Hormonal regulation of IGF-binding protein-2 expression in proliferating C₂C₁₂ myoblasts

No studies have investigated the hormonal regulation of IGF-binding protein-2 (IGFBP-2) secretion and mRNA expression in myoblasts. In this study, cells of the C₂C₁₂ mouse myoblast cell line were used to examine the effects of various agents on the hormonal regulation of IGFBP-2. Conditioned medium (CM) was collected and cells were harvested at 2, 4, 6, 15 and 24 h after exposure to treatment media containing porcine insulin (pINS) or recombinant human IGF-I (rhIGF-I), and at 6, 15 and 24 h after exposure to treatment media containing dexamethasone (DEX) or prostaglandin E₂ (PGE₂). Relative abundance of a single 1.8 kb IGFBP-2 mRNA transcript was determined by Northern analysis using total cellular RNA and a labeled cDNA specific for rat IGFBP-2. IGFBP-2 was detected in CM by probing Western blots with ¹²⁵I-IGF-I (ligand blot analysis). We have previously shown by immunoblot analysis that the predominant 32 000 M_r protein on ligand blots is IGFBP-2. Treatment with 10^-9 or 10^-6 M pINS led to a rapid reduction (P<0.01) in relative IGFBP-2 mRNA abundance and protein secretion relative to controls. Treatment with 7 × 10^-10 10 or 7 × 10^-9 M (5 or 50 ng/ml) rhIGF-I increased IGFBP-2 mRNA abundance and protein secretion (P<0.01). Cultures treated with 10^-8 M DEX exhibited significantly increased (P<0.001) IGFBP-2 mRNA and protein. IGFBP-2 secretion was not affected by 10^-6 M PGE₂ but mRNA levels were higher than controls at 24 h (P<0.01). These findings suggest that multiple factors, including growth factors and metabolic hormones, are involved in regulating IGFBP-2 expression in C₂C₁₂ myoblasts.