Gene Expression Profiles Induced by Growth Factors in In Vitro Cultured Myocytes

Disruption in the normal adolescent growth spurt can cause the spinal deformities that result in idiopathic scoliosis. Effects of insulin-like growth factor 1 (IGF1) and fibroblast growth factor 2 (FGF2) on the expression of genes involved in the proliferation and differentiation of myocytes in culture were analyzed. Human myocytes in in vitro culture were treated with IGF1 or FGF2 (10 ng/ml) for 24 hours. Experiments were performed during the exponential growth phase with approximately 1e7 cells per 75 cm 2 flask. mRNA was reverse transcribed directly and analyzed using RT-PCR Taqman assays. Expression levels of key genes involved in cell growth and differentiation (CHD 7, HDAC 5, ACTA 1, LEF1, WNT5A, COL1A1, COL2A1, ACAN, FGF7 and VCAN) were monitored using RT-PCR with gene-specific Taqman probes. Two patterns of response to the growth factors were observed: Five genes (CHD7, HDAC5, COL1A1, ACAN, LEF1) were stimulated in their level of expression by IGF-1 with lesser or no effects of FGF2, and one gene (WNT5) was even down regulated by the addition of FGF2. Only ACTA1 showed an increased expression level that was augmented higher by FGF2 than by IGF-1.In summary, we could explicate the feasibility of our myocyte culture system to study genes with possible implication in the development of scoliosis. Growth factor addition to these cells exhibit differential effects simulating eventually the changing growth factor environment during puberty. Any disturbance of the intricate pattern of the various pathways studied might have long lasting effects on skeletal muscle development leading to human disease and might be probed in affected individuals.