长链非编码RNA MIR4435-2HG靶向微小RNA-1-3p调控信号通路磷脂酰肌醇激酶/蛋白激酶B对胰腺癌细胞增殖、侵袭和迁移的机制研究  

Mechanism of LncRNA MIR4435-2HG regulated signaling pathway PI3K/Akt on proliferation,invasion and migration of pancreatic cancer cells by regulating miR-1-3p expression

作　　者：倪文[1] 王曦[1] 王奇胜[1] 苏俊 NI Wen;WANG Xi;WANG Qisheng;SU Jun(Hanchuan People's Hospital,Xiaogan,Hubei 431600,China)

机构地区：[1]汉川市人民医院,湖北孝感431600

出　　处：《安徽医药》2021年第3期467-473,共7页Anhui Medical and Pharmaceutical Journal

摘　　要：目的探讨长链非编码RNA MIR4435-2HG(LncRNA MIR4435-2HG)通过调控微小RNA-1-3p(miR-1-3p)的表达对胰腺癌细胞增殖、迁移及侵袭的影响及其可能作用机制。方法体外培养正常胰腺细胞系hTERT-HPNE与胰腺癌细胞系PANC-1、SW1990、PaTu8988、BxPC-3,将胰腺癌PaTu8988细胞按照随机数字表法分为si-MIR4435-2HG组(转染MIR4435-2HG siRNA)、si-con组(转染siRNA Control)、miR-1-3p组(转染miR-1-3p mimics)、miR-con组(转染miR-1-3p阴性对照)、si-MIR4435-2HG+an⁃ti-miR-1-3p组(共转染MIR4435-2HG siRNA与miR-1-3p抑制剂)、si-MIR4435-2HG+anti-miR-con组(共转染MIR4435-2HG siR⁃NA与miR-1-3p抑制剂的阴性对照),同时将未经任何处理的细胞作为NC组。采用实时荧光定量逆转录聚合酶链反应(qRTPCR)检测细胞中MIR4435-2HG与miR-1-3p的表达;双荧光素酶报告基因检测MIR4435-2HG与miR-1-3p的相互作用。四甲基偶氮唑盐微量酶反应比色法(MTT法)分析敲低MIR4435-2HG及上调miR-1-3p表达对胰腺癌细胞增殖的影响;Transwell迁移及侵袭实验检测MIR4435-2HG及上调miR-1-3p表达对胰腺癌细胞迁移及侵袭能力的影响。蛋白质印迹法(Western blotting)检测细胞周期蛋白D1(cyclin D1)、基质金属蛋白酶2(MMP2)、基质金属蛋白酶9(MMP9)及磷脂酰肌醇激酶(PI3K)/蛋白激酶B(Akt)信号通路相关蛋白的表达。结果MIR4435-2HG在胰腺癌细胞中的表达水平明显高于正常胰腺细胞;MIR4435-2HG可特异性结合miR-1-3p并调控其表达活性;敲低MIR4435-2HG与上调miR-1-3p表达后,可抑制胰腺癌PaTu8988细胞增殖,明显减弱细胞迁移及侵袭能力[其中NC组、miR-con组、si-MIR4435-2HG组增殖活性分别为48 h:(1.21±0.10)、(1.18±0.12)、(0.86±0.09),细胞迁移数量分别为(90.56±9.16)、(88.56±8.91)、(26.49±2.10),细胞侵袭数量分别为(160.79±16.12)、(155.09±15.49)、(69.23±7.10),均P<0.05],下调PaTu8988细胞中cyclin D1、MMP2、MMP9、p-Akt、PI3Kp110α、PI3Kp110β的表达;抑制miR-1-3p表达可促�Objective To investigate the effects of long-chain non-coding RNA MIR4435-2HG(LncRNA MIR4435-2HG)on the pro⁃liferation,migration and invasion of pancreatic cancer cells by regulating the expression of microRNA-1-3p(miR-1-3p)and its possible mechanism.Methods The normal pancreatic cell line hTERT-HPNE and pancreatic cancer cell lines PANC-1,SW1990,PaTu8988 and BxPC-3 were cultured in vitro,pancreatic cancer PaTu8988 cells were divided into si-MIR4435-2HG groups(transfection MIR4435-2HG siRNA),si-con group(transfection siRNA Control),miR-1-3p group(transfection miR-1-3p mimics),mir-con group(transfection negative control),si-MIR4435-2HG anti-miR-1-3p group(co-transfection MIR4435-2HG siRNA and miR-1-3),si-MIR4435-2HG anti-miR-con group(co-transfection MIR4435-2HG siRNA negative control with miR-1-3p inhibitors)according to ran⁃dom number table method p inhibitors,while cells without any treatment were taken as the NC group.The expression of MIR4435-2HG and miR-1-3p was detected by real-time fluorescent quantitative PCR.The dual luciferase reporter gene detected the interaction of MIR4435-2HG with miR-1-3p.MTT assay was used to analyze the effect of knockdown of MIR4435-2HG and up-regulation of miR-1-3p expression on proliferation of pancreatic cancer cells.Transwell migration and invasion assays were performed to detect the effects of MIR4435-2HG and up-regulation of miR-1-3p expression on migration and invasion of pancreatic cancer cells.Western blotting was used to detect the expression of cyclin D1,MMP2,MMP9 and phosphatidylinositol kinase(PI3K)/protein kinase B(Akt)signaling pathway-associated proteins.Results The expression level of MIR4435-2HG in pancreatic cancer cells was significantly higher than that of normal pancreatic cells.MIR4435-2HG could bind specifically to miR-1-3p to modulate the expression of miR-1-3p.Knock⁃down of MIR4435-2HG and up-regulation of miR-1-3p expression inhibited the proliferation of pancreatic cancer PaTu8988 cells,sig⁃nificantly attenuated cell migration and invasion[The

关 键 词：胰腺肿瘤 RNA 长链非编码 RNA MIR4435-2HG 微小RNA-1-3p 磷脂酰肌醇激酶/蛋白激酶B信号通路 增殖 迁移 侵袭 

分 类 号：R735.9[医药卫生—肿瘤]