仿生电刺激在心肌细胞诱导胚胎干细胞向心肌样细胞分化中的作用

Effect of bionic electrical stimulation on the differentiation of embryonic stem cells into cardiomyocytes in the presence myocardial cells in vitro

目的探讨仿生电刺激对离体心肌微环境诱导胚胎干细胞(ESCs)向心肌样细胞分化的作用。方法原代分离培养SD大鼠乳鼠心肌细胞及昆明小鼠ESCs,根据是否给予ESCs仿生电刺激及心肌微环境诱导分为5组:对照组、电刺激组、心肌细胞组、电刺激+心肌细胞条件培养液组、电刺激+心肌细胞组。分别于实验第5、7、14天在相差显微镜下观察各组细胞生长情况,实验结束时行肌钙蛋白T(cTnT)免疫荧光染色检测,计算cTnT阳性细胞百分率。结果与其他组相比,电刺激+心肌细胞组ESCs分化而来的心肌样细胞搏动更快,更趋于一致,ESCs诱导分化后的心肌样细胞cTnT蛋白荧光染色阳性率最高,约为40.0%±2.39%,显著高于对照组(2.00%±1.60%)、电刺激组(3.00%±2.00%)、心肌细胞组(28.70%±4.06%)及电刺激+心肌细胞条件培养液组(17.10%±2.23%,P<0.05)。结论模拟心脏跳动的仿生电刺激有助于ESCs在体外心肌微环境下分化为功能完备的心肌样细胞。

Objective To investigate the effects of electrical stimulation on the differentiation of embryonic stem cells（ESCs） into cardiomyocytes in the presence of myocardial cells in vitro.Methods ESCs and neonate rat cardiomyocytes were isolated and cultured.These cells of primary culture were divided into 5 groups according to whether or not electric stimulation was given and the presence of cardiomyocytes： control group,stimulation group,cardiomyocytes group,stimulation＋ cardiomyocyte conditioned medium group,and stimulation＋cardiomyocytes group.Expression of troponin T（cTnT） in the differentiated cells from ESCs was examined by immunofluoresence on the 5th,7th and 14th day.Results In the group co-cultured with myocardial cell and electrical stimulation,the differentiating ratio of cardiomyocytes derived from ESCs and expressing cTnT was 40.00%±2.39%,and it was higher than that in control group（2.00%±1.60%）,stimulation group（3.00%±2.00%）,cardiomyocytes group（28.70%±4.06%）,stimulation＋cardiomyocyte conditioned medium group（17.10%±2.23%,P0.05）.Conclusion Bionic electric stimulation promotes the differentiation of ESCs into cardiomyocyte in a microenvironment consisting of myocardial cells.