下调lncRNA HEIH负调控miR-214-3p逆转乳腺癌细胞的顺铂耐药性

Down-regulating lncRNA HEIH reverses cisplatin resistance in breast cancer cells by negatively regulating miR-214-3p

目的探讨长链非编码RNA(long non-coding RNA,lncRNA)HEIH对顺铂(cisplatin,DDP)耐药乳腺癌细胞耐药性的影响及作用机制。方法收集30例乳腺癌患者癌组织和对应的癌旁组织,实时定量聚合酶链反应(quantitative real time-polymerase chain reaction,qRT-PCR)检测组织中lncRNA HEIH和miR-214-3p表达量。体外培养乳腺癌细胞MCF-7及对DDP耐药乳腺癌细胞MCF-7/DDP,qRT-PCR检测细胞中lncRNA HEIH表达量。将MCF-7/DDP细胞分为si-NC组、si-HEIH组、si-HEIH+anti-miR-NC组和si-HEIH+anti-miR-214-3p组,细胞计数试剂盒-8(cell counting kit-8,CCK-8)法和流式细胞仪分别检测细胞增殖和凋亡,蛋白质印迹法检测细胞中细胞周期蛋白D1(CyclinD1)、p21、B淋巴细胞瘤-2(B lymphocyte tumor-2,Bcl-2)和B淋巴细胞瘤-2相关蛋白(B lymphocyte tumor-2 related protein,Bax)蛋白表达量。双荧光素酶报告基因实验验证lncRNA HEIH和miR-214-3p的调控关系。结果与癌旁组织比较,乳腺癌组织中lncRNA HEIH的表达量升高(P<0.05),miR-214-3p的表达量降低(P<0.05)。与MCF-7细胞比较,MCF-7/DDP细胞中lncRNA HEIH的表达量升高(P<0.05)。与si-NC组比较,si-HEIH组MCF-7/DDP细胞OD值及CyclinD1、Bcl-2蛋白表达降低(P<0.05),凋亡率及p21、Bax蛋白表达升高(P<0.05)。si-HEIH+anti-miR-214-3p组MCF-7/DDP细胞OD值及CyclinD1、Bcl-2蛋白表达高于si-HEIH+anti-miR-NC组(P<0.05),而凋亡率及p21、Bax蛋白表达低于si-HEIH+anti-miR-NC组(P<0.05)。lncRNA HEIH在MCF-7/DDP细胞中负调控miR-214-3p表达。结论下调lncRNA HEIH表达可能通过负调控miR-214-3p逆转对乳腺癌细胞对DDP的耐药性。

Objective To investigate the effect and mechanism of lncRNA HEIH on drug resistance of cisplatin(DDP)-resistant breast cancer cells.Methods 30 cases of breast cancer tissues and corresponding adjacent tissues were collected,and then qRT-PCR was used to detect the expression of lncRNA HEIH and miR-214-3p in the tissues.MCF-7 and MCF-7/DDP were cultured in vitro,and qRT-PCR was used to detect the expression level of lncRNA HEIH in the cells.MCF-7/DDP cells were divided into si-NC group,si-HEIH group,si-HEIH+anti-miR-NC group and si-HEIH+anti-miR-214-3p group.And then CCK-8 method and flow cytometry were used to detect the cell proliferation and apoptosis,respectively.Western blot was used to detect the protein expression of CyclinD1,P21,Bcl-2,and Bax in cells.The regulatory relationship between lncRNA HEIH and miR-214-3p was verified by dual luciferase reporter gene experiments.Results Compared with adjacent tissues,the expression level of lncRNA HEIH in breast cancer tissues increased(P<0.05),but the expression level of miR-214-3p decreased(P<0.05).Compared with MCF-7 cells,the expression level of lncRNA HEIH in MCF-7/DDP cells increased(P<0.05).Compared with the si-NC group,the OD value of MCF-7/DDP cells and the protein expression of CyclinD1 and Bcl-2 in si-HEIH group decreased(P<0.05),but the apoptosis rate and the protein expression of p21 and Bax increased(P<0.05).The OD value of MCF-7/DDP cells and the protein expression of CyclinD1 and Bcl-2 in si-HEIH+anti-miR-214-3p group were higher than those in the si-HEIH+anti-miR-NC group(P<0.05),but the apoptosis rate and the protein expression of p21 and Bax were lower than those in si-HEIH+anti-miR-NC group(P<0.05).lncRNA HEIH negatively regulated miR-214-3p expression in MCF-7/DDP cells.Conclusion Down-regulating lncRNA HEIH expression may reverse the resistance of breast cancer cells to DDP by negatively regulating miR-214-3p.