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绿色荧光蛋白标记的小鼠胚胎干细胞系的建立及向心肌细胞分化(PDF)

《心脏杂志》[ISSN:1009-7236/CN:61-1268/R]

期数:
2006年第4期
页码:
392-395
栏目:
基础研究
出版日期:
2006-08-25

文章信息/Info

Title:
Establishment of murine embryonic stem cell line carrying green fluorescent protein and its differentiation into cardiomyocytes
作者:
罗浩; 江洪; 蔡军; 曾彬
武汉大学人民医院心内科，湖北 武汉 430060
Author(s):
LUO Hao;  JIANG Hong;  CAI Jun;  ZENG Bin
Department of Cardiology, Renmin Hospital, Wuhan University, Wuhan，Hubei 430060, China
关键词:
胚胎干细胞; 绿色荧光蛋白; 转染; 分化; 心肌细胞
Keywords:
embryonic stem cells;  green fluorescence protein;  transfection;  differentiation;  cardiomyocytes
分类号:
Q254
DOI:
-
文献标识码:
A
摘要:
目的 建立能够稳定表达绿色荧光蛋白 (green fluorescent protein，GFP)的小鼠胚胎干细胞系，并诱导其向心肌细胞分化。方法 质粒pEGFPN1脂质体复合体转染小鼠胚胎干细胞，经G418筛选后选取GFP强阳性克隆进行扩增建系。对稳定表达GFP的胚胎干细胞系进行畸胎瘤形成检测，观察其多向分化潜能。诱导GFP阳性胚胎干细胞向心肌细胞分化。免疫荧光及RTPCR检测心肌细胞特异性标志物。结果 转染后胚胎干细胞经20次传代后仍然表达GFP，裸鼠皮下接种胚胎干细胞后30~50 d均可形成畸胎瘤。GFP阳性胚胎干细胞成功向心肌细胞分化。结论 成功建立表达GFP的小鼠胚胎干细胞系，并可诱导其向心肌细胞分化。
Abstract:
AIM To establish a murine embryonic stem (ES) cell line carrying green fluorescent protein (GFP) and differentiate it into cardiomyocytes. METHODS Plasmid pEGFPN1 liposome complex was introduced into the murine ES cells and the transfected clones were filtered in the presence of G418. Strong fluorescent GFP clones were singly picked out and further proliferated. Teratoma formation was performed to analyze the differentiation status and potential of the GFP ES cells. GFP ES cells differentiated into cardiomyocytes. Immunocytochemistry staining and RTPCR indicated that the cells derived from ES cells were cardiomyocytes based on the presence of cardiacspecific molecular marker. RESULTS One line of GFP ES cells subcultured twenty passages and still carried the GFP DNA. GFP positive ES cells were injected subcutaneously into the nude mice and tumors formed after about 30-50 days. The stable GFP ES cell line successfully differentiated into cardiomyocytes in vitro. CONCLUSION A murine ES cell line carrying GFP is established and the GFP positive ES cell line can differentiate into cardiomyocytes.

参考文献/References

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［2］ Doetschman TC, Eistetter H, Katz M, et al. The in vitro development of blastocyst derived embryonic stem cell lines: formation of visceral yolk sac, blood island myocardium［J］. J Embryol Exp Morphol, 1985, 87: 27-45.

［3］Caspi O,Gepstein L. Potential applications of human embryonic stem cellderived cardiomyocytes［J］. Ann N Y Acad Sci, 2004, 1015: 285-298.

［4］ Keirstead HS, Nistor G, Bernal G, et al. Human embryonic stem cellderived oligodendrocyte progenitor cell transplants remyelinate and restore locomotion after spinal cord injury［J］. J Neurosci, 2005, 25(19): 4694-4705.

［5］ Maltsev VA, Wobus AM, Rohwedel J, et al. Cardiomyocytes differentiated in vitro from embryonic stem cells developmentally express cardiacspecific genes and ionic currents［J］. Circ Res, 1994, 75(2): 233-244.

［6］ Rolletschek A, Chang H, Guan K, et al. Differentiation of embryonic stem cellderived dopaminergic neurons is enhanced by survival promoting factors［J］. Mech Dev, 2001, 105(1-2): 93-104.

［7］ Boheler KR, Czyz J, Tweedie D, et al. Differentiation of pluripotent embryonic stem cells into cardiomyocytes［J］. Circ Res, 2002, 91(3): 189-201.

备注/Memo

备注/Memo:
收稿日期：2005-08-01.通讯作者：江洪, 教授, 主要从事介入性心脏病学研究 Tel:(027)88041911 作者简介：罗浩， 硕士生 Tel：(027)88328605 Email：mouse9801@163.com
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