miR-19b沉默对P19细胞分化及线粒体功能的影响  

Effects of miR-19b knockdown on cardiac differentiation in mouse embryonic carcinoma P19 cells

作　　者：刘雪华[1,2] 朱莎莎[2] 余章斌[2] 朱春[2] 李萌萌[2] 韩树萍[2] 胡晓山[2] 朱金改[2] 彭宇竹[1,2] 

机构地区：[1]南京医科大学,江苏南京210029 [2]南京医科大学附属南京妇幼保健院儿科,江苏南京210004

出　　处：《东南大学学报（医学版）》2015年第5期704-709,共6页Journal of Southeast University(Medical Science Edition)

基　　金：国家自然科学基金资助项目(81070500);南京市卫生青年人才基金资助项目(QRX11107)

摘　　要：目的:通过沉默微小RNA-19b(miR-19b)研究miR-19b对P19细胞分化及线粒体功能的影响。方法:转染miR-19b沉默质粒与空载质粒进入P19细胞建立起稳定的细胞系;线粒体功能分析用以检测细胞凋亡;诱导分化P19细胞,实时定量荧光PCR检测miR-19b沉默后细胞分化关键基因表达水平。结果:Target Scan 5.1软件及双荧光素酶报告基因系统共同证实,Wnt1为miR-19b在P19细胞中发挥作用的靶点(P<0.05);测定第0、4、6、8、10、12天c Tn T、NKX2.5、GATA4等基因的核酸表达量显示,心肌分化相关标志基因逐渐增高,miR-19b沉默组明显低于对照组(P<0.05);于分化第10天检测线粒体功能,发现miR-19b沉默组线粒体DNA拷贝数的相对含量高于对照组,其活性氧水平显著低于对照组,ATP水平也高于对照组(均P<0.05)。结论:miR-19b沉默可抑制细胞凋亡,抑制心肌细胞分化。Objective：To explore the function of microRNA-19b（miR-19b） knockdown on differentiation in P19 cells.Methods：We transfect miRNA-19b knockdown plasmid and vector into P19 cells and stable cell lines were selected by puromycin and conformed by dual luciferase reporter gene system.Mitochondria function assays were adopted to analyze apoptosis respectively.Real time PCR detected P19 cell differentiation markers.Results：TargetScan 5.1 software and dual luciferase reporter gene system confirmed that Wnt1 was the target gene of miR-19b（P〈0.05）;Expressions of all the marker genes gradually grew during the process of differentiation;the genes showed significantly lower degrees of expression in miR-19b-knockdow cells.Compared with the vector group, mtDNA copy number was significantly higher in the miR-19b knockdown group, so as to ATP levels.ROS levels in miR-19b-knockdown cells were much lower than those in control cells （ all P 〈0.05）.Conclusion： miR-19b knockdown could inhibit apoptosis and differentiation in P19 cells.

关 键 词：微小RNA-19b沉默 P19细胞 线粒体 分化 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]