多能干细胞来源的心脏祖细胞的分离及功能特性研究
Isolation and functional characterization of pluripotent stem cell-derived cardiac progenitor cells.
作者信息
Huang Xiaojing, Wu Sean M
机构信息
Cardiovascular Research Center, Division of Cardiology, Massachusetts General Hospital, Boston, Massachusetts, USA.
出版信息
Curr Protoc Stem Cell Biol. 2010 Sep;Chapter 1:Unit 1F.10. doi: 10.1002/9780470151808.sc01f10s14.
Abstract
The use of transgenic markers in pluripotent stem cells allows the facile isolation of transient cell populations that appear at certain phases of embryonic development. Here, we describe a procedure for deriving cardiac progenitors from mouse pluripotent stem cells carrying a GFP reporter under the control of an Nkx2.5 enhancer sequence. The cells are propagated under standard conditions and are differentiated using the hanging-droplet method with medium optimized for commitment to the cardiac lineage. Cardiac progenitors are isolated from the differentiation culture using fluorescence-activated cell sorting (FACS) and can be cultured further for functional characterization and experimentation. The protocols described here can be applied to both embryonic and induced pluripotent stem cells and can easily be adapted to transgenic lines carrying other cardiac cell lineage reporters.
摘要
在多能干细胞中使用转基因标记物能够轻松分离出在胚胎发育特定阶段出现的瞬时细胞群体。在此，我们描述了一种从携带在Nkx2.5增强子序列控制下的绿色荧光蛋白（GFP）报告基因的小鼠多能干细胞中获得心脏祖细胞的方法。细胞在标准条件下进行传代培养，并使用悬滴法进行分化，所用培养基针对向心脏谱系的定向分化进行了优化。利用荧光激活细胞分选（FACS）从分化培养物中分离出心脏祖细胞，并可进一步培养以进行功能表征和实验。这里描述的方案可应用于胚胎干细胞和诱导多能干细胞，并且可以很容易地适用于携带其他心脏细胞谱系报告基因的转基因品系。
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