成人脂肪间充质干细胞体外定向诱导分化为心肌样细胞的实验观察_医学论文.docVIP

成人脂肪间充质干细胞体外定向诱导分化为心肌样细胞的实验观察_医学论文 成人脂肪间充质干细胞体外定向诱导分化为心肌样细胞的实验观察_医学论文 作者：张卫泽，陈跃武，哈小琴，陈永清，秦勉，马凌，郭建巍，洪志斌 【关键词】 成人脂肪 【Abstract】 AIM: To explore the methods of isolating and culturing mesenchymal stem cells from adult adipose tissue （ADMSCs） and inducing them to differentiate into cardiomyocytes in vitro, so as to establish a new ideal source of cell transplantation for myocardial regeneration. METHODS: MSCs were isolated and cultured from adult adipose tissue. 5azacytidine (5aza) induced those cells to differentiate into cardiomyocytes. Then the immunocytochemical method and the RTPCR method were used to identify cardiomyocytes after 14, 21, 28 d of additional culture respectively. RESULTS: The morphology of MSCs induced for 14 d was like cardiomyocytes under a phasecontrast microscope. Cardiac troponinI, desmin and cardiac βmyosin heavy chain gene were positively expressed 28 d later by immunocytochemistry and RTPCR. CONCLUSION: Adult adipose tissue contains plenty of mesenchymal stem cells which can be easily isolated and cultured, and then induced to differentiate into cardiomyocytes by 5aza in vitro. ADMSCs might be a new ideal source of cell transplantation for myocardial regeneration. 【Keywords】 mesenchymal stem cell；adipose tissue；cell differentiation；cardiomyocytes 【摘要】 目的：探索成人脂肪组织来源的间充质干细胞（ADMSCs）体外分离、培养方法，并化学诱导使其向心肌细胞分化，为心肌再生医学提供理想的种子细胞来源. 方法：体外分离成人ADMSCs并进行传代培养，5氮胞苷（5aza）诱导其向心肌细胞分化. 分别于诱导后14，21，28 d时用免疫细胞化学染色、RTPCR法进行心肌细胞鉴定. 结果： ADMSCs经5aza诱导后14 d时细胞形态趋向于心肌细胞，诱导28 d免疫细胞化学显示心肌特异性肌钙蛋白I（cTNI）、结蛋白（Desmin）表达阳性，RTPCR检测心肌β肌球蛋白重链（Cardiac βMHC）基因表达阳性. 结论：成人脂肪组织存在丰富的间充质干细胞且易于分离扩增，体外5aza诱导可使其向心肌细胞分化，可做为心肌再生医学的理想种子细胞. 【关键词】 间充质干细胞；脂肪组织；细胞分化；心肌细胞 0引言 近年有关干细胞移植技术的研究进展对临床治疗心肌终末期疾病带来了前所未有的希望. 国外已有动物实验及小规模临床试验显示自体骨髓间充质干细胞（bone marrow mesenchymal stem cells，BMMSCs）移植入坏死心肌后可改善心功能［1-2］. 但由于BMMSCs来源有限，其广泛临床应用受到限制. 新近研究发现，同样来源于中胚层的脂肪组织中亦含有一种多向分化潜能干细胞，这种细胞群更易分离，细胞数量多，可以发挥替代BMMSCs的作用［3-6］. 本实验我们通过体外分离脂肪组织源性间充质干细胞(adiposederived mesenchymal stem cells，ADMSCs)并传代扩增，在体外特定条件下定向诱导