miR-let-7c-5p通过靶向HMGA2抑制膀胱癌细胞侵袭和迁移  

miR-let-7c-5p inhibits invasion and migration of bladder cancer cells by targeting HMGA2

作　　者：姚越 张冲 熊言骏 韩兵 高幸福 汪盛[1] YAO Yue;ZHANG Chong;XIONG Yanjun;HAN Bing;GAO Xingfu;WANG Sheng(Department of Urology,First Affiliated Hospital of Bengbu Medical College,Anhui 233000,China)

机构地区：[1]蚌埠医学院第一附属医院泌尿外科,安徽蚌埠233000

出　　处：《南方医科大学学报》2021年第7期1022-1029,共8页Journal of Southern Medical University

基　　金：安徽省自然科学基金(1808085MH293);蚌埠医学院2020科研创新计划项目(Byycx20030)。

摘　　要：目的探讨miR-let-7c-5p能否调控HMGA2抑制膀胱癌细胞侵袭和迁移。方法生物信息学方法确定miR-let-7c-5p的关键基因;RT-qPCR和Western blot检测膀胱癌和癌旁组织中miR-let-7c-5p mRNA和HMGA2蛋白相对表达量。以人正常膀胱细胞SV-HUC-1作为对照,RT-qPCR和Western blot检测膀胱癌细胞系T24,UM-UC-3,5637细胞中miR-let-7c-5p mRNA和HMGA2蛋白的相对表达量。对UM-UC-3细胞中miR-let-7c-5p分别进行上调和下调,上调实验设模拟物阴性对照组(mimic NC)、miR-let-7c-5p模拟物组(mimicmiR-let-7c-5p)、下调实验设阴性对照组(inhibitor NC)及miR-let-7c-5p抑制剂组(si-miRlet-7c-5p),双荧光素酶实验、RT-qPCR和Western blot法共同验证miR-let-7c-5p和HMGA2的靶向关系;Transwell小室检测单独上调或下调miR-let-7c-5p表达及同时下调miR-let-7c-5p和HMGA2表达UM-UC-3细胞侵袭和迁移的变化;Western blot检测上皮间质转化(EMT)相关蛋白(E-cadherin,N-cadherin,vimentin,Snail)相对表达量。结果HMGA2为miR-let-7c-5p的靶基因之一;与癌旁组织相比,膀胱癌组织中miR-let-7c-5pmRNA相对表达量降低(P<0.05),HMGA2蛋白相对表达量增加(P<0.05);与SV-HUC-1细胞相比,UM-UC-3细胞中miR-let-7c-5p相对表达量显著降低,HMGA2显著增加(P=0.01)。上调miRlet-7c-5p表达,UM-UC-3细胞侵袭和迁移能力均显著下降(P<0.05);下调miR-let-7c-5p表达,UM-UC-3细胞侵袭和迁移能力均显著增加(P<0.05);当同时下调miR-let-7c-5p和HMGA2表达UM-UC-3细胞侵袭和迁移能力显著下降(P<0.05)。Western blot结果显示,上调miR-let-7c-5p表达,E-cadherin表达增加,N-cadherin,vimentin,Snail蛋白表达下降;下调miR-let-7c-5p表达,Ecadherin表达下降,N-cadherin,vimentin,Snail蛋白表达增加;同时下调miR-let-7c-5p和HMGA2表达能够抑制UM-UC-3细胞EMT(P<0.05)。结论miR-let-7c-5p通过靶向HMGA2抑制EMT进而抑制UM-UC-3细胞侵袭和迁移。Objective To investigate whether miR-let-7 c-5 p inhibits invasion and migration of bladder cancer cells by regulating HMGA2.Methods We used bioinformatics methods to determine the key genes of miR-let-7 c-5 p.RT-qPCR and Western blotting were used to detect the expressions of miR-let-7 c-5 p mRNA and HMGA2 protein in bladder cancer and adjacent tissues.With human normal bladder SV-HUC-1 cells as the control,we detected the expression levels of miR-let-7 c-5 p mRNA and HMGA2 protein in bladder cancer cell lines T24,UM-UC-3 and 5637 with RT-qPCR and Western blotting.We observed the effects of miR-let-7 c-5 p upregulation(by transfection with a miR-let-7 c-5 p mimic),miR-let-7 c-5 p downregulation(using a miR-let-7 c-5 p inhibitor),and knockdown of both HMGA2 and miR-let-7 c-5 p on invasion,migration and expressions of epithelial-mesenchymal transition(EMT)-related proteins(E-cadherin,N-cadherin,vimentin,and Snail)in UM-UC-3 cells.Dual luciferase assay,RT-qPCR and Western blotting were used to verify the targeting relationship between miR-let-7 c-5 p and HMGA2.Results HMGA2 was identified as one of the target genes of miR-let-7 c-5 p.Compared with the adjacent tissues,bladder cancer tissues showed a significantly decreased expression of miR-let-7 c-5 p and an increased expression of HMGA2 protein(P<0.05).In UM-UC-3 cells,the expression of miR-let-7 c-5 p was significantly reduced and that of HMGA2 was significantly increased as compared with those in SV-HUC-1 cells(P=0.001).Up-regulating miR-let-7 c-5 p expression significantly lowered the invasion and migration abilities of UM-UC-3 cells,and down-regulating miR-let-7 c-5 p expression obviously promoted the invasion and migration of UM-UC-3 cells(P<0.05).Knockdown of both miR-let-7 c-5 p and HMGA2 expression significantly lowered the invasion and migration(P<0.05)and inhibited the expressions of EMT-related proteins of UM-UC-3 cells(P<0.05).Conclusion miR-let-7 c-5 p inhibits EMT of bladder cancer UM-UC-3 cells by targeting HMGA2,thereby inhibiting the cell invasio

关 键 词：膀胱癌 miR-let-7c-5p HMGA2 上皮间质转化 侵袭迁移 

分 类 号：R737.14[医药卫生—肿瘤]