微环境对体外间充质干细胞向心肌样细胞分化的作用

Role of microenvironment on in vitro differentiation of mesenchymal stem cells into cardiomyogenic cells

目的研究心肌组织中何种细胞对骨髓间充质干细胞(MSCs)向心肌样细胞定向分化起决定性作用。方法分离培养骨髓MSCs、心肌细胞(CM)和内皮细胞(EC)。MSCs经BrdU标记后与CM、EC分别共培养和单独培养,通过形态学、免疫细胞化学和双重免疫细胞化学鉴定。结果分离培养的MSCs、CM和EC纯度高于95%。标记MSCs与CM共培养后,细胞具有心肌细胞样形态。4周时,有44%的细胞BrdU与肌动蛋白(sarcomeric actin)或连接蛋白-43(connexin-43)双重免疫细胞化学染色阳性。与EC共培养以及单独培养时,MSCs形态变化不明显,无双染阳性细胞出现。结论CM对骨髓MSCs向心肌样细胞的定向分化具有重要作用。

Objective To find the cells in myocardium controlling in cardiomyogenic differentiation of marrow derived-mesenchymal stem cells （MSCs）. Methods Rat marrow derived-MSCs, cardiomyocytes （CM） and endothelial cells （EC） were isolated and cultured, then MSCs were incubated with the CM or EC or cultured alone after labeled by 5-Bromo-2-deoxyuridine （BrdU） for 72 hours. The cells were identified by morphology, immunocytochemistry and double immunocytochemistry. Results More than 95% of the ceils isolated and cultured belonged to MSCs, CM or EC. Incubated with CM, most MSCs labeled by BrdU showed cardiomyocyte-like morphology. After 4 weeks of cultivation, positive staining of double immunocytochemistry of BrdU and Sarcomeric Actin or Connexin- 43 （CX-43） were observed in 44% cells. However, there was no obvious change in morphology and no positive staining of double immunocytochemistry was recorded in MSCs incubated with EC or cultured alone after 1,2,3 and 4 weeks of cultivation. Conclusion MSCs can differentiate into cardiomyogenic cells in cardiomyocyte microenvironment and cardiomyocytes may play a crucial role in the differentiation.