[1]朱静,冯川,李莉,等.干扰组蛋白乙酰化引发间充质干细胞心肌定向分化特定蛋白的异常表达[J].第三军医大学学报,2008,30(04):303-306.
 ZHU Jing,FENG Chuan,LI Li,et al.ShRNA on histone acetylation induces unusual cardiac-specific protein expression in MSCs differentiation into cardiomyocytes[J].J Third Mil Med Univ,2008,30(04):303-306.
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干扰组蛋白乙酰化引发间充质干细胞心肌定向分化特定蛋白的异常表达(/HTML )
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《第三军医大学学报》[ISSN:1000-5404/CN:51-1095/R]

卷:
30卷
期数:
2008年第04期
页码:
303-306
栏目:
论著
出版日期:
2008-02-28

文章信息/Info

Title:
ShRNA on histone acetylation induces unusual cardiac-specific protein expression in MSCs differentiation into cardiomyocytes
作者:
朱静冯川李莉邓兵田杰张晓萍
重庆医科大学:儿科研究所,儿童医院心血管内科
Author(s):
ZHU Jing FENG Chuan LI Li DENG Bing TIAN Jie ZHANG Xiao-ping
Pediatric Research Institute, Department of Cardiology, Children’s Hospital, Chongqing Medical University
关键词:
乙酰化修饰RNA干扰质粒间充质干细胞分化
Keywords:
acetylation RNA interference plasmid mesenchymal stem cells differentiation
分类号:
R329.2;R341;R394.3
文献标志码:
A
摘要:
目的 利用组蛋白乙酰化修饰关键因子Gcn5 shRNA重组质粒转染经5-氮杂胞苷(5-azacytidine,5-aza)诱导后的间充质干细胞(mesenchymal stem cells,MSCs)并检测心肌早期发育相关结构和功能蛋白表达情况,探讨组蛋白乙酰化修饰在MSCs定向分化过程中的调控作用。  方法 ①以组蛋白乙酰化修饰关键因子Gcn5为靶点,用带绿色荧光蛋白(green fluorescence protein,GFP)基因的pGenSil-1作为载体,构建Gcn5 shRNA质粒。②MSCs体外扩增,5-aza诱导后,不同的时间点用脂质体包载Gcn5 shRNA共转染大鼠MSCs,24 h后相差显微镜下观察细胞生长状况,电镜下观察染色质的形态变化,采用Western blot法分别检测Gcn5、心肌结构重链蛋白(myosin heavy chain,MHC)和连接蛋白(Connexin 43, Cx43)的表达状况。  结果 电镜示经通用阴性质粒HK转染的MSCs细胞,形态与正常MSCs无明显差别,染色质仍以边集状态为主;经干扰质粒转染的MSCs细胞在形态上与正常MSCs细胞无明显差别,但其染色质出现明显的浓缩紧密现象。以β-actin为内参,比较干扰组和阴性对照组Gcn5、Cx43、MHC蛋白,计算出沉默效率分别为86%、58%和67%。  结论 转染Gcn5 shRNA质粒可干扰组蛋白乙酰化平衡,进而影响MSCs体外分化中心肌发育特定蛋白的表达。
Abstract:
Objective To construct recombinant plasmid that can inhibit acetyltransferase Gcn5 expression through RNA interference, then transfect it into MSCs induced by 5-azacytidine (5-aza), and analyze the role of acetylases in the process of MSCs differentiation.   Methods MSCs were isolated from bilateral thigh bones and tibias of Wistar rats, then treated with 5-aza. Gcn5 shRNA recombinant plasmid vector was transfected into MSCs via lipofectin. The expressions of Gcn5, myosin heavy chain (MHC) and Connexin 43 (Cx43) were investigated by Western blot.   Results Under electron microscope, MSCs transfected with Gcn5 shRNA recombinant plasmid displayed obvious heterochromatin, but MSCs and MSCs transfected HK plasmid showed euchromatin. The expressions of Gcn5, Cx43 and MHC decreased significantly after Gcn5 shRNA recombinant plasmid transfection. Taking β-actin as inner criterion, the silent ratio of Gcn5, MHC and Cx43 were 86%, 58% and 67%, respectively.   Conclusion The Gcn5 shRNA plasmids we constructed block acetylation during the process of MSCs differentiation into cardiomyocytes. Acetylation plays a very important role in the regulation of MSCs differentiation.

参考文献/References:

朱静, 冯川, 李莉,等. 干扰组蛋白乙酰化引发间充质干细胞心肌定向分化特定蛋白的异常表达[J].第三军医大学学报,2008,30(4):303-306.

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更新日期/Last Update: 2008-04-29