长链非编码RNA LINC01234通过靶向微小RNA-940调控前列腺癌细胞的增殖、凋亡、迁移及侵袭

Long non-coding RNA LINC01234 regulates proliferation,apoptosis,migration and invasion of prostate cancer cells by targeting microRNA-940

目的探讨长链非编码RNA(lncRNA)LINC01234对前列腺癌细胞的增殖、凋亡、迁移及侵袭的影响及其分子机制。方法采用实时定量反转录聚合酶链反应(RT-qPCR)法检测收集2017年5月至2018年7月郑州大学第一附属医院收治的20例前列腺癌患者的前列腺癌组织及癌旁组织中LINC01234、微小RNA(miRNA,miR)-940的表达水平。分别将LINC01234小干扰RNA(si-LINC01234)、乱序无意义阴性序列(si-NC)、微小RNA-940模拟物(miR-940 mimics)、阴性对照基因(miR-NC)、si-LINC01234+阴性对照(anti-miR-NC)、si-LINC01234+微小RNA 940特异性寡核苷酸抑制剂(anti-miR-940)转染至DU145细胞。分别记作si-NC、si-LINC01234、miR-NC、miR-940、si-LINC01234+anti-miR-NC、si-LINC01234+anti-miR-940组。噻唑蓝(MTT)检测各组细胞的活性;流式细胞术检测各组细胞凋亡率;Transwell法检测各组细胞的迁移及侵袭;双荧光素酶报告实验验证LINC01234与miR-940的靶向关系。两组间比较采用独立样本t检验,多组间比较采用单因素方差分析。结果LINC01234在前列腺癌组织中的表达水平高于癌旁组织(1.00±0.08比2.61±0.25,t=27.430,P<0.05),miR-940在前列腺癌组织中的表达水平低于癌旁组织(1.00±0.09比0.56±0.05,t=19.112,P<0.05),差异有统计学意义;转染si-LINC01234、miR-940 mimics后细胞活力显著降低(1.29±0.11比0.71±0.06;1.24±0.09比0.79±0.07,t=13.886,P<0.05),迁移细胞数[(84.36±8.12)个比(39.41±4.22)个;(86.25±8.14)个比(44.37±4.48)个]与侵袭细胞数[(67.25±6.74)个比(28.44±3.58)个;(69.48±6.25)个比(33.47±4.18)个]显著减少(t=14.735、15.309,P<0.05),细胞凋亡率[(6.58±0.61)%比(20.36±2.14)%;(7.11±0.71)%比(18.26±1.36)%]显著升高(t=18.577,P<0.05),差异均有统计学意义;双荧光素酶报告实验证实miR-940过表达可抑制野生型(WT)-LINC01234细胞的荧光活性,LINC01234可负向调控miR-940的表达(t=328.298,P<0.05),差异有统计学意义;干扰miR-940表达可逆转抑制LINC01234表达对前列腺癌细胞增殖、迁移、侵袭的抑制作用及其对凋亡的促进作用。结论lncRNA LINC01234可能通过靶向miR-940促进前列腺癌细胞增殖、迁移及侵袭,抑制细胞凋亡。

Objective To investigate the effects of long non-coding RNA(lncRNA)LINC01234 on proliferation,apoptosis,migration and invasion of prostate cancer cells and its molecular mechanism.Methods The expression levels of LINC01234 and microRNA(miRNA,miR)-940 in prostate cancer tissues and adjacent tissues were detected by real-time quantitative reverse transcriptase-polymerase chain reaction(RT-qPCR).The si-LINC01234,si-NC,miR-940 mimics,miR-NC,si-LINC01234+anti-miR-NC,si-LINC01234+anti-miR-940 were transfected into DU145 cells,respectively.Methyl thiazolyl tetrazolium(MTT)assay was used to detect the activity of cells in each group.Flow cytometry was used to detect the apoptosis rate of each group.Transwell method was used to detect the migration and invasion of cells in each group.The dual luciferase report experiment verified the targeting relationship between LINC01234 and miR-940.Results The expression level of LINC01234 in prostate cancer tissues was significantly higher than that in adjacent tissues(1.00±0.08 vs.2.61±0.25,t=27.430,P<0.05),and that of miR-940 in prostate cancer tissues was significantly lower than that in adjacent tissues(1.00±0.09 vs.0.56±0.05,t=19.112,P<0.05).After transfection with si-LINC01234and miR-940 mimics,cell viability was significantly reduced(1.29±0.11 vs.0.71±0.06;1.24±0.09 vs.0.79±0.07,t=13.886,P<0.05),number of migrating cells(84.36±8.12 vs.39.41±4.22;86.25±8.14 vs.44.37±4.48)and the number of invasive cells(67.25±6.74 vs.28.44±3.58;69.48±6.25 vs.33.47±4.18)were significantly reduced(t=14.735,15.309,P<0.05),the apoptosis rate[(6.58±0.61)%vs.(20.36±2.14)%;(7.11±0.71)%vs.(18.26±1.36)%]was significantly increased(t=18.577,P<0.05).The dual luciferase report experiment confirmed that miR-940 overexpression could inhibit the fluorescence activity of wild-type WT-LINC01234 cells,and LINC01234 could negatively regulate the expression of miR-940(t=328.298,P<0.05).Interfering with the expression of miR-940 could reverse the inhibition of LINC01234 expression on the proliferation,migration and invasion of prostate cancer cells and its promoting effect on apoptosis.Conclusion LncRNA LINC01234 promotes proliferation,migration and invasion of prostate cancer cells,and inhibits apoptosis probably through targeting miR-940.