The Effects of Imbalance Between Histone Acetylation and Deacetylation on C2C12 Myoblasts Differentiation

Abstract

Objective To explore the effects of histone acetylation and deacetylation on C2C12 myoblasts differentiation. Methods Based on the differentiation of C2C12 myoblasts into myotubes using high glucose dulbecco's modified eagle medium (DMEM) containing 2% horse serum in vitro, valproic acid(VPA) was given to C2C12 myoblasts during differentiation with different concentrations, which contained 1 mmol/L VPA, 2 mmol/L VPA, 4 mmol/L VPA and 8 mmol/L VPA in the final concentrations with 2% horse serum and high glucose DMEM. So that this experiment was scheduled into 6 groups as control group(contain 10% fetal bovine serum in growth medium), horse serum induced differentiation group, 1 mmol/L VPA group, 2 mmol/L VPA group, 4 mmol/L VPA group and 8 mmol/L VPA group according to different growth medium. There were 6 samples in each group. The myotube differentiation rate were compared in different concentration VPA groups and horse serum induced differentiation group. Besides, mRNA and protein expression levels of muscle-related proteins(including Myosin, Troponin I-SS, myogenic differentiation 1) and histone deacetylases (HDAC, including HDAC1, 2, 3) were also evaluated with real time polymerase chain reaction (RT-PCR) and Western blotting. The mRNA and protein expression levels of them were compared and analyzed. Results ①The mRNA and protein expression levels of muscle-related proteins of horse serum induced differentiation group were significantly higher than those of control group, and the differences were statistically significant (P 0.05). ②The myotube differentiation rate in every concentration VPA group compared with horse serum induced differentiation group were significantly lower, and the differences were statistically significant (P<0.05). ③ The mRNA and protein expression levels of muscle-related proteins and HDAC in 4 mmol/L VPA group or 8 mmol/L VPA group compared with horse serum induced differentiation group were significantly decreased, and the differences were statistically significant (P<0.05). ④The impacts of VPA on the expression of muscle-related proteins and HDAC in mRNA level and protein level during C2C12 differentiation were identified as a concentration dependent pattern, indicated a negative linear correlation(r=-0.92, -0.89, -0.93, -0.84, -0.93, -0.97; r=-0.99, -0.86, -0.92, -0.95, -0.91, -0.96; P<0.05). Conclusions Histone acetylation and deacetylation played an important role in skeletal muscle cell differentiation. The inhibition of HDAC could result in the reduction of transcription and expression of muscle-related proteins and suppressed the differentiation of myoblasts. Key words: Histone acetyltransferases; Histone deacetylases; Cell differentiation; Valproic acid