兔骨髓间充质干细胞来源的心肌(样)细胞的诱导分化研究

Differentiation of Marrow Bone Mesenchymal Stem Cell into Cardiomyocytes

目的体外诱导骨髓间充质干细胞(Mesenchymal stem cells,MSCs)向肌源性细胞分化,探索诱导后的MSCs移植于心肌梗死区的存活和分化情况。方法提取、分离、培养兔的MSCs。经5-氮胞苷诱导后,进行免疫组化,电镜观察。4',6二乙酞基-2-苯基吲哚(DAPI)标记MSCs,建立兔心肌梗死模型。实验动物随机分两组:实验组(n=10)在心梗区域注入经诱导后的MSCs;对照组(n=10)在心梗区域注入不含MSCs的培养液。移植4周后,进行病理标本观察和免疫组化检测。结果5-氮胞苷诱导MSCs4周,部分细胞表达肌钙蛋白T(troponin T),电镜观察到肌丝形成。MSCs在体外用DAPI标记,用荧光显微镜观察细胞发蓝色荧光。移植4周后,在实验组中用荧光显微镜观察可见梗死区组织标本中可见DAPI标记带蓝色荧光的供体细胞核,移植细胞表达troponin T。结论MSCs经5-氮胞苷诱导后可向心肌细胞转化。移植细胞可在心肌存活,并向心肌细胞(样)转化。

Objective To induce the differentiation of rabbit bone mesenchymal stem cells（MSCs） into myogenic cells in vitro, and to investigate transplanted MSCs＇ survival and differentiation in myocardiac infarction. Methods MSCs from rabbit were abstracted, isolated and cultured. Then MSCs were induced by 5- azacytidine for 24 hours. After culturing for 4 weeks, MSCs＇ differentiation was studied by irmnunohistochemistry and electron microscopy technique. The model of acute myocardiac infarction was established by ligating left anterodecendant arteries（LAD） of 20 rabbits. These rabbits were randomly divided into two groups： the experiment one（ n = 10） and the control one（ n = 10）. MSCs induced were labeled with 46-dimidino-2-phenylindone （DAPI） and autogenously transplanted into myocardiac infarction in the experiment group. Non-cell cultural liquid was transplanted into myoeardiac infarction in the control group. After transplanting for 4 weeks, MSCs＇ differentiation was studied by immunohistoehemistry. Results Some of MSCs induced by 5-azaeytidine expressed troponin T. Electron microscopy showed myofilaments. Through fluorescent microscope, MSCs laboled with DAPI show blue fluorescence. After 4 weeks of transplantation, the isehemie myoeardium in experiment group can be seen MSCs with blue fluorescence. Donor cells derived from MSCs demonstrate myogenic differentiation with the expression of troponin T. Conclusion MSCs can be induced into eardiomyoeytes by 5-azaeytidine in vitro. MSCs transplanted into myoeardium can survive and show myogenic differentiation.