微小RNA-181a-3p表达下调对甲状腺癌细胞放射敏感性的影响及其作用机制  

Effect of down-regulated microRNA-181a-3p expression on radiosensitivity of thyroid cancer cells and its action mechanism

作　　者：王红梅 谢斌[2] 陈寒超[1] 关虹 李春芸[1] WANG Hong-mei;XIE Bin;CHEN Han-chao;GUAN Hong;LI Chun-yun(Department of Clinical Laboratory,Haikou Affiliated Hospital of Central South University Xiangya School of Medicine,Haikou 570203,China;Department of Critical Care Medicine,Haikou Affiliated Hospital of Central South University Xiangya School of Medicine,Haikou 570203,China)

机构地区：[1]中南大学湘雅医学院附属海口医院检验科,海南省海口市570203 [2]中南大学湘雅医学院附属海口医院重症医学科,海南省海口市570203

出　　处：《广西医学》2021年第16期1959-1964,共6页Guangxi Medical Journal

基　　金：海南省自然科学基金(817405)。

摘　　要：目的探讨下调微小RNA(miRNA)-181a-3p表达对甲状腺癌细胞放射敏感性的影响,并分析其可能的分子生物学机制。方法(1)体外培养甲状腺癌细胞TPC-1、BCPAP和正常甲状腺上皮细胞Nthyori3-1,并以不同剂量(0、2、4、6、8 Gy)X射线照射TPC-1细胞,采用定量PCR法检测各细胞中miRNA-181a-3p的表达量。(2)将inhibitor control、miRNA-181a-3p inhibitors分别转染TPC-1细胞(anti-miRNA-NC组、anti-miRNA-181a-3p组),另设对照组(细胞中只加入转染试剂),检测不同剂量(0、2、4、6、8 Gy)X射线照射后的细胞活性和经4 Gy X射线照射后的细胞克隆数。(3)应用TargetScan在线工具预测miRNA-181a-3p和丝裂原活化蛋白激酶激酶激酶5(MAP3K5)的靶向关系并采用双荧光素酶报告基因实验验证。(4)将空载质粒、shRNA-MAP3K5、shRNA-MAP3K5对照质粒Vector和miRNA-181a-3p inhibitors、shRNA-MAP3K5和miRNA-181a-3p inhibitors分别转染至TPC-1细胞,设为Scrambled组、shMAP3K5组、Vector+anti-miRNA-181a-3p组和shMAP3K5+anti-miRNA-181a-3p组。检测不同剂量(0、2、4、6、8 Gy)X射线照射后的细胞活性和经4 Gy X射线照射后的细胞克隆数。结果(1)TPC-1、BCPAP细胞中miRNA-181a-3p的表达量高于Nthyori3-1细胞(均P<0.05);不同剂量X射线照射能上调TPC-1细胞中miRNA-181a-3p的表达量,且表达量与照射剂量呈剂量依赖性(均P<0.05)。(2)与对照组和anti-miRNA-NC组比较,经4、6、8 Gy X射线照射的anti-miRNA-181a-3p组细胞活力降低,经4 Gy X射线照射后anti-miRNA-181a-3p组细胞克隆数减少(均P<0.05)。(3)TargetScan在线预测和双荧光素酶报告基因实验结果表明,MAP3K5是miRNA-181a-3p的靶基因。(4)与Vector+anti-miRNA-181a-3p组比较,MAP3K5+anti-miRNA-181a-3p组经4、6、8 Gy X射线照射后的细胞活力和经4 Gy X射线照射后的细胞克隆数均增加(P<0.05)。结论miRNA-181a-3p在甲状腺癌细胞中表达上调,抑制miRNA-181a-3p表达或通过靶向调控MAP3K5或可增强甲状腺癌TPC-1细胞的放Objective To investigate the effect of down-regulated microRNA(miRNA)-181a-3p on the radiosensitivity of thyroid cancer cells,and to analyze the possible molecular biological mechanism.Methods(1)Thyroid cancer cells TPC-1 and BCPAP,and normal thyroid epithelial cells Nthyori3-1 were cultured in vitro,and X-ray radiation with different doses(0,2,4,6 and 8 Gy)was conducted on TPC-1 cells,then the expression of miRNA-181a-3p was detected in all cells by quantitative PCR.(2)Inhibitor control and miRNA-181a-3p inhibitors were transfected into TPC-1 cells of anti-miRNA-NC group and anti-miRNA-181a-3p group,respectively,and control group(added with transfection reagent alone)was established,then the viability of TPC-1 cells after X-ray radiation with different doses(0,2,4,6 and 8 Gy)and the clone formation of TPC-1 cells after X-ray radiation with 4 Gy were detected.(3)The targeting relationship between miRNA-181a-3p and mitogen-activated protein kinase kinase kinase 5(MAP3K5)was predicted by TargetScan online tool and verified by dual-luciferase reporter gene assay.(4)The TPC-1 cells were transfected by empty plasmid,shRNA-MAP3K5,shRNA-MAP3K5 control plasmids Vector and miRNA-181a-3p inhibitors,shRNA-MAP3K5 and miRNA-181a-3p inhibitors,and were defined as Scrambled group,shMAP3K5 group,Vector+anti-miRNA-181a-3p group and shMAP3K5+anti-miRNA-181a-3p group,respectively.The viability of TPC-1 cells after X-ray radiation with different doses(0,2,4,6 and 8 Gy)and the clone formation of TPC-1 cells after X-ray radiation with 4Gy were detected.Results(1)The expression of miRNA-181a-3p in TPC-1 and BCPAP cells was significantly higher than that in Nthyori3-1 cells(all P<0.05);the expression of miRNA-181a-3p in TPC-1 cells was up-regulated by X-ray radiation with different doses in a dose-dependence manner(all P<0.05).(2)By comparison with the control and anti-miRNA-NC groups,the anti-miRNA-181a-3p group yielded decreased cell viability after X-ray radiation with 4,6 and 8 Gy,and reduced clone formation after X-ray radiation wi

关 键 词：甲状腺癌 微小RNA-181a-3p 丝裂原活化蛋白激酶激酶激酶5 放射敏感性 

分 类 号：R736.1[医药卫生—肿瘤]