四种诱导剂诱导骨髓间充质干细胞向心肌样细胞的分化

背景：不同诱导剂诱导骨髓间充质干细胞分化形成心肌样细胞功能学方面的研究未见报道。目的：观察5-氮杂胞苷、血管紧张素Ⅱ、骨形态发生蛋白2和P53特异性抑制剂对骨髓间充质干细胞向心肌样细胞分化的影响。方法：分离大鼠骨髓间充质干细胞,取第3代细胞分为5组,即正常对照组,5-氮杂胞苷组、血管紧张素Ⅱ组、骨形态发生蛋白2组和P53特异性抑制剂组。采用流式细胞仪技术鉴定骨髓间充质干细胞表面特定抗原表达情况,Western blot法检测诱导4周后Cx43、肌钙蛋白Ⅰ表达情况,fluo3/AM钙离子探针激光共聚焦检测诱导4周后钙瞬变能力。结果与结论：骨髓间充质干细胞表面抗原CD29、CD44、CD45的阳性表达率分别为89.1%,90.4%和1.9%。Western blot结果显示,各诱导组均有肌钙蛋白Ⅰ的表达,P53特异性抑制剂组较其余各组表达多（P〈0.05）,骨形态发生蛋白2组较其余各组表达少（P〈0.05）。Cx43蛋白在各组之间均有表达,正常对照组较各实验组明显较少（P〈0.05）,P53特异性抑制剂组较其余各组表达多（P〈0.05）,血管紧张素Ⅱ组较其余各组表达少（P〈0.05）。钙瞬变功能测定结果显示,荧光强度血管紧张素Ⅱ〉P53特异性抑制剂〉5氮杂胞苷〉骨形态发生蛋白2〉正常对照组（P〈0.01）。提示不同诱导剂诱导骨髓间充质干细胞之后,各实验组均能向心肌样细胞分化,表达心肌特异性蛋白。而不同诱导剂发挥作用的通路不同,从而对诱导骨髓间充质干细胞分化形成心肌样细胞的功能产生不同影响。

Induced differentiation of bone marrow mesenchymal stem cells into cardiomyocyte-like cells by four inductors

BACKGROUND：It has been rarely reported that different inductors are used to induce bone marrow mesenchymal stem cells to differentiate into cardiomyocyte-like cells.OBJECTIVE：To investigate the effects of 5-azacytidine,angiotensin Ⅱ,bone morphogenetic protein 2,pifithrin-α on induced differentiation of bone marrow mesenchymal stem cells into cardiomyocyte-like cells.METHODS：Passage 3 rat bone marrow mesenchymal stem cells were divided into five groups：control,5-azacytidine,angiotensin Ⅱ,bone morphogenetic protein 2,pifithrin-α.Surface antigen expression of bone marrow mesenchymal stem cells was identified by flow cytometry.After 4 weeks of induction,Cx43 and troponin Ⅰ expression was detected by western blot method and calcium transient capability of induced bone marrow mesenchymal stem cells was detected by laser confocal scanning microscopy with flou3/AM.RESULTS AND CONCLUSION：Cell surface antigen CD29,CD44,CD45 positive rate was 89.1%,90.4%,1.9% respectively.Troponin Ⅰ expression appeared in each group and the expression was significantly higher in the pifithrin-α group（P 0.05）,as well as significantly lower in the bone morphogenetic protein 2 group（P 0.05）,compared with the remaining groups.Cx 43 expression appeared in each group,and the expression was significantly lower in the control group（P 0.05） compared with the remaining groups.In the four experimental groups,Cx43 expression was highest in the pifithrin-α group and lowest in the angiotensin Ⅱ group（P 0.05）.Calcium transient capability determined by fluorescence intensity was arranged as follows：angiotensin Ⅱ group pifithrin-α group 5-azacytidine group bone morphogenetic protein 2 group control group（P 0.01）.These findings suggest that after induced by 5-azacytidine,angiotensin Ⅱ,bone morphogenetic protein 2,and pifithrin-α,bone marrow mesenchymal stem cells differentiated towards cardiomyocyte-like cells and expressed myocardium-specific protein.Different inductors exert different effects on differentiation of bone marrow mesenchymal stem cells into cardiomyocyte-like cells which occurs because of different pathways.