lncRNA FLVCR1-AS1靶向miR-381-3p对结直肠癌细胞增殖、凋亡的影响及其分子机制  

Effects of lncRNA FLVCR1-AS1 targeting miR-381-3p on proliferation and apoptosis of colorectal cancer cells and molecular mechanism

作　　者：许国彩[1] 刘芝兰[1] 逯艳艳[1] XU Guocai;LIU Zhilan;LU Yanyan(Qinghai Provincial People′s Hospital,Xining 810002,China)

机构地区：[1]青海省人民医院,西宁810002

出　　处：《山东医药》2020年第29期5-9,共5页Shandong Medical Journal

摘　　要：目的探讨长链非编码RNA FLVCR1-AS1(lncRNA FLVCR1-AS1)调控微小RNA-381-3p(miR-381-3p)表达影响结直肠癌(CRC)细胞增殖、凋亡的作用及机制。方法取正常结肠上皮细胞FHC及CRC细胞SW480、DLD-1、HCT116,采用qRT-PCR法检测细胞中的FLVCR1-AS1与miR-381-3p。将FLVCR1-AS1表达最高的CRC细胞随机分为7组:NC组不处理,si-FLVCR1-AS1组、si-con组分别转染FLVCR1-AS1小干扰RNA及乱序无意义序列,miR-381-3p组、miR-con组分别转染miR-381-3p模拟物(mimics)及阴性对照mimic NC序列(miR-con),si-FLVCR1-AS1+anti-miR-381-3p组、si-FLVCR1-AS1+anti-miR-con组分别共转染si-FLVCR1-AS1与miR-381-3p特异性寡核苷酸抑制剂及其阴性对照;采用MTT法测算细胞存活率,流式细胞术测算细胞凋亡率,Western blotting法检测细胞周期蛋白D1(Cyclin D1)、活化的含半胱氨酸的天冬氨酸蛋白水解酶3(Cleaved-Caspase-3)。先用Starbase对FLVCR1-AS1和miR-381-3p结合进行预测,再以双荧光素酶报告基因检测FLVCR1-AS1与miR-381-3p的相互作用。结果与FHC比较,SW480、DLD-1、HCT116中FLVCR1-AS1表达水平均升高而miR-381-3p表达水平均降低(P均<0.05),其中SW480的FLVCR1-AS1表达最高。与NC组、si-con组、miR-con组比较,si-FLVCR1-AS1组、si-FLVCR1-AS1+anti-miR-con组细胞存活率降低、细胞凋亡率升高,细胞中Cyclin D1蛋白表达量降低、Cleaved-Caspase-3蛋白表达量升高(P均<0.05);miR-381-3p组细胞存活率降低、细胞凋亡率升高,细胞中Cyclin D1蛋白表达量降低、Cleaved-Caspase-3蛋白表达量升高(P均<0.05);NC组、si-con组、miR-con组两两比较,差异均无统计学意义;与si-FLVCR1-AS1+anti-miR-con组比较,si-FLVCR1-AS1+anti-miR-381-3p组细胞存活率升高、细胞凋亡率降低,细胞中Cyclin D1蛋白表达量升高、Cleaved-Caspase-3蛋白表达量低(P均<0.05)。Starbase预测显示,FLVCR1-AS1与miR-381-3p存在靶向结合位点;双荧光素酶报告实验显示,在转染野生型载体WT-FLVCR1-AS1的SW480细胞中,共转染miR-381-3Objective To investigate the effects and mechanism of long non-coding RNA FLVCR1-AS1(lncRNA FLVCR1-AS1)regulating microRNA-381-3p(miR-381-3p)expression on the proliferation and apoptosis of colorectal cancer(CRC)cells.Methods We took the normal colonic epithelial cells FHC and CRC cells SW480,DLD-1,HCT116,and used qRT-PCR to detect FLVCR1-AS1 and miR-381-3p in the cells.The CRC cells with the highest FLVCR1-AS1 expression were randomly divided into 7 groups:cells in the NC group were not treated,cells in the si-FLVCR1-AS1 group and si-con group were transfected with FLVCR1-AS1 small interfering RNA and disordered nonsense sequence,the cells in the miR-381-3p group and miR-con group were transfected with miR-381-3p mimics(mimics)and negative control mimic NC sequence(miR-con),respectively,and the cells in the si-FLVCR1-AS1+anti-miR-381-3p group and si-FLVCR1-AS1+anti-miR-con group were co-transfected with si-FLVCR1-AS1 and miR-381-3p specific oligonucleotide inhibitors and their negative controls,respectively.The cell survival rate was measured by MTT.Flow cytometry was used to measure the apoptosis rate.Western blotting was used to detect Cyclin D1(Cyclin D1)and activated cysteine-containing aspartate proteolytic enzyme 3(Cleaved-Caspase-3).First,Starbase was used to predict the binding of FLVCR1-AS1 and miR-381-3p,and then the dual luciferase reporter gene was used to detect the interaction between FLVCR1-AS1 and miR-381-3p.Results Compared with the FHC cells,the expression level of FLVCR1-AS1 increased in the SW480,DLD-1,and HCT116 cells,while the expression level of miR-381-3p decreased(all P<0.05),and the expression of FLVCR1-AS1 in SW480 cells was the highest.Compared with the NC group,si-con group and miR-con group,si-FLVCR1-AS1 group,and si-FLVCR1-AS1+anti-miR-con group had lower cell survival rates and higher cell apoptosis rates,the protein expression of Cyclin D1 in cells decreased,and the protein expression of Cleaved-Caspase-3 increased(all P<0.05).In the miR-381-3p group,the cell survival rate decrea

关 键 词：结直肠癌 长链非编码RNA FLVCR1-AS1 miR-381-3p 细胞增殖 细胞凋亡 

分 类 号：R735[医药卫生—肿瘤]