Apelin13对人胚胎干细胞定向心肌细胞分化效率的影响

发布时间：2018-03-11 14:35

本文选题：人胚胎干细胞　切入点：Apelin　出处：《山东医药》2017年33期 　论文类型：期刊论文

【摘要】：目的探讨Apelin13对人胚胎干细胞(hESCs)定向心肌细胞分化效率的影响。方法将hESCs单层培养后经EDTA消化传代,应用明确的诱导分化培养基对观察组(加Apelin13)和对照组(不加Apelin13)定向诱导hESCs向心肌细胞分化。于诱导第2天(中胚层阶段)、第4天(心肌分化初始阶段)、第7天(心肌分化末阶段)作为时间节点,采用实时荧光定量PCR检测两组Brachury T、Mesp1、NKx2.5、APJ mRNA表达水平,采用Western blotting检测两组Brachury T、Mesp1、NKx2.5、APJ、Nanog、Sox2蛋白表达;同时于诱导第7天采用细胞免疫荧光技术检测两组心肌细胞标志物肌钙蛋白T2(TNNT2)、α-辅肌蛋白(α-actinin)表达。结果诱导第7天观察组hESCs定向心肌分化程度高于对照组,免疫荧光染色倒置显微镜下可清楚看到成熟心肌细胞的肌结及心肌细胞TNNT2、α-actinin表达。在3个诱导时间节点,观察组Brachury T、Mesp1、NKx2.5、APJ mRNA及其蛋白表达均高于对照组(P均0.05),观察组hESCs标志蛋白Nanog、Sox2低于对照组(P均0.05)。结论 Apelin13可提高hESCs向心肌细胞的分化效率。
[Abstract]:Objective to investigate the effect of Apelin13 on the differentiation efficiency of human embryonic stem cells (hESCs) oriented cardiomyocytes. Methods hESCs monolayer was cultured and subcultured with EDTA. HESCs was induced to differentiate into cardiomyocytes in the observation group (plus Apelin 13) and the control group (without Apelin 13). On the second day of induction (mesoderm stage, day 4, myocardial differentiation initial stage, day 7), the differentiation of cardiomyocytes was induced on the specific differentiation medium of the observation group (plus Apelin 13) and the control group (without Apelin 13). End stage of myocardial differentiation) as a time node, Real time fluorescence quantitative PCR was used to detect the expression of Brachury Mesp1 nkx2.5APJ mRNA, and Western blotting was used to detect the expression of Brachury Tet Mesp1 NKx2.5APG Sox2 protein. At the same time, on the 7th day after induction, the expression of troponin T _ 2 and 伪 -actinin (伪 -actinin) was detected by cellular immunofluorescence technique. Results on the 7th day of induction, the degree of hESCs directional myocardial differentiation in the observation group was higher than that in the control group. The myocyte node and the expression of TNNT2, 伪 -actinin in the mature cardiomyocytes were clearly observed by immunofluorescence staining under inverted microscope, and the expression of TNNT2, 伪 -actinin was observed at three induced time nodes. The expression of mRNA and its protein in Brachury Tet Mesp1 NKx2.5 Brachury in the observation group was higher than that in the control group (P 0.05), and the hESCs marker protein Nanogan Sox2 in the observation group was lower than that in the control group (P 0.05). Conclusion Apelin13 can increase the differentiation efficiency of hESCs into cardiomyocytes.
【作者单位】：安徽医科大学海军临床学院;中国人民解放军海军总医院;
【基金】：国家自然科学基金资助项目(81370238) 北京市自然科学基金资助项目(7142156)
【分类号】：R54

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