肾透明细胞癌中 miR －200c 负性调控 ZEB2基因表达的机制研究

摘要

目的：探讨肾透明细胞癌中 miR －200c 调控 ZEB2基因表达的分子机制。方法：实时定量 PCR 法检测肾透明细胞癌及癌旁组织中 miR －200c 及 ZEB2基因的表达水平。培养肾透明细胞癌 Caki －1细胞，将miR －200c 的前体转染 Caki －1细胞，Western blot 法检测 ZEB2蛋白的表达改变，荧光素酶报告基因表达分析实验验证 miR －200c 与 ZEB2基因的3＇非翻译区（3＇UTR）的结合及调控作用。结果：实时定量 PCR 表达检测显示，与癌旁组织相比，miR －200c 在肾透明细胞癌组织中的表达显著下调，而 ZEB2 mRNA 在肾透明细胞癌组织中的表达则呈现明显上调。Pearson 相关性分析结果表明，在肾透明细胞癌及癌旁组织中 miR －200c 与ZEB2基因的表达均显示负性相关。荧光素酶报告基因表达分析实验明确 miR －200c 能够与 ZEB2基因的3＇UTR 特异性地结合，并抑制荧光素酶的表达。miR －200c 前体能够显著下调 Caki －1细胞中 ZEB2蛋白的表达。结论：miR －200c 与 ZEB2基因的表达异常与肾透明细胞癌相关，在肾透明细胞癌细胞中 miR －200c 能够负性调节其靶基因 ZEB2的表达。%Objective:To explore the molecular mechanism of miR -200c regulating ZEB2 gene in renal clear cell carcinoma.Methods:Real -time PCR was used to detect the expression of miR -200c and ZEB2 mRNA in renal clear cell carcinoma and its adjacent tissues.Western blot assay was used to investigate the ZEB2 protein level and lu-ciferase assay was used to verify the binding ability of miR -200c on ZEB2 after miR -200c precursor transfected in-to Caki -1 cells.Results:Real -time PCR results showed that miR -200c down -regulated in renal clear cell carci-noma tissues compared to its adjacent tissues,and ZEB2 mRNA up -regulated in renal clear cell carcinoma tissues. The Pearson relative analysis showed that miR -200c was negatively related to the ZEB2 gene in renal clear cell car-cinoma and its adjacent tissues.Western blot and luciferase assay revealed that miR -200c could negatively regulate ZEB2 protein expression by binding to the 3'UTR of ZEB2 gene.Conclusion:The abnormal expression of miR -200c and ZEB2 involved in renal clear cell carcinoma and miR -200c could negatively target and silence ZEB2 gene ex-pression in renal clear cell carcinoma cells.