脉冲电刺激对H9c2细胞增殖及分化的影响

Effect of pulsed electrical stimulation on the proliferation and differentiation of H9c2 cells

目的观察脉冲电刺激对H9c2细胞增殖的抑制作用,探讨脉冲电刺激对H9c2细胞分化的诱导作用。方法用不同频率(0、1、5、10、50、100 Hz)、电压(0、5、10、20、40、50 V)、时间(0、1、2、4、6、8 h/d)的组合脉冲电刺激作用于大鼠H9c2细胞,以5-氮胞苷作为干预因素对照。采用MTT法检测H9c2细胞增殖率。采用免疫荧光细胞化学染色检测心肌肌钙蛋白T(cTNT)及八聚体结合转录因子4(Oct4)的表达。采用RT-PCR法检测cTNT、肌球蛋白重链α(α-MHC)mRNA的表达。结果脉冲电刺激能抑制H9c2细胞增殖(P<0.05),脉冲电刺激的参数为频率1 Hz、4 h/d、10 V时对H9c2细胞增殖的抑制作用最大且不致细胞死亡。用此参数脉冲电刺激作用于H9c2细胞2周后细胞Oct4表达减少(P<0.05),而cTNT和α-MHC表达仍十分明显。结论脉冲电刺激能抑制大鼠胚胎心脏组织成肌细胞株H9c2细胞的增殖,同时cTNT和α-MHC表达增强,Oct4表达减弱,对H9c2细胞具有诱导分化作用。

Objective To observe the inhibition of pulsed electric stimulation on the proliferation of H9c2 cells and the induction on the differentiation of the cells. Methods We applied a set of pulse electrical stimulations of different frequencies （0, 1, 5, 10, 50, 100 Hz）, different voltage （0, 5, 10, 20, 40, 50 V） and different time （0, 1, 2, 4, 6, 8 h/d） on H9c2 cells in rats. 5-azacytidine was used as an intervention factor control. We detected the proliferation rate of H9c2 cells through M＇IF assay, the expressions of cardiac troponin T （cTNT） and Oct4 through immunofluorescence cytochemical staining, and the expressions of cTNT and （x-MHC mRNA using RT-PCR method. Results Pulsed electric stimulation inhibited the proliferation of H9c2 cells （P〈0.05）. The pulsed electric stimulation of 1 Hz, 4 h/d and 10 V was proved optimized in inhibiting the growth of H9c2 cells to the greatest extent but not inducing the death of cells. Two weeks after the optimized pulsed elctric stimulation on H9c2 cells, the expression of Oct4 was reduced （P 〈 0.05）, while the expressions of cTNT and α-MHC were still strong. Conclusion Pulsed electric stimulation can not only inhibit the reproduction of myoblast cell line H9c2 in heart tissue of rat embryo, but also induce the differentiation of H9c2 cells. During it, the expressions of cTNT and a-MHC are enhanced but Oct4 is weakened.