Improved medium with EGF and BSA for differentiated human skeletal muscle cells

Epidermal growth factor (EGF) and bovine serum albumin (BSA) are both required for serum-free clonal growth of human muscle satellite cells (HMSC). However, neither inhibits differentiation of HMSC, and when both are added to a minimal serum-free differentiation medium, they enhance survival and maintenance of human myotubes. A combination of 10 ng/mL EGF and 0.5 mg/mL BSA, added to MCDB 120 plus 10 micrograms/mL insulin, increases both total protein per dish and total creatine kinase activity, and keeps the myotubes in good condition for a longer period of time. The myotubes become cross-striated and exhibit frequent spontaneous twitching. Substantial amounts of neonatal myosin heavy chain and the MM isozyme of creatine kinase are expressed, together with detectable amounts of adult fast myosin heavy chain. With regular feeding, these cultures can be maintained for at least 3 weeks with no overgrowth by mononucleate cells, and with far less degeneration than with insulin as the only supplement.