心肌微环境对骨髓间充质干细胞的诱导分化作用

目的通过与心肌细胞(CMs)共同培养和采用含有CMs裂解液的培养基两种方法体外模拟心肌微环境,探讨心肌微环境对骨髓间充质干细胞(MSCs)分化的诱导作用。方法自新生乳鼠的心脏分离CMs,自成年大鼠的骨髓分离MSCs,将MSCs与CMs按1∶4的比例共同培养1周,观察细胞形态的改变,并通过免疫荧光方法检测共培养后MSCs表达心脏特异性肌钙蛋白T(cTnT)及CD31的情况;将分离的CMs反复冻融制成CMs裂解液,将MSCs在含有4倍CMs裂解液的培养基中培养1周,观察细胞形态的改变,并通过免疫化学方法检测MSCs表达心脏特异性肌钙蛋白T(cTnT)及CD31的情况;以仅用普通培养基所培养的MSCs作为对照。结果与CMs共培养的MSCs和CMs裂解液培养的MSCs逐渐伸展变长,形成肌细胞形态,培养1周后经抗cTnT和抗CD31免疫染色均呈阳性;对照组MSCs没有明显形态变化,抗cTnT和抗CD31免疫染色成阴性。结论与CMs共培养和采用含有CMs裂解液的培养基两种方法均可在体外模拟心肌微环境,诱导MSCs向心肌样细胞和内皮样细胞方向分化。

Effect of myocardial microenvironment on the differentiation of bone marrow mesenchymal stem cells

Objective In this study, we try to understand the effects of microenvironment on the differentiation of mesenchymal stem cells (MSCs) by coculturing MSCs with mature cardiomyocytes or culturing MSCs in cardiomyocyte-lysate, in this study. Methods MSCs isolated from mature rats were either cocultured with cardiomyocytes isolated from new born rats with the ratio of 1 to 4, or cultured in the medium containing 4-fold cardiomyocyte-lysate obtained by repeated freezing and defrosting of rat myocardial cells. The morphology of MSCs under light microscopy were observed daily for 7 days and immunostaining against cTnT and CD31 was performed on the 7~ th day. MSCs cultured in ordinary medium were observed as the control. Results Both MSCs cocultured with cardiomyocytes and cultured in cardiomyocyte-lysate were differentiated into myogenic cells and expressed cTnT and CD31 at the 7th day of cultivation. The MSCs in the control group did not change in morphology and express cTnT or CD31. Conclusion Both myocardial cell coculturing system and cardiomyocyte-lysate system can be used to induce bone marrow MSCs to differentiate into cardiomyocyte-like cells and endotheliocyte-like cells.