lncRNA LUCAT1对膀胱癌细胞增殖、凋亡的影响及其机制研究

Effect of lncRNA LUCAT1 on the proliferation and apoptosis of bladder cancer cells and its mechanism

目的探讨lncRNA LUCAT1对膀胱癌细胞的增殖和凋亡的影响以及潜在的作用机制。方法培养正常膀胱上皮细胞SV-HUC-1和膀胱癌细胞J82、5637、BIU-87,将膀胱癌细胞5637随机分为control组、si-NC组,si-LUCAT1组、pcDNA-NC组、pcDNA-LUCAT1组、si-LUCAT1+anti-miR-NC组、si-LUCAT1+anti-miR-493-5p组。qRT-PCR法检测miR-493-5p和LUCAT1表达水平;Western Blot检测蛋白表达;MTT法检测细胞活性;流式细胞术检测细胞凋亡;双荧光素酶报告实验验证LUCAT1和miR-493-5p的靶向关系。结果与膀胱上皮细胞SV-HUC-1相比,膀胱癌细胞J82、5637、BIU-87中LUCAT1的表达水平明显升高,miR-493-5p表达水平明显降低,差异均有统计学意义(P<0.05);与si-NC组相比,si-LUCAT1组细胞活性降低,凋亡率升高,CyclinD1表达水平降低,Caspase-3表达水平升高,p-JAK2和p-STAT蛋白的表达水平降低,差异均有统计学意义(P<0.05);miR-493-5p mimics与LUCAT1野生型报告质粒共转染的细胞荧光素酶活性降低,差异有统计学意义(P<0.05);而miR-493-5p mimics与LUCAT1突变型报告质粒共转染的细胞荧光素酶活性无显著变化,差异无统计学意义(P>0.05);与si-LUCAT1+anti-miR-NC组相比,si-LUCAT1+anti-miR-493-5p组细胞活性升高,细胞凋亡率降低,Cyclin D1表达水平升高,Caspase-3表达水平降低,p-JAK2和p-STAT蛋白表达水平升高,差异均有统计学意义(P<0.05)。结论lncRNALUCAT1可抑制膀胱癌细胞增殖,促进其凋亡,其机制可能与miR-493-5p及JAK/STAT信号通路有关。

Objective To investigate the effect of lncRNA LUCAT1 on the proliferation and apoptosis of bladder cancer cells and its potential mechanism.Methods Culture normal bladder epithelial cells SV-HUC-1 and bladder cancer cells J82,5637,BIU-87,and bladder cancer cells 5637 were randomly divided into the control group,si-NC group,si-LUCAT1 group,pcDNA-NC group,pcDNA-LUCAT1 group,si-LUCAT1+anti-miR-NC group,si-LUCAT1+anti-miR-493-5p group.qRT-PCR method was used to detect the expression of miR-493-5p and LUCAT1;Western blot was used to detect protein expression;MTT(3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide)method was used to detect cell viability;flow cytometry was used to detect apoptosis;dual luciferase report experiment was used to verify the targeting relationship between LUCAT1 and miR-493-5p.Results Compared with bladder epithelial cells SV-HUC-1,the expression level of LUCAT1 in bladder cancer cells J82,5637,and BIU-87 was significantly increased,and the expression level of miR-493-5p was significantly reduced,with statistically significant differences(all P<0.05).Compared with the si-NC group,the cell activity of the si-LUCAT1 group was decreased,the apoptosis rate was increased,the expression level of CyclinD1 was decreased,the expression level of Caspase-3 was increased,and the expression levels of p-JAK2 and p-STAT protein were decreased,with statistically significant differences(all P<0.05);the luciferase activity of cells co-transfected with miR-493-5p mimics and LUCAT1 wild-type reporter plasmid was decreased,and the difference was statistically significant(P<0.05);the luciferase activity of cells co-transfected with miR-493-5p mimics and LUCAT1 mutant reporter plasmid did not change significantly,and the difference was not statistically significant(P>0.05);compared with the si-LUCAT1+anti-miR-NC group,in the si-LUCAT1+anti-miR-493-5p group,cell viability was increased,cell apoptosis rate was decreased,cyclin D1 expression level was increased,caspase-3 expression level was decreased,and p-JAK2 and p-STAT protein expression levels were increased,with statistically significant differences(all P<0.05).Conclusion lncRNA LUCAT1 can inhibit the proliferation and promote the apoptosis of bladder cancer cells,and its mechanism may be related to miR-493-5p and JAK/STAT signaling pathway.