参麦体外诱导人胚胎生殖细胞向心肌细胞分化

摘要

Objective: To evaluate the feasibility of shenmai on myocardiac differentiation of human embryonic germ cells (hEGC) and establish a method of in vitro differentiation of hEGC towards cardiomyocytes. Methods: hEGC were obstained through tissue culturing. Five days after suspension, embryoid bodies formed from hEGC were seeded in 24-well plates and incubated with differentiation medium containing shenmai in various concentration. Immunocytochemical staining was used to identify the expression of primordial myocardium-specific transcription factor GATA-4 and cardiac troponin T (cTnT). Results:The maximum cardiac differention rate of hEGC reached at a concentration of 1 g/L shenmai, while the positive cells were found in 57. 0%±3. 25% after three weeks. There was siginificant difference compared with the control. After shenmai induction, hEGC became fusiform shape, the cell processes affiliated with each other and the cells formed cell lines. Just after 3 days, GATA-4 appeared weak positive and was up to the maximum after 3 weeks. The expression of cTnT was observed 2 weeks after induction with shenmai, and it increased gradually in 3 weeks and 4 weeks after induction. Conclusion: Shenmai can enhance the differentiation of hEGC into cardiomyocytes, so we can establish a method of in vitro differentiation of hEGC towards cardiomyocytes.%目的:探讨参麦对人胚胎生殖细胞(hEGC)向心肌细胞诱导分化的作用.方法:取5～10周人胚胎生殖腺嵴,进行组织块体外培养,用直接悬浮法使hEGC形成拟胚体(EBs),用不同浓度参麦的培养基对其进行诱导分化,然后取不同时间的细胞做免疫细胞化学显色,鉴定细胞的心肌特异转录因子GATA-4和心肌肌钙蛋白-T(cTnT)表达情况.结果:参麦诱导hEGC分化为心肌细胞的最佳浓度为1g/L,诱导第3周时分化率达57 0%±3 25%,显著高于不添加任何诱导剂的对照组.诱导后细胞形态变成梭形,3周细胞突起相互连接成条索状,且排列方向趋于一致;诱导后第3天即开始出现GATA-4弱表达,第3周时表达最强;诱导后2周,细胞内开始表达cTnT,3周强阳性表达,4周表达明显增强.结论:参麦能够促进hEGC向心肌细胞分化,从而得以建立一种体外诱导hEGC分化为心肌细胞的方法.