微小核糖核酸592调控神经胶质瘤细胞株U251细胞凋亡  

miR-592 induces apoptotic cell death in human U251 cells

作　　者：陈照亮[1] 刘红[1] 杨会利[1] 高玉凯[1] 张娟[1] 吉文玉[2] 

机构地区：[1]滨州市中心医院肿瘤科,滨州251700 [2]新疆医科大学第一附属医院儿外一科

出　　处：《中国神经精神疾病杂志》2017年第4期234-238,共5页Chinese Journal of Nervous and Mental Diseases

摘　　要：目的研究微小核糖核酸592(miR-592)对神经胶质瘤细胞株U251凋亡的影响。方法首先通过定量聚合酶链反应分析miR-592在神经胶质瘤组织和癌旁组织中的表达变化;随后向U251细胞转染miR-592的拟合物,并通过流式细胞技术分析miR-592过表达对U251细胞凋亡的影响;通过生物信息学分析,找到miR-592的潜在靶分子,并通过荧光素酶双报告实验以及蛋白免疫印迹法等进行验证;进一步,转染U251细胞Runx2的下调si RNA,绘制细胞的生长曲线,并对U251细胞的凋亡进行分析。结果定量PCR结果分析发现,miR-592在肿瘤组织中明显低表达(t=2.752,P=0.013);miR-592过表达能明显抑制U251细胞的生长,与对照组比较,培养36 h、48 h后存在统计学差异(t=2.127,P=0.031;t=2.284,P=0.026);流式细胞分析结果显示,miR-592显著促进U251细胞凋亡:对照组晚期凋亡率为7.2%±0.68%,而转染miR-592组晚期凋亡率为17.47%±1.45%,存在统计学差异(t=3.294,P=0.007);荧光素酶双报告实验以及蛋白免疫印迹法实验结果发现miR-592直接靶向Runx2的3’-UTR来抑制Runx2蛋白的水平;检测下调Runx2对U251细胞生长的影响,结果显示转染了Runx2 si RNA的细胞生长明显比对照组低(t=3.124,P=0.011),流式细胞技术对周期的检测显示,Runx2下调表达上调U251细胞的凋亡率。通过绘制肿瘤生长曲线,发现miR-592过表达明显抑制肿瘤的生长。同时,Runx2的下调表达也明显抑制肿瘤的生长。结论 miR-592通过直接靶向Runx2来诱导神经胶质瘤细胞凋亡,进而抑制细胞的生长。Objective To investigate the role of miR-592 in the Gliorna. Methods We first analyzed the expression of rniR-592 in Glioma tissues from patients by quantitative PCR. We transfeeted U251 cells with rniR-592 mimics and then detected the growth of ceils by MTr assay. We performed dual-lueiferase reporter assay and western blot assay to examine whether Runx2 was the direct target of miR-592 in U251 cells. In order to test whether Runx2 was the func- tional target of miR-592, we determined the cell growth curve by down-regulating the level of Runx2. Moreover, we al- so detected the apoptosis of U251 after Runx2 knockdown. Results The expression of miR-592 was significantly re- duced in gliorna tissues （t=2.752,P=0.013）. Over-expression miR-592 remarkably increased the apoptotic rate of U251 cells compared with the control group （t=2.127,P=0.031; t=2.284,P=0.026）. Flow cytornetry analysis showed that MiR- 592 significantly promoted apoptotic cell death of U251 cells Apoptosis rate was 7.2%＋0.68% in rniR-592 group and 17.47%±1.45% in control group （t=3.294,P=0.007）. The results of double lueiferase assay and Western blot assay showed that miR-592 directly targeted the 3 ＂Runx2 of -UTR to inhibit the level of Runx2 protein. The effect of down- regulation of Runx2 on the growth of U251 cells was detected, the results showed that growth was significantly slower in the cells transfected with Runx2 siRNA than in those without Runx2 siRNA （t=3.124,P=0.011）. Detection of cycle by flow cytometry showed that mnx2 down-regulated the apoptosis rate of U251 cells. Tumor growth curve showed that overexpression of miR-592 significantly inhibited tumor growth and the down regulation of Runx2 expresssion also sig- nificantly inhibited tumor growth. Conclusion miR-592 suppresses the growth and promotes the apoptotic rate of U251 ceils by targeting Runx2.

关 键 词：神经胶质瘤 U251 miR-592 凋亡 RUNX2 

分 类 号：R739.41[医药卫生—肿瘤]