LINC00665靶向调控miR-34a-5p对食管癌细胞增殖、迁移和侵袭能力的影响

The effects of LINC00665 targeting miR-34a-5p on the proliferation,migration and invasion of esophageal cancer cells

目的:探讨LINC00665靶向调控微小RNA(miR)-34a-5p对食管癌细胞增殖、迁移和侵袭能力的影响。方法:选取食管癌病人肿瘤组织及癌旁组织,采用qRT-PCR法检测组织中LINC00665、miR-34a-5p表达水平。以食管癌细胞株EC9706为研究对象,设置对照组、阴性转染(siNC)组、LINC00665小干扰RNA载体(siLINC00665)组、siLINC00665+inhibitor阴性转染(anti-NC)组、siLINC00665+miR-34a-5p inhibitor(anti-miR-34a-5p)组,CCK-8法检测各组细胞增殖能力;Transwell实验检测细胞侵袭、迁移数;Western blotting检测基质金属蛋白酶2(MMP-2)、细胞周期素D1(CyclinD1)、p21蛋白表达水平;双荧光素酶实验验证LINC00665和miR-34a-5p的靶向关系。结果:食管癌病人肿瘤组织LINC00665表达水平较癌旁组织明显增加,miR-34a-5p表达水平明显下降(P<0.01)。与对照组、siNC组相比,siLINC00665组细胞OD值(24、48 h)、侵袭和迁移数、CyclinD1、MMP-2蛋白、LINC00665表达均降低,p21蛋白、miR-34a-5p表达均增加(P<0.05);与siLINC00665+anti-NC组相比,siLINC00665+anti-miR-34a-5p组细胞OD值(24、48 h)、侵袭和迁移数、CyclinD1、MMP-2蛋白、LINC00665表达均增加,p21蛋白、miR-34a-5p表达均降低(P<0.05)。双荧光素酶实验证实LINC00665和miR-34a-5p靶向关系。结论:沉默LINC00665可靶向上调miR-34a-5p表达,进而抑制食管癌细胞增殖、迁移和侵袭能力。

Objective:To investigate the effects of LINC00665 targeting and regulating micrornas(miR)34a-5p on the proliferation,migration and invasion of esophageal cancer cells.Methods:The cancer tissues and adjacent tissues of patients with esophageal cancer were collected,the expression levels of LINC00665 and miR-34a-5p in tissues were detected by qRT-PCR.The esophageal cancer cell line EC9706 were divided into the control group,negative transfection(siNC)group,LINC00665 small interfering RNA vector(siLINC00665)group,siLINC00665+inhibitor negative transfection(anti-NC)group and siLINC00665+miR-34a-5p inhibitor(anti-miR-34a-5p)group.The proliferation ability of cells was detected by CCK-8 method,the numbers of cell invasion and migration were detected by Transwell assay;the protein expression levels of matrix metalloproteinase 2(MMP-2),cyclin D1(CyclinD1)and p21 were measured by Western blotting,and the targeting relationship between LINC00665 and miR-34a-5p was verified by dual-luciferase experiment.Results:The expression level of LINC00665 in cancer tissues of patients with esophageal cancer was obviously higher than that in adjacent tissues,and the expression level of miR-34a-5p in cancer tissues was obviously lower(P<0.01).Compared with the control group and siNC group,the OD value(24,48 h),numbers of invasion and migration,expression levels of CyclinD1,MMP-2 proteins and LINC00665 in siLINC00665 group were obviously lower,and the expression levels of p21 protein and miR-34a-5p was obviously higher(P<0.05).Compared with the siLINC00665+anti-NC group,the OD value(24,48 h),numbers of invasion and migration,expression levels of CyclinD1,MMP-2 proteins and LINC00665 in siLINC00665+anti-miR-34a-5p group were obviously higher,and the expression levels of p21 protein and miR-34a-5p were obviously lower(P<0.05).The targeting relationship between LINC00665 and miR-34a-5p was confirmed by dual-luciferase reporter gene experiment.Conclusions:Silencing LINC00665 can target and up-regulate the expression of miR-34a-5p,thereby inhibiting the proliferation,migration and invasion of esophageal cancer cells.