miR-199b-3p对肾癌细胞增殖、凋亡、侵袭的影响及其机制探讨  

Effects of miR-199b-3p on proliferation,apoptosis,and invasion of renal carcinoma cells

作　　者：刘晨溪 赵红娟[1] 田伟[1] 刘超 徐洋涛[1] 潘岩[2] 尤校雷[3] 周洪月[4] LIU Chenxi;ZHAO Hongjuan;TIAN Wei;LIU Chao;XU Yangtao;PAN Yan;YOU Xiaolei;ZHOU Hongyue(Urology Surgery,Baoding Second Central Hospital,Baoding 072750,China;不详)

机构地区：[1]保定市第二中心医院泌尿外科,河北保定072750 [2]保定市第二中心医院肿瘤科 [3]邯郸市中心医院泌尿外科 [4]河北大学附属医院泌尿外科

出　　处：《山东医药》2022年第4期26-30,35,共6页Shandong Medical Journal

基　　金：河北省医学科学研究课题计划项目(20210072)。

摘　　要：目的探讨微小RNA-199b-3p(miR-199b-3p)对肾癌细胞增殖、凋亡、侵袭的影响,并分析其调控机制。方法应用RT-qPCR检测人正常肾小管上皮细胞株HK-2细胞、肾癌细胞株786-O中的miR-199b-3p。取肾癌细胞株786-O,分别转染孔质粒、miR-199b-3p mimics建立miR-NC组和miR-199b-3p mimics组,分别应用CCK-8增殖实验、克隆形成实验、细胞流式实验、Transwell实验观察各组细胞增殖、凋亡、侵袭能力。应用生物信息学方法分析miR-199b-3p可能作用的靶基因。取肾癌细胞株786-O,分别转染孔质粒、miR-199b-3p mimics(10 nmol/L)、miR-199b-3p mimics(50 nmol/L)、成纤维细胞生长因子2(FGF2)-siRNA,建立miR-NC组、miR-199b-3p mimics 10 nmol/L组、miR-199b-3p mimics 50 nmol/L组、FGF2-siRNA组,应用Western blotting法检测FGF2及PI3K/AKT信号通路关键分子表达蛋白。结果786-O细胞中miR-199b-3p表达低于HK-2细胞(P<0.05)。CCK-8增殖实验、克隆形成实验验显示miR-199b-3p mimics组细胞增殖能力低于miR-NC组(P均<0.05)。细胞流式实验显示,miR-199b-3p mimics组细胞凋亡率高于miR-NC组(P<0.05)。Transwell实验结果显示,miR-199b-3p mimics组786-O细胞侵袭能力低于miR-NC组(P<0.05)。TargetScan7.2软件在线预测显示FGF23′非编码区与miR-199b-3p具有结合位点。Western boltting法实验结果显示,miR-NC组、miR-199b-3p mimics 10 nmol/L组、miR-199b-3p mimics 50 nmol/L组FGF2蛋白表达分别为0.87±0.11、0.42±0.06、0.23±0.04,三组两两比较P均<0.05。miR-199b-3p mimics组、FGF2-siRNA组细胞中FGF2、p-AKT、p-ERK蛋白表达低于miR-NC组,AKT、ERK蛋白表达高于miR-NC组(P均<0.05)。结论miR-199b-3p在肾癌细胞中低表达,可以抑制肾癌细胞增殖、侵袭并促进其凋亡,其作用机制可能与miR-199b-3p负靶向调控FGF2、PI3K/AKT信号通路有关。Objective To investigate the effects of microRNA-199b-3p(miR-199b-3p)on proliferation,apoptosis,and invasion of renal carcinoma cells and to analyze its regulation mechanism.Methods The miR-199b-3p in the human normal renal tubular epithelial cell line HK-2 and renal carcinoma cell line 786-O was detected by RT qPCR.Renal carcinoma cell line 786-O was transfected with porogen and miR-199b-3p mimics to establish the miR NC group and miR-199b-3p mimics group.CCK-8 proliferation experiment,clone formation experiment,cell flow cytometry,and Transwell experiment were used to observe the cell proliferation,apoptosis,and invasion abilities of cells in each group.Bioinformatics methods were used to analyze the possible target genes of miR-199b-3p.Renal carcinoma cell line 786-O was transfected with porogen,10 nmol/L miR-199b-3p mimics,50 nmol/L miR-199b-3p mimics and fibroblast growth factor 2(FGF2)-siRNA,respectively,which were then taken as the miR-NC group,10 nmol/L miR-199b-3p mimics group,50 nmol/L miR-199b-3p mimics group,and FGF2-siRNA group.The protein of key molecules of FGF2 and PI3K/AKT signaling pathway was detected by Western blotting.Results The expression of miR-199b-3p in 786-O cells was significantly lower than that in HK-2 cells(P<0.05).CCK-8 proliferation test and clone formation assay showed that the proliferation of 786-O cells in miR-199b-3p mimics group was significantly lower than that in the miR-NC group(P<0.05).The apoptosis of 786-O cells in the miR-199b-3p mimics group was significantly higher than that in the miR-NC group(P<0.05).Transwell assay showed that the invasive ability of 786-O cells in the miR-199b-3p mimics group was significantly lower than that in the miR-NC group(P<0.05).Targetscan 7.2 software online prediction showed that FGF23′URT had binding sites with miR-199b-3p.Western blotting showed that the expression levels of FGF2 protein in the miR-NC group,10 nmol/L miR-199b-3p mimics group,and 50 nmol/L miR-199b-3p mimics group were 0.87±0.11,0.42±0.06,and 0.23±0.04,respectiv

关 键 词：微小RNA-199b-3p 肾癌 细胞增殖 细胞凋亡 细胞侵袭能力 成纤维细胞生长因子2 PI3K/AKT信号通路 

分 类 号：R737.11[医药卫生—肿瘤]