摘要
先前的研究表明,miR-150-5p发挥抑癌基因的作用,调控肿瘤细胞的侵袭与转移。然而,关于其在乳腺癌细胞侵袭与转移中的机制尚不明确。本实验旨在研究miR-150-5p负向调控Rab1A在乳腺癌细胞上皮-间质转化(epithelial-mesenchymal transition,EMT)中的作用。双荧光素酶的结果显示,miR-150-5p可负向调控Rab1A。荧光定量PCR(qRT-PCR)结果显示,miR-150-5p在乳腺癌细胞MCF-7及MDA-MB-231(MDA-231)中的表达水平明显低于正常乳腺上皮细胞MCF-10A;在MDA-231中过表达miR-150-5p后,qRT-PCR结果显示,Rab1A mRNA的表达水平明显降低。Western印迹结果显示,过表达miR-150-5p后,miR-150-5p组细胞中的Rab1A、波形蛋白(vimentin)及N-钙黏着蛋白(N-cadherin)的表达水平相对于对照组(NC)细胞明显降低,而E-钙黏着蛋白(Ecadherin)的表达水平明显增加。Transwell侵袭和划痕实验显示,与miR-150-5p+Con组细胞相比,miR-150-5p+Rab1A组细胞的侵袭和迁移能力明显增加。qRT-PCR结果显示,miR-150-5p+Rab1A组细胞的Rab1A mRNA表达水平明显增加。Western印迹结果显示,miR-150-5p+Rab1A组细胞中的波形蛋白、N-钙黏着蛋白表达水平明显增加,而E-钙黏着蛋白表达明显降低,过表达Rab1A后显著逆转了miR-150-5p对EMT的影响。综上所述,miR-150-5p可以通过负向调控Rab1A抑制EMT,进而抑制乳腺癌细胞的侵袭和迁移。
Previous studies have showed that miR-150-5p acts as a tumor suppressor gene and regulates the invasion and metastasis of tumor cells. However,its mechanism in the invasion and migration of breast cancer cells is unclear. The purpose of this study was to investigate the role of miR-150-5p in the negative regulation of Rab1A in epithelial-mesenchymal transition（ EMT） of breast cancer cells. The results of double luciferase assay showed that miR-150-5p could negatively regulate Rab1A. The resultsof fluorescence quantitative real-time PCR（ qRT-PCR） showed that the expression of miR-150-5p in breast cancer cells MCF-7 and MDA-MB-231（ MDA-231） was significantly lower than that in normal breast epithelial cells MCF-10 A. The result of qRT-PCR after overexpressing miR-150-5p in MDA-231 cells showed that the expression level of Rab1A mRNA was significantly increased. Western blotting showed that the expression level of Rab1A,Vimentin and N-cadherin were significantly decreased and the expression of E-cadherin was significantly increased in miR-150-5p-overexpression group cells compared with the control group cells. Transwell invasion and scratch assays revealed that the capacity of invasion and migration were obviously increased in miR-150-5p ＋ Rab1A group cells compared with miR-150-5p＋ Con group cells. qRT-PCR revealed that the expression of Rab1A mRNA was prominently increased in miR-150-5p ＋ Rab1A group cells. Western blotting indicated that the expression levels of vimentin and N-cadherin were up-regulated in miR-150-5p ＋ Rab1A group cells,but the expression of E-cadherin was down-regulated. Overexpressed Rab1A notably reversed the effects of miR-150-5p in EMT. In summary,these results illustrated that miR-150-5p suppresses EMT by targeting Rab1A,thus inhibiting the invasion and migration of breast cancer cells.
作者
李平
任斌
李洪利
尹崇高
LI Ping;REN Bin;LI Hong-Li;YIN Chong-Gao(Department of Pathology,Weifang Nursing Vocational College;Department of Surgery,Clinical College of Weifang Medical University;Medical Research Center,Weifang Medical University;College of Nursing,Weifang Medical University Weifang 261053,Shandong,China)
出处
《中国生物化学与分子生物学报》
CAS
CSCD
北大核心
2018年第9期997-1003,共7页
Chinese Journal of Biochemistry and Molecular Biology
基金
国家自然科学基金青年基金项目(No.81702932
No.81641111)
山东省自然科学基金(No.ZR2015HL065)~~
