lncRNA H19靶向miR-194对骨肉瘤细胞增殖和侵袭的影响及其作用机制  

Effect of lncRNA H19 targeting miR-194 on proliferation and invasion of osteosarcoma cells and its mechanism

作　　者：李盈义 王勇[2] 邱冠臻 李洪敬[3] LI Yingyi;WANG Yong;QIU Guanzhen;LI Hongjing(Department of Orthopedics,The Affiliated Dalian Fifth People’s Hospital of Dalian Medical University,Dalian 116021,China;Department of Orthopedics,The Affiliated Central Hospital of Shenyang Medical College,Shenyang 110075,China;Department of Joint Surgery,The First Affiliated Hospital of Dalian Medical University,Dalian 116011,China)

机构地区：[1]大连医科大学附属大连市第五人民医院骨外二科,辽宁大连116021 [2]沈阳医学院附属中心医院骨科,沈阳110075 [3]大连医科大学附属第一医院关节外科,辽宁大连116011

出　　处：《中国医科大学学报》2023年第3期212-218,共7页Journal of China Medical University

基　　金：国家自然科学基金(81972522)。

摘　　要：目的探讨长链非编码RNA(lncRNA)H19靶向微RNA 194(miR-194)对骨肉瘤细胞增殖和侵袭的影响及其作用机制。方法收集2018年1月至2019年6月大连医科大学附属大连市第五人民医院骨外二科行骨肉瘤切除术患者的骨肉瘤组织及对应癌旁组织标本各30例。采用实时荧光定量PCR(qRT-PCR)检测30例骨肉瘤组织、对应的癌旁组织以及人正常成骨细胞hFOB1.19和骨肉瘤细胞SAOS-2、U2OS中lncRNA H19和miR-194的表达水平。利用脂质体转染法将lncRNA H19小干扰RNA(si-H19组)、RNA干扰无意义序列(si-NC组)、miR-194模拟物(miR-194 mimics组)、miRNA阴性对照(miR-NC组)分别转染SAOS-2细胞,MTT法检测各组细胞活力,细胞侵袭实验检测细胞侵袭能力,Western blotting检测细胞N-钙黏蛋白(CDH2)和1型胰岛素样生长因子受体(IGF1R)蛋白的表达。生物信息学分析、双荧光素酶报告基因实验和qRT-PCR验证lncRNA H19和miR-194的靶向调控关系。结果与癌旁组织比较,骨肉瘤组织中lncRNA H19的表达水平升高,miR-194的表达水平降低(均P<0.05);与人正常成骨细胞h FOB1.19比较,骨肉瘤细胞SAOS-2、U2OS中lncRNA H19表达水平升高(P<0.05),miR-194表达水平下降(P<0.05)。与si-NC组比较,si-H19组SAOS-2细胞活力降低,侵袭能力降低,CDH2和IGF1R蛋白表达水平降低(均P<0.05);与miR-NC组比较,miR-194 mimics组SAOS-2细胞的活力降低,侵袭能力降低,CDH2和IGF1R蛋白表达水平降低(P<0.05)。生物信息学分析及双荧光素酶报告基因实验结果显示,lncRNA H19可以靶向调控miR-194。干扰miR-194的表达逆转了抑制lncRNA H19的表达对骨肉瘤SAOS-2细胞增殖和侵袭的影响。结论lncRNA H19在骨肉瘤组织和细胞系中的表达升高,miR-194在骨肉瘤组织和细胞系中的表达下降,沉默骨肉瘤细胞SAOS-2中lncRNA H19的表达,能够抑制骨肉瘤细胞的增殖和侵袭,其机制可能与上调SAOS-2细胞中miR-194表达,进一步下调CDH2和IGF1R的表达有关。Objective To investigate the effect of long non-coding RNA(lncRNA)H19 targeting microRNA 194(miR-194)on the proliferation and invasion of osteosarcoma cells and its molecular mechanism.Methods Thirty osteosarcoma tissue specimens and 30 corresponding adjacent cancer tissue specimens were collected from patients who underwent osteosarcoma resection in the Second Department of Orthopedics,the Affiliated Dalian Fifth People’s Hospital of Dalian Medical University,from January 2018 to June 2019.The expressions of lncRNA H19 and miR-194 in 30 osteosarcoma tissues and corresponding adjacent tissues,human normal osteoblasts hFOB1.19 and osteosarcoma cells SAOS-2 and U2OS,were detected using qRT-PCR.lncRNA H19 small interfering RNA(si-H19),small interfering RNA negative control(si-NC),miR-194 mimics(miR-194 mimics),and miRNA negative control(miR-NC)were transfected into SAOS-2 cells by liposome transfection.Cell viability was detected by MTT assay,the invasive ability was detected by Transwell assay,and the expressions of cadherin 2(CDH2)and insulin-like growth factor 1 receptor(IGF1R)were detected by Western blotting.The relationship between lncRNA H19 and miR-194 were verified using bioinformatics,dual-luciferase reporter gene experiments,and qRT-PCR.Results The expression level of lncRNA H19 in osteosarcoma tissues was increased,and the expression level of miR-194 was decreased(all P<0.05)compared with those of adjacent tissues.The expression level of lncRNA H19 in SAOS-2 and U2OS osteosarcoma cells was increased(P<0.05),and the expression level of miR-194 was decreased(P<0.05)compared with those of hFOB1.19.The viability of SAOS-2 cells in the si-H19 group decreased,invasion ability decreased,and expression levels of CDH2 and IGF1R proteins decreased(P<0.05)compared with those of the si-NC group.The cell activity of SAOS-2 cells in the miR-194 mimic group decreased,invasion ability decreased,and expression levels of CDH2 and IGF1R proteins decreased(P<0.05)compared with those of the miR-NC group.Bio­informatic ana

关 键 词：长链非编码RNA H19 微RNA 194 骨肉瘤细胞 增殖 侵袭 

分 类 号：R738.1[医药卫生—肿瘤]