiled for 5 min and analyzed by SDS-PAGE on 5-12% gradient gels. After electrotransfer and membrane blocking as previously described (Silverman et al 1995), membranes were treated with an antiphosphotyrosine antibody coupled to horseradish peroxidase at a 1:1000 dilution in blocking buffer overnight at 4 C. Membranes were subsequently washed three times in blocking buffer, and immunoreactive bands were visualized using the enhanced chemiluminescence (ECL) detection system (Amersham ECL kit RPN 2108). Autoradiography was subsequently carried out.…”

Nutritional and hormonal control of skeletal-muscle cell growth and differentiation

“…At least five of the IGFBP appear to be produced by muscle cells in culture, with IGFBP-2, -3, -4, -5 and -6 being found at varying levels in different muscle cell types. For instance, the C2 muscle cell line appears to only secrete IGFBP-5 (Tollefsen et al 1989;James et al 1993 (Silverman et al 1995), as does secretion of IGFBP-5 in C2 cells (James et al 1993(James et al , 1996. The relative effects of these different binding proteins on the processes of differentiation are still at an early stage, but enhancement of IGFBP-5 production via sense transfection methods results in a decrease in differentiation of C2 cells, whereas transfection of the anti-sense construct for IGFBP-5 results in decreased secretion of IGFBP-5 and the cells differentiate prematurely and to a greater extent (James et al 1996).…”