Abstract:

BACKGROUND: The proliferation and differentiation of skeletal muscle cells are strictly regulated by multiple factors. Recently, more and more studies have indicated that microRNAs and myogenic regulatory factors as gene regulators play important roles in myogenesis.
OBJECTIVE: To explore the temporal expression patterns of muscle-specific microRNAs (miR-1, miR-133, and miR-206) in C2C12 cells during differentiation and the regulatory function of myogenic factors.
METHODS: C2C12 myogenic differentiation was induced in DMEM containing 2% horse serum. Inverted microscope was used to observe the alteration of their appearance, and immunofluorescence for myosin heavy chain was utilized to identify the differentiated C2C12 cells. At 0, 1, 2, 3, 4, 6 days after differentiation, the cellular totaI RNA of differentiated C2C12 cells was extracted. After reverse transcription, real-time polymerase chain reaction was used to determine the expression levels of three muscle-specific microRNAs as well as three myogenic regulatory factors (MyoD, myogenin and pax7) in differentiated C2C12 cells．
RESULTS AND CONCLUSION: After C2C12 cells were induced for 3 days, myotubes and MHC-positive cells began to form. Later on, more and more myotubes and MHC-positive cells appear and peaked at 6 days. The expression levels of miR-1, miR-133, miR-206, MyoD and myogenin were up regulated at first during C2C12 differentiation, and then recovered close to the levels of pre-differentiation. However, pax7 was not altered. The expression levels of muscle-specific microRNAs, MyoD and myogenin during C2C12 myogenic differentiation alter so strikingly that they might have effects on C2C12 differentiation; MyoD and myogenin might be involved in the induction of muscle-specific microRNAs.