胚胎干细胞在体外模拟心肌生长环境中向心肌分化  

Differentiation of Embryonic Stem Cells towards Cardiomyocytes in Mimic Cardiac Microenvironment in vitro

作　　者：欧东波[1] 陈瑞[2] 郑强荪[1] 郭菁菁[1] 郭万刚[1] 刘雄涛[1] 

机构地区：[1]第四军医大学唐都医院心内科,陕西西安710038 [2]第四军医大学唐都医院整形烧伤科,陕西西安710038

出　　处：《现代生物医学进展》2009年第4期622-626,共5页Progress in Modern Biomedicine

摘　　要：目的:探讨获得大量成熟同质有功能的心肌细胞(cardiomyocytes,CMs)的方法。方法:胚胎干细胞(embryonic stem cells,ESCs)经悬滴培养法形成拟胚体(embryoid bodies,EBs),将EBs用抗坏血酸处理后与天然心肌细胞共培养诱导ESCs的分化,采用免疫组化、流式细胞技术、RT-PCR、超微分析等方法分析分化细胞(ESC-derived CMs,ESCMs)的结构和功能。结果:共培养组,常规组和对照组中有(90.1±3.40)%,(29.6±6.03)%和(20.1±3.82)%的EBs表现节律性收缩,差异有统计学意义(P<0.05)。在体外模拟的心脏微环境中,不但跳动EBs的数目明显增加,且单个跳动EBs中含有功能CMs的百分比也明显增加。兴奋收缩的自律性能维持更长时间,在后期分化得到的有功能ESCMs在细胞形态上接近于天然的心肌细胞。结论:心脏微环境是促使ESCs特定地分化为CMs的关键因素,抗坏血酸处理和共培养诱导策略可使ESCs定向分化为CMs。Objeetive： To explore a method to obtain a large number of more mature and homogenous cell types from ESCs. Methods： A novel protocol was developed by combining ascorbic acid and co-cultured inducing strategies to control the differentiation of embryonic stem cells （ESCs）, and then evaluated the structural and functional properties of these ESC-derived CMs （ESCMs） by im- munohistochemistry, flow cytometric analysis, RT-PCR, and transmission electron microscopy. Results： The majority （average, 90.1 ＋ 3.40） % of ESCs-derived aggregates （embryoid bodies, EBs） in the co-culture systems exhibited spontaneous contractions, as ESCs cultured alone, or only with ascorbic acid, the average percentage of beating aggregates was （20.1±3.82） % and （29.6±6.03） %. Both the number of beating EBs and the percentage of functional CMs were increased significantly in mimic cardiac microenvironment. The automaticity of beating also maintained for more time and cellular morphology of beating cells at late stage was similar to native CMs. Conclusion： The cardiac microenvironment is the key inducer in directing the site-specific differentiation ofESCs. Ascorbic acid and co-cultured inducing strategies can make ESCs directionally differentiate into Cardiomyocytes（CMs）.

关 键 词：胚胎干细胞 细胞分化 抗坏血酸 心脏微环境 心肌细胞 

分 类 号：Q813[生物学—生物工程] Q132.7