miR-145靶向Six1对胃癌侵袭的影响及分子机制  

Effect of miR-145 targeting Six1 on invasion of gastric cancer and its molecular mechanism

作　　者：蒋丽[1] 李辉[1] 张桂英[2] 穆仪冰[1] 刘海涛 Jiang Li;Li Hui;Zhang Guiying;Mu Yibing;Liu Haitao(Department of Gastroenterology,Fourth Hospital of Changsha of Hunan Province,Changsha 410006,China)

机构地区：[1]长沙市第四医院消化内科,410006 [2]中南大学湘雅医院消化内科

出　　处：《国际肿瘤学杂志》2018年第8期470-477,共8页Journal of International Oncology

基　　金：湖南省卫生计生委科研计划（B2016221）

摘　　要：目的探讨微小RNA-145（mill-145）、Six1在胃癌组织及胃癌细胞株中的表达，miR-145靶向Six1对人胃癌细胞株侵袭的影响及分子机制。方法选取2017年1月至11月长沙市第四医院经胃镜＋病理组织学确诊的进展期胃癌患者60例，采用实时荧光定量PCIR（qRT—PCR）检测miR-145mRNA及Six1在60例胃癌及配对癌旁组织中的表达，并分析其相关性。在胃癌细胞株MKN-45、BGC．823和SGC-7901中转染miR-145模拟物，qRT-PCR法检测3种胃癌细胞株中miR-145mRNA相对水平，选取miR-145mRNA表达水平最高的SGC-7901胃癌细胞株行后续实验。设模拟物组（转染mill-145模拟物）、NC组（转染miR-145阴性对照）和空载体组（不加入任何寡聚核苷酸），采用Transwell实验检测胃癌细胞侵袭能力；小管形成实验验证促血管形成能力；Westernblotting法检测胃癌细胞株SGC-7901下游蛋白Six1、上皮钙黏着蛋白（E-cadherin）和波形蛋白（vimentin）表达水平。MirTrap系统及荧光素酶报告实验验证miR-145是否靶向Six1基因。结果胃癌组织及癌旁组织中miR-145mRNA分别为0．579±0．086、1．009±0．121，差异有统计学意义（t=-22．498，P〈0．001）；Six1分别为2．516±0．208、1．041±0．227，差异有统计学意义（t=37．119，P〈0．001）；miR-145mRNA与Six1表达呈负相关（r=-0．728，P〈0．001）。过表达miR-145后，模拟物组、NC组和空载体组中，胃癌细胞穿膜细胞数分别为48．550±3．716、82．800±3．797、87．467±8．023，差异有统计学意义（F=87．789，P〈0．001），模拟物组与Nc组及空载体组相比差异均有统计学意义（均P〈0．001）。模拟物组、NC组和空载体组中，人脐静脉内皮细胞的成管数分别为24．333±1．211、34．167±2．041、36．500±3．209，差异有统计学意义（F=47．103，P〈0．001），模拟物组与Nc组及空载体组相比差异均有统计学意义（均P〈0．001）。模拟物组�Objective To investigate the expressions of microRNA-145 （miR-145） and Six1 in gastric cancer tissues and gastric cancer cell lines, and to investigate the effect of miR-145 targeting Six1 on invasion of human gastric cancer cells and its molecular mechanism. Methods Sixty patients with advanced gastric cancer confirmed by gastroscopy and pathology in the Fourth Hospital of Changsha were selected, from January to November, 2017. The expressions of miR-145 mRNA and Six1 in 60 cases of gastric cancer and their matched adjacent tissues were detected by real-time fluorescence quantitative PCR （qRT-PCR） and their correlations were analyzed, miR-145 mimics were transfected into MKN-45, BGC-823 and SGC-7901 gastric cancer cell lines. The relative levels of miR-145 mRNA in three gastric cancer cell lines were detected by qRT-PCR. The SGC-7901 gastric cancer cell lines with the highest expression level of miR-145 mRNA were selected for subsequent experiments. The mimic group （transfected with miR-145 mimic）, NC group （transfected with miR-145 negative control） and empty carrier group （no addition of any oligonucleotides） were set up. The invasive ability of gastric cancer cells was detected by Transwell test. The ability of angiogenesis was verified by microtu- bule formation experiment. The expression levels of downstream proteins of the gastric cancer cell lines SGC-7901 such as Six1, E-cadherin and vimentin were detected by Western blotting method. The MirTrap system and the luciferase report test verified whether miR-145 targeted the Six1 gene. Results The expressions of miR-145 mRNA in gastric cancer tissues and in para cancerous tissues were 0. 579 ± 0. 086 and 1. 009 ±0. 121, respectively. The difference was statistically significant （ t = - 22. 498, P 〈 0. 001 ）. The expressions of Six1 were 2. 516 .± 0.208 and 1. 041 ± 0. 227, respectively. The difference was statistically significant （t = 37.119, P 〈 0.001 ）. The expressions of miR-145 mRNA and Six1 were negatively correlated

关 键 词：胃肿瘤 微RNAS 肿瘤侵润 Six1 

分 类 号：R735.2[医药卫生—肿瘤]