小鼠胚胎干细胞来源的心肌细胞系的建立  

Establishment of a cardiomyocyte line from mouse embryonic stem cell-derived embryoid bodies

作　　者：秦洁[1,2] 薛兆英[2] 郭新[1] 桂耀庭[1] 余振东[1] 蔡志明[1] 

机构地区：[1]北京大学深圳医院男性生殖与遗传广东省重点实验室深圳北京大学香港科技大学医学中心,广东深圳518036 [2]北京大学第一医院,北京100034

出　　处：《基础医学与临床》2009年第9期916-919,共4页Basic and Clinical Medicine

基　　金：广东省自然科学基金(7300931;7008952);广东省医学科研基金(B2007158)

摘　　要：目的建立模拟细胞微环境、促进细胞分化的三维培养体系,以体外诱导和培养小鼠胚胎干细胞（mES-D3）来源的心肌细胞。方法将mES细胞通过悬浮培养获得拟胚体（EBs）后转入Matrigel^TM Matrix3D培养体系中培养。用形态学观察计数收缩集落的数量和频率;用免疫组织化学和RT-PCR检测心肌细胞分化相关基因的表达。结果在MatrigelTM中培养5 d左右,可观察到自发节律收缩的EBs,频率约为40~50次/min;继续培养3 d左右,频率约为80-90次/min;到45 d左右大部分集落最终收缩。分化为心肌细胞所表达的特异性蛋白cTnT、Desmin和Actin表达呈阳性,并表达心肌特异性基因cTnT、NKX2E、GATA-4和MLC-2v。结论mES细胞来源的EBs在MatrigelTM培养体系中能够分化为心肌细胞,籍此我们建立了一株细胞系。Objective To establishment a defined three-dimensional culture system of cardiomyocytes from mouse embryonic stem cell-derived embryoid bodies （EBs）. Methods The EBs were derived from mES by suspending culture and were plated to MatrigelTM Matrix three-dimensional system. The mouse fibroblast cells served as negative control to identify genes expression of contracting EBs. Results The first beating was irregular after 5 days of culture, but became consistent after extra 3 days with at an average frequency of 80 - 100 beats per minute, mES expressed cardiac specific protein of cTnT, Desmin and Actin. The expression of genes associated cardiomyocytes differentiation as cTnT, NKX2E, GATA-4 and MLC-2v were detected by RT-PCR. Conclusion By using the MatrigelTM three-dimensional system, mES-derived EBs may spontaneously differentiate to cardiomyocytes effectively, and possess necessary biological characters of cardiomyocytes.

关 键 词：小鼠胚胎干细胞(mES) 拟胚体(EBs) 分化 心肌细胞 

分 类 号：R541[医药卫生—心血管疾病]