摘要
目的 探讨长链非编码RNA(lncRNA)MINCR在喉癌细胞增殖、迁移、侵袭和顺铂耐药性中的作用,通过其对miR-584-3p的调节确定LncRNA MINCR的潜在机制。方法 采用实时定量聚合酶链式反应(RT-qPCR)检测喉癌患者癌组织和喉癌细胞系中,LncRNA MINCR和miR-584-3p表达情况。将喉癌M4E细胞分为NC组(对照,未转染)、si-con组、si-LncRNA MINCR组、miR-con组、miR-584-3p组、anti-miR-con+si-LncRNA MINCR组、anti-miR-584-3p+si-LncRNA MINCR组。采用Transwell法检测细胞迁移、侵袭,Western blot检测多药耐药相关蛋白1(MRP1)、基质金属蛋白酶(MMP)2、MMP9表达,二苯基四氮唑溴盐(MTT)法检测细胞增殖活力和计算顺铂的半数抑制浓度(IC50)变化,双荧光素酶活性检测分析LncRNA MINCR和miR-584-3p的结合。结果喉癌患者癌组织中LncRNA MINCR的表达量比癌旁组织高(分别为3.140±0.330、1.000±0.150,P<0.01),且在喉癌细胞系M4E、Tu177、AMC-HN-8中的表达量(分别为3.010±0.240、2.840±0.260、2.340±0.220)比鼻咽上皮细胞NP69(1.000±0.110)增加(P<0.01)。喉癌患者癌组织中miR-584-3p表达量比癌旁组织低(分别为0.450±0.050、1.000±0.130,P<0.01),且在喉癌细胞系M4E、Tu177、AMC-HN-8中的表达量(分别为0.410±0.040、0.490±0.040、0.540±0.050)比鼻咽上皮细胞NP69(1.000±0.120)低(P<0.01)。低表达LncRNA MINCR后,细胞吸光度值、迁移细胞数、侵袭细胞数、IC50值减少,且MRP1、MMP2及MMP9蛋白表达降低(P<0.01)。过表达miR-584-3p后,细胞吸光度值、迁移细胞数、侵袭细胞数、IC50值减少,且MRP1、MMP2及MMP9蛋白表达降低(P<0.01)。LncRNA MINCR可作为ceRNA吸附调控miR-584-3p的表达。和anti-miR-con+si-LncRNA MINCR组比较,anti-miR-584-3p+si-LncRNA MINCR组细胞吸光度值、迁移细胞数、侵袭细胞数、IC50值减少,且MRP1、MMP2及MMP9蛋白表达降低(P<0.01)。结论 低表达LncRNA MINCR通过靶向miR-584-3p,降低喉癌细胞对顺铂的耐药性。
Objective To investigate the role of long non-coding RNA MINCR(LncRNA MINCR)in laryngeal cancer cell proliferation,migration,invasion and cisplatin resistance,and determine the potential mechanism of LncRNA MINCR through its regulation of miR-584-3p.Methods LncRNA MINCR and miR-584-3p in cancer tissues and laryngeal carcinoma cell lines of patients with laryngeal carcinoma was analyzed by real-time quantitative polymerase chain reaction(RT-qPCR).Divided laryngeal cancer M4E cells into NC group(control,untransfected),si-con group,si-LncRNA MINCR group,miR-con group,miR-584-3p group,anti-miR-con+si-LncRNA MINCR Group,anti-miR-584-3p+si-LncRNA MINCR group.Transwell method monitored cell migration and invasion.Expression of multidrug resistance-related protein 1(MRP1),matrix metalloproteinase(MMP)2,and MMP9 was assessed by Western blot.The 3-(4,5-Dimethylthazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT)method was used to detect cell proliferation activity and calculates the change of the half inhibitory concentration(IC50)of cisplatin,and the dual luciferase activity detection analyzed the targeted binding of LncRNA MINCR and miR-584-3p.Results The expression level of LncRNA MINCR in laryngeal cancer tissues was higher than that in para-cancerous tissues(3.140±0.330 and 1.000±0.150,respectively,P<0.01),and its expression in laryngeal cell lines M4E,Tu177,and AMC-HN-8(3.010±0.240,2.840±0.260 and 2.340±0.220,respectively)was higher than that in nasopharyngeal epithelial cell NP69(1.000±0.110)(P<0.01).The expression level of miR-584-3p in cancerous tissues of laryngeal cancer patients was lower than that in para-cancerous tissues(0.450±0.050 and 1.000±0.130,respectively,P<0.01),and its expression in laryngeal cell lines M4E,Tu177,and AMC-HN-8(0.410±0.040,0.490±0.040 and 0.540±0.050,respectively)was lower than that in nasopharyngeal epithelial cell NP69(1.000±0.120)(P<0.01).After knocking down LncRNA MINCR expression,cell absorbance values,migration cell numbers,invasion cell numbers,and IC50 values were decreased,and the expression of MRP1,MMP2,and MMP9 proteins also were decreased(P<0.01).After overexpressing miR-584-3p,cell absorbance values,migration cell numbers,invasion cell numbers,and IC50 values were decreased,and the expression of MRP1,MMP2,and MMP9 proteins also were decreased(P<0.01).LncRNA MINCR acted as a ceRNA to regulate the expression of miR-584-3p.Compared with the anti-miR-con+si-LncRNA MINCR group,the anti-miR-584-3p+si-LncRNA MINCR group had reduced cell absorbance values,migration cell numbers,invasion cell numbers,and IC50 values,as well as reduced expression of MRP1,MMP2 and MMP9(P<0.01).Conclusion The low expression of LncRNA MINCR can reduce the resistance of laryngeal cancer cells to cisplatin by targeting miR-584-3p.
作者
褚丹丹
施涛
许莉
薛飞
陈伟
程友
CHU Dan-dan;SHI Tao;XU Li;XUE-Fei;CHEN Wei;CHENG You(Department of Otolaryngology-Head and Neck Surgery,General Hospital of Eastern Theater Command,Nanjing,Jiangsu Province 210002 China)
出处
《解剖学研究》
2025年第4期330-336,343,共8页
Anatomy Research
基金
江苏省科技项目(BK20161388)
东部战区总医院军事医学面上项目(YYMS2021021)。
