Expression of insulin-like growth factor II (IGF-II), IGF binding protein-2 and myogenin during differentiation of myogenic satellite cells derived from the turkey.

Myogenic satellite cells are essential for the development of postnatal skeletal muscle. The proliferation and differentiation of these cells are, in part, regulated by the insulin-like growth factors (IGFs), and it has been shown that the IGF binding proteins (IGFBPs) are capable of modulating the actions of IGFs. We have examined the endogenous expression of IGF-II, IGFBP-2 and the myogenic regulatory factor, myogenin, during differentiation of clonally derived turkey muscle satellite cells. Cells were harvested at approximately 80% of confluent density. Additional cultures were rinsed, fed differentiation medium and harvested when approximately 20%, 60% and 80% differentiated (fused). Northern blot analyses were performed using total cellular RNA and labeled rat cDNAs specific for IGF-II, IGFBP-2 and myogenin. A single IGF-II mRNA transcript of approximately 4.0 kb was observed. The relative mRNA abundance was highest in proliferating cultures and decreased with the onset of differentiation, to approximately 60% of initial levels where it remained throughout differentiation. Use of the IGFBP-2 cDNA probe indicated a single mRNA transcript of approximately 2.0 kb. The level of expression of IGFBP-2 mRNA was highest in proliferating cells and decreased to 25%, 16% and 11% of initial levels as differentiation progressed. A single 1.8 kb mRNA transcript was detected with the myogenin probe. Expression of myogenin was undetectable in proliferating cultures and increased significantly as differentiation progressed. Serum-free medium was conditioned for 24 h (CM) at each time point and collected from similar cultures. An IGFBP species of M(r) approximately 30,000 was detected in CM by probing western blots with [125I] IGF-I (ligand blot analysis). The intensity of this band decreased with differentiation to 35%, 24% and 18% of the level for proliferating cultures. Western blots were also probed with an antibody raised against the M(r)-34,000 bovine IGFBP-2. This antibody specifically bound to the M(r)-30,000 IGFBP, and the level of antibody binding decreased as differentiation progressed. It therefore appears that IGF-II, IGFBP-2 and myogenin are expressed in a differentiation-dependent manner by turkey myogenic satellite cells and may thus be involved in the process of differentiation of avian muscle cells.