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【目的】体外诱导大鼠骨髓间充质干细胞(MSCs)分化为心肌细胞并探讨其内向整流钾电流(IK1)特征。【方法】采用密度梯度离心法和贴壁培养法分离、纯化大鼠骨髓MSCs,在第3代细胞以5-氮杂胞苷进行诱导,用免疫细胞化学染色和RT-PCR方法鉴定诱导细胞,并采用全细胞膜片钳技术检测IK1表达。【结果】诱导后细胞体积较诱导前增大,呈长梭形。14d后细胞之间出现连接,排列方向渐趋一致。免疫细胞化学染色显示Desmin、α-sarcomericactin和心肌特异性cTnI呈阳性反应,RT-PCR结果表明诱导细胞有心肌β肌球蛋白重链表达。诱导细胞IK1表达呈明显的不均一性,部分细胞IK1与正常心室肌细胞相似,但IK1电流密度总体上低于正常心室肌细胞。【结论】5-氮杂胞苷能在体外诱导MSCs向心肌细胞分化;诱导细胞IK1表达不均一性提示可能有致心律失常潜能。
【Objective】 Rat bone marrow mesenchymal stem cells (MSCs) were induced to differentiate into cardiomyocytes in vitro and their characteristics of inward rectifier potassium current (IK1) were explored. 【Method】 Rat bone marrow MSCs were isolated and purified by density gradient centrifugation and adherent culture. MSCs were induced by 5-azacytidine in the third generation of cells. The induced cells were identified by immunocytochemical staining and RT-PCR. The whole cell patch clamp technique was used to detect IK1 expression. 【Result】 The results showed that the volume of cells after induction increased compared with that before induction and became fusiform. After 14 days, the cells appeared to be connected and the alignment direction became more and more consistent. Immunocytochemical staining showed that Desmin, α-sarcomericactin and myocardial specific cTnI were positive, and RT-PCR results showed that the induced cells had myocardial β-myosin heavy chain expression. The expression of IK1 was obviously inhomogeneous. IK1 was similar to normal ventricular myocytes in some cells, but the current density of IK1 was lower than that of normal ventricular myocytes. 【Conclusion】 5-azacytidine can induce the differentiation of MSCs into cardiomyocytes in vitro. The inhomogeneous expression of IK1 induces the possible arrhythmia potential.