摘要
目的:利用经典的悬滴培养法形成拟胚体(EBs),诱导小鼠胚胎干细胞(m ESCs)在体外分化为心肌细胞,观察维生素C作为诱导剂对m ESCs体外诱导分化为心肌细胞的影响,进而寻找一种高效、安全的诱导方法。方法:复苏m ESCs,传代培养后,利用悬滴法使m ESCs形成拟胚体,用含0.1mg/m L维生素C的分化培养基对其进行诱导分化,以不添加任何诱导剂作为对照组,观察出现搏动拟胚体的时间,搏动拟胚体的数量,计算分化比率,统计搏动频率,并进行免疫荧光染色测定心肌肌钙蛋白T(c Tn T)的表达。结果:小鼠胚胎干细胞可自发分化为心肌细胞,但效率较低,0.1mg/m L的维生素C能明显提高m ESCs分化为心肌细胞的效率,约83.3%的EBs出现搏动,显著高于未添加任何诱导剂组,搏动频率为(81.2±7.8)次/分钟。两组搏动心肌细胞c Tn T免疫染色阳性。结论:维生素C能够显著提高m ESCs向心肌细胞分化效率,应用维生素C体外诱导m ESCs向心肌细胞分化是一种较为理想的体外诱导方式。
Objective： Using the hanging drop method to induce differentiation of mouse embryonic stem cells into cardiomyocytes in vitro and investigate the effect of ascorbic acid on the cardiomyocytes differ- entiation from mouse embryonic stem cells and find a high efficient and safety method. Methods： mESCs were recovered, passaged and form embryoid bodies using the hanging drop method. The embryoid bodies were in- duced by differentiation medium that containing ascorbic acid （0. 1mg/mL）. Control group did not add any inducer. Observing the time of appearing beating cardiomyocytes and counting the percentage of beating colo- nies and the beating frequency of cardiomyocytes; investigating the effect of ascorbic acid on the cardiomyo- cytes differentiation from mouse embryonic stem ceils. Results： Beating area was found in 83.3 % of embry- oid bodies when using differentiation medium that containing ascorbic acid （0. 1 mg/mL） , significantly greater than control group. Beating cardiomyocytes of two groups were positive to cTnT staining. Conclusion ： Ascor- bic acid can markedly improve the efficiency of mESCs differentiation of cardiomyocytes, and special cardiac proteins cTnT were detected. It is an ideal method using the ascorbic acid to induce mESCs differentiate into cardiomyocytes in vitro.
出处
《心肺血管病杂志》
CAS
2015年第10期774-777,共4页
Journal of Cardiovascular and Pulmonary Diseases
基金
北京市朝阳区科委社会发展可持续发展与城乡一体化促进计划项目基金资助(SF1417)
首都医科大学学生科研创新项目(XSKY2015192)
关键词
维生素C
小鼠胚胎干细胞
心肌细胞
细胞分化
Ascorbic acid
Mouse embryonic stem cells
Cardiomyocyte
Induced differentiation
