长链非编码RNA-MALAT1调控miR-22/snail轴对胃癌增殖、迁移及侵袭的影响  

Effects of long non-coding RNA MALAT1 on the proliferation,migration,and invasion of gastric cancer through regulating the miR-22/snail axis

作　　者：陈虹宇 廖盛涛 郑茜[1] 邱烈旺[1] 余柯岐 吕琳[1] 梅浙川[1] Chen Hongyu;Liao Shengtao;Zheng Qian;Qiu Liewang;Yu Keqi;Lii Lin;Mei Zhechuan(Department of Gastroenterology,the Second Affiliated Hospital of Chongqing Medical University)

机构地区：[1]重庆医科大学附属第二医院消化内科

出　　处：《重庆医科大学学报》2019年第11期1408-1416,共9页Journal of Chongqing Medical University

摘　　要：目的:探讨长链非编码RNA MALAT1(long non-coding RNA MALAT1,lnc RNA MALAT1)调控miR-22/snail轴对胃癌增殖、迁移及侵袭的影响。方法:采用siRNA干扰技术抑制MALAT1表达,同时过表达miR-22和Snail;采用RT-PCR检测MALAT1,miR-22和snail在胃癌细胞系中的表达水平(n=3);采用Western blot检测NC组、si-MALAT1组中snail蛋白的表达水平(n=3);采用CCK-8法在12、24、48、72 h检测NC组、si-MALAT1组、miR-22组、miR-22+snail组、si-MALAT1+miR-22组中细胞吸光度值(n=3);采用划痕实验检测NC组、si-MALAT1组、miR-22组迁移速度(n=3);采用Transwell检测NC组、siMALAT1组、miR-22组、miR-22+snail组、si-MALAT1+miR-22组穿过膜细胞的个数(n=3);采用双荧光素酶检测NC+pMIRMALAT1-WT组、NC+pMIR-MALAT1-MUT组、miR-22+pMIR-MALAT1-WT组、miR-22+pMIR-MALAT1-MUT、NC+pMIRSnail-WT组、NC+pMIR-Snail-MUT组、miR-22+pMIR-Snail-WT组、miR-22+pMIR-Snail-MUT组的荧光活性(n=3)。两独立样本间的比较采用t检验,多组数据之间的比较采用单因素方差进行分析,涉及时间因素时采用重复测量方差分析。结果:MALAT1在胃癌细胞系中的表达水平明显高于正常胃黏膜上皮细胞(均P=0.000);低表达MALAT1后,AGS和SGC-7901增殖能力(P1=0.001,P2=0.005),迁移能力(P1=0.018,P2=0.007),侵袭能力(P1=0.024,P2=0.015)均降低;同时双荧光素酶结果显示MALAT1能够靶向结合miR-22(P=0.001),miR-22能够靶向结合snail(P=0.001);miR-22在胃癌细胞中的表达水平明显低于正常胃黏膜上皮细胞(均P<0.05);过表达miR-22后,AGS和SGC-7901增殖能力(P1=0.004,P2=0.009),迁移能力(P1=0.034,P2=0.005),侵袭能力(P1=0.037,P2=0.011)均降低;在低表达MALAT1和过表达miR-22后能够更进一步增加低表达MALAT1后对AGS和SGC-7901的增殖(P1=0.022,P2=0.006)和侵袭(P1=0.024,P2=0.015)的抑制作用;同时过表达miR-22和snail后能够反转miR-22对AGS和SGC-7901的增殖(P1=0.003,P2=0.004)和侵袭(P1=0.015,P2=0.01)的抑制作用。结论:MALAT1在胃癌细胞中明显�Objective:To investigate the effects of long non-coding RNA MALAT1 on the proliferation,migration,and invasion of gastric cancer through regulating the miR-22/snail axis.Methods:The expression of MALAT1 was inhibited by siRNA technology,and miR-22 and snail were overexpressed;real-time PCR was used to determine the expression levels of MALAT1,miR-22,and snail in gastric cancer cell lines(n=3);Western blot was used to determine the expression level of snail protein in NC group and si-MALAT1 group(n=3);CCK8 assay was used to determine the cell absorbance values in NC group,si-MALAT1 group,miR-22 group,mi R-22+snail group,and si-MALAT1+miR-22 group at 12 h,24 h,48 h,and 72 h(n=3);wound healing assay was used to determine the migration rate in NC group,si-MALAT1 group,and miR-22 group(n=3);transwell assay was used to determine the number of transmembrane cells in NC group,si-MALAT1 group,miR-22 group,miR-22+snail group,and si-MALAT1+miR-22 group(n=3);dual luciferase assay was used to determine the fluorescence activity in NC+pMIR-MALAT1-WT group,NC+pMIR-MALAT1-MUT group,miR-22+pMIR-MALAT1-WT group,miR-22+pMIR-MALAT1-MUT group,NC+pMIR-Snail-WT group,NC+pMIR-SnailMUT group,miR-22+pMIR-Snail-WT group,and miR-22+pMIR-Snail-MUT group(n=3).The t-test was used for comparison between two independent samples;one-way analysis of variance was used for comparison between multiple sets of data,and repeated measures analysis of variance was used for time-associated factors.Results:The expression level of MALAT1 in gastric cancer cell lines was significantly higher than that in normal gastric mucosal epithelial cells(all P=0.000);after underexpression of MALAT1,AGS and SGC-7901 had significantly decreased proliferative ability,migration ability,and invasion ability(P1=0.001,0.018,and 0.024,respectively;P2=0.005,0.007,and 0.015,respectively);meanwhile,the dual luciferase assay results showed that MALAT1 could targetedly bind to miR-22,and miR-22 could targetedly bind to snail(both P=0.001).The expression level of miR-22 in gastric c

关 键 词：长链非编码RNA MALAT1 miR-22 SNAIL 增殖 迁移 侵袭 

分 类 号：R735.2[医药卫生—肿瘤]