摘要
目的探讨长链非编码RNA(lncRNA)MYOSLID靶向微小RNA(miR)-339-5p调控肺癌细胞生物学行为的分子机制。方法收集2016年5月至2018年4月于本院接受手术治疗的肺癌组织及癌旁组织标本20例,采用实时荧光定量聚合酶链反应(RT-qPCR)法检测肺癌组织及癌旁组织中MYOSLID与miR-339-5p的表达。体外培养肺癌细胞株A549并分组转染,用噻唑蓝(MTT)、双荧光素酶报告实验验证MYOSLID对miR-339-5p的靶向调控作用;通过细胞转染及分组,流式细胞术检测细胞凋亡,Transwell实验检测细胞迁移及侵袭,双荧光素酶报告基因检测,蛋白质印迹法(Western blot)检测,细胞周期蛋白D1(Cyclin D1),负向调控因子P21(p21),B淋巴细胞瘤2(bcl-2)、bcl-2相关X蛋白(bax)、金属蛋白酶2(MMP)-2、MMP-9蛋白表达用Image J软件分析各条带灰度值。结果抑制lncRNA MYOSLID表达对肺癌A549细胞活力及Cyclin D1蛋白相对表达量,si-MYOSLID组低于si-NC组[1.00±0.09比0.42±0.04,t=17.667,P<0.05;0.74±0.06比0.31±0.03,t=19.230,P<0.05];p21蛋白si-MYOSLID组高于si-NC组[0.28±0.03比0.69±0.06,t=18.335,P<0.05,],差异有统计学意义。抑制lncRNA MY OSLID表达对肺癌A549细胞凋亡率(%)及bax的表达水平,si-MYOSLID组明显高于si-NC组[6.32±0.64比19.48±1.83,t=20.317,P<0.05;0.32±0.03比0.71±0.07,t=15.362,P<0.05];bcl-2蛋白的表达水平,si-MYOSLID组明显高于si-NC组[0.62±0.06比0.23±0.03,t=17.441,P<0.05],差异有统计学意义。抑制lncRNA MYOSLID表达对肺癌A549细胞迁移与侵袭细胞数及MMP-2、MMP-9的表达水平,si-MYOSLID组低于si-NC组[0.22±0.06比0.43±0.01,t=14.441,P<0.05;0.15±0.02比0.37±0.02,t=10.410,P<0.05;0.36±0.02比0.48±0.02,t=16.041,P<0.05],差异有统计学意义。miR-NC组肺癌A549细胞增殖、凋亡率低于miR-339-5p组[(1.00±0.09)%比(2.58±0.25)%,t=17.839,P<0.05;(7.32±0.74)%比(17.45±1.63)%,t=16.041,P<0.05];迁移细胞数及侵袭细胞数,miR-NC组高于miR-339-5p组(98.36±9.54比52.64±5.28,t=12.579,P<0.05;84.33±8.
Objective To investigate the molecular mechanism of long non-coding RNA Myoslid targeting microRNA-339-5p regulating the biological behavior of lung cancer cells.Methods Totally,20 cases of lung cancer tissues and paracancerous tissues that received surgical treatment in our hospital from May 2016 to April 2018 were collected,and the expression of Myoslid and miR-339-5p in lung cancer tissues and paracancerous tissues was detected by real-time fluorescence quantitative polymerase chain reaction(RT-qPCR).The lung cancer cell line A549 cells were cultured in vitro and transfected.The targeted regulation effect of Myoslid on miR-339-5p was verified by thiazolium blue methyl thiazolyl tetrazolium(MTT)and double luciferase report assays.Cell apoptosis was detected by flow cytometry,cell migration and invasion were detected by Transwell assay,double luciferase reporter gene was detected,and protein expression levels of Cyclin D1,B cell lymphoma/leukemia-2(bcl-2),matrix metalloproteinase(MMP)-2,MMP-9,p21 and bcl-2 associated X protein(bax)were detected by Western blotting.Results The activity of A549 cells and the relative expression of Cyclin D1 protein were significantly reduced in si-myoslid group than in si-NC group[1.00±0.09,0.42±0.04,t=17.667,P<0.05;0.74±0.06,0.31±0.03,t=19.230,P<0.05].P21 protein in si-myoslid group was significantly higher than that in si-NC group[0.28±0.03,0.69±0.06,t=18.335,P<0.05].Inhibition of lncRNA MYOSLID expression could significantly increase the apoptosis rate of A549 cells and the expression level of bax,and the apoptosis rate and expression of bax in si-myoslid group were significantly increased as compared with those in si-NC group[6.32±0.64,19.48±1.83,t=20.317,P<0.05;0.32±0.03,0.71±0.07,t=15.362,P<0.05].The expression level of bcl-2 protein in si-myoslid group was significantly higher than that in si-NC group[0.62±0.06,0.23±0.03,t=17.441,P<0.05].Inhibition of lncRNA Myoslid expression had significant effects on the number of migrating and invasive A549 cells as well as
作者
韩宝义
张悦
黄伟坚
黎昆伟
贾杰
胡祖荣
张力为
Han Baoyi;Zhang Yue;Huang Weijian;Li Kunwei;Jia Jie;Hu Zurong;Zhang Weiwei(Surgery Center,Maternal and Child Health Hospital of Guangdong Province,Guangdong Children's Hospital,Guangzhou 511400,China;Department of Thoracic Surgery,the First Affiliated Hospital of Xinjiang Medical University,Urumqi 830000,China)
出处
《中华实验外科杂志》
CAS
北大核心
2021年第8期1417-1422,共6页
Chinese Journal of Experimental Surgery
基金
广东省医学科学技术研究基金项目(B2019110)
广东省软科学研究计划项目(2017A040405033)。
关键词
长链非编码RNA
微小RNA
肺癌
增殖
迁移
侵袭
凋亡
Long non-coding RNA
MicroRNA
Lung cancer
Proliferation
Migration
Invasion
Apoptosis
