中国组织工程研究Issue(19)：3069-3074,6.DOI:10.3969/j.issn.2095-4344
钠钙交换体启动子促进诱导多能干细胞向心肌细胞转化
Sodium-calcium exchanger promoter promotes differentiation of mouse induced pluripotent stem cells into cardiomyocytesin vitro
摘要
Abstract
BACKGROUND:Inducing pluripotent stem cels has been considered as a promising treatment for ischemic heart disease. However, an ideal inducing method has not been found yet. <br> OBJECTIVE:To investigate the role of sodium-calcium exchanger (NCX1) promoter in the differentiation of mouse induced pluriptent stem cels (miPS) into cardiomyocytes. <br> METHODS: The pLVX-IRES-ZsGreen1 vectors which contain NCX1 promoter constructed by recombinant DNA technology were co-transfected to 293FT cels with ViraPowerTM Lentiviral Packaging Mix. The recombinant lentiviruses infected with miPS were selected and purified by puromycin. miPS were recovered and passaged to form embryoid bodies. The embryoid bodies were induced by differentiation medium containing various concentrations of the virus titer. The number of beating embryoid bodies were calculated. The expression profiles of the myocardial intra-markers were tested to determine the differentiation efficiency of iPSC by RT-PCR and immunofluorescence analysis. <br> RESULTS AND CONCLUSION:pLVX-IRES-ZsGreen1 vectors which contain NCX1 promoter were constructed and confirmed by PCR. Virus could be packaged, purified and concentrated successfuly. The recombinant lentivirus to transduce miPS was sorted by flow cytometry. In contrast to NCX1-/GFP- cels, NCX1+/GFP+ cels were differentiated and developed prominent beating areas with sustained contractile activity for additional 4 days, and demonstrated positive expression of gap communication marker CX43 and cardiac troponin. The expressions of GATA4, MEF2c and Nkx2.5 in the NCX1+ cels were 4.2, 7.5, and 2.5 times those in NCX1- cels. Results showed the NCX1 promoter can promote the cardiac differentiation of miPS .关键词
干细胞/诱导/小鼠诱导多能干细胞/钠钙交换体启动子/慢病毒/国家自然科学基金Key words
stem cels/multipotent stem cels/mice/sodium-calcium exchanger/lentivirus分类
医药卫生引用本文复制引用
戴波,Wang Yi-gang,李洪涛,刘泽,肖定璋,余细勇,陈斌,何建新,向定成,邱健..钠钙交换体启动子促进诱导多能干细胞向心肌细胞转化[J].中国组织工程研究,2014,(19):3069-3074,6.基金项目
国家自然科学基金面上项目(81270189)；广东省自然科学基金博士启动项目(S2011040000852) （81270189）
