外源性miR-34a抑制H1299细胞生长的机制研究  

Cell Growth Inhibition by Exogenous miR-34a in H1299 Cell

作　　者：段广新[1] 段卫明[2] 许雅香[1] 张燕娟[3] 周新文[3] 

机构地区：[1]苏州大学医学部基础医学与生物科学学院,苏州215123 [2]苏州大学附属第一医院,苏州215123 [3]苏州大学医学部放射医学与防护学院,苏州215123

出　　处：《中国细胞生物学学报》2013年第5期637-642,共6页Chinese Journal of Cell Biology

基　　金：国家自然科学基金(批准号:81071878)资助的课题~~

摘　　要：将合成的miR-34a成熟序列转染p53–/–的非小细胞肺癌H1299细胞,探究外源性的miR-34a是否影响p53缺失细胞的生长、凋亡和衰老以及其作用机制。MTT检测细胞的生长与存活,藻红B染色检测细胞的死亡,Annexin V/PI染色检测细胞凋亡,β-半乳糖苷酶染色试剂盒检测细胞的衰老,Western blot检测与细胞凋亡和周期相关的Bcl-2、Puma、Cdk4和E2F3蛋白的表达。结果显示,miR-34a转染组和阴性对照组相比,存活率明显降低,且48 h较24 h更显著,24 h和48 h存活率分别为79.94%、64.83%;细胞死亡和凋亡的分析结果表明,外源性miR-34a可以促进细胞死亡和凋亡;此外,细胞还出现了明显的衰老,并且检测到凋亡、衰老相关的Bcl-2、Puma、E2F3和Cdk4蛋白的表达下调。miR-34a可以下调p53下游相应靶蛋白的表达,通过部分补救p53通路或p53非依赖途径促进细胞的凋亡、衰老并抑制细胞的增殖。To elucidate the function and mechanism of exogenous miR-34a involved in growth, apopto- sis and senescence in the p53-/- H1299, miR-34a mimics was used to transfect into H1299. Viablity cell rate was analyzed by MTT. Erythrosine B cells staining was used to count the rate of cell death. Apoptosis was detected by Annexin V/PI staining. Senescence was observed by senescence associated 13-galactosidase kit staining. The expres- sion of the Bcl-2, Puma, E2F3 and Cdk4 was resolved by Western blot. The group transfection with miR-34a has a lower cell viablity rate than the group of negative control. The decrease of cell viablity of 48 h is more notable than 24 h. The cell viablity rate of 24 h and 48 h are 79.94%, 64.83%, respectively. The rate of cell death and apoptosis is higher and the senescence staining more marked when transfected with miR-34a. The expression of Bcl-2, E2F3 and Cdk4 significantly deci＇eases and Puma increases when induction ofmiR-34a. So miR-34a can decrease it＇s tar- get proteins of p53 downstream and induce apoptosis and cell death, senescence, inhibit cell growth of H1299 cell by partly saving the p53 pathway and p53 independent pathway.

关 键 词：MIR 34a 凋亡 衰老 E2F3 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]