lncRNA TPTEP1通过抑制miR-129-5p影响膀胱癌T24细胞的增殖和侵袭

lncRNA TPTEP1 affects the proliferation and invasion of bladder cancer T24 cells by inhibiting miR-129-5p

目的:分析长链非编码RNA(long-chain non-coding RNA,lncRNA)TPTEP1在膀胱癌组织和细胞的表达,观察其对膀胱癌细胞增殖和侵袭的影响及其作用机制。方法:收集2017年8月至2019年10月在武汉市东西湖人民医院泌尿外科接受手术治疗的43例膀胱癌患者的癌与癌旁组织标本,采用实时荧光定量聚合酶链式反应(qPCR)检测膀胱癌组织、膀胱癌细胞系(T24、BIU-87、5637、J82、UM-UC-3)中lncRNA TPTEP1的表达;选择表达最低的膀胱癌细胞为实验对象,分别转染阴性对照质粒(对照组)和lncRNA TPTEP1过表达质粒(实验组),采用MTT法和Transwell实验检测上调lncRNA TPTEP1对细胞增殖和侵袭的影响。采用生物信息学技术预测lncRNA TPTEP1可能的靶分子,qPCR和WB检测lncRNA TPTEP1下游分子的表达水平。结果:与癌旁组织比较,膀胱癌组织中lncRNA TPTEP1的表达下调(P<0.01);与正常膀胱上皮细胞比较,各膀胱癌细胞系中lncRNA TPTEP1的表达均下调(均P<0.05),以T24细胞中的表达最低(P<0.01)。上调lncRNATPTEP1可抑制T24细胞的增殖(P<0.05)和侵袭(P<0.01)。生物信息学技术显示,lncRNA TPTEP1可互补结合miR-129-5p,miR-129-5p可互补结合EMP3。上调lncRNA TPTEP1可抑制T24细胞中miR-129-5p的表达(P<0.01),从而间接促进EMP3 mRNA和蛋白的表达(P<0.01),但p-MEK、p-ERK1/2、p-AKT、p-PI3K等MAPK/ERK信号通路相关蛋白表达均降低(均P<0.01)。结论:上调膀胱癌细胞系中低表达的lncRNA TPTEP1可抑制膀胱癌T24细胞系的增殖和侵袭,其作用机制与其通过下调miR-129-5p的表达间接促进EMP3基因表达有关。

Objective: To observe the expression of long-chain non-coding RNA(lncRNA) TPTEP1 in bladder cancer tissues and cells, and to observe its effect on the proliferation and invasion of bladder cancer cells and its molecular mechanism. Methods: From August 2017 to October 2019, 43 cases of bladder cancer tissues and paracancer tissues from the patients treated by surgery in the Department Urology, People’s Hospital of Dongxihu Distric of Wuhan City. Real-time fluorescence quantitative polymerase chain reaction(qPCR) was used to detect the expression of lncRNA TPTEP1 in bladder cancer tissues and bladder cancer cell lines(T24, BIU-87, 5637, J82, UM-UC-3). The bladder cancer cells with the lowest lncRNA TPTEP1 expression were selected as the experimental object, and transfected with the negative control plasmid(the control group) and lncRNA TPTEP1 over-expression plasmid(the experimental group), respectively. The effect of lncRNA TPTEP1 upregulation on cell proliferation and invasion was detected by MTT method and Transwell experiment. Bioinformatics techniques were used to predict the possible target molecules of lncRNA TPTEP1.qPCR and WB were used to detect the expression levels of lncRNA TPTEP1 downstream molecules. Results: Compared with adjacent tissues, the expression of lncRNA TPTEP1 in bladder cancer tissues was down-regulated(P<0.01). Compared with normal bladder epithelial cells, the expression of lncRNA TPTEP1 in bladder cancer cell lines was down-regulated(P<0.05), and its expression in T24 cells was the lowest(P<0.01). Up-regulation of lncRNA TPTEP1 could inhibit the proliferation(P<0.05) and invasion(P<0.01) of T24 cells. Bioinformatics technology showed that lncRNA TPTEP1 could bind with miR-129-5 p, and miR-129-5 p could bind with EMP3;up-regulating lncRNA TPTEP1 could inhibit the expression of miR-129-5 p in T24 cells(P<0.01), and indirectly promote the mRNA and protein expressions of EMP3(P<0.01) in T24 cells. The expression of MAPK/ERK signaling pathway related proteins such as p-MEK, p-ERK1/2, p-AKT and p-PI3 K decreased(P<0.01). Conclusion: Up-regulating the low-expressed lncRNA TPTEP1 in bladder cancer cell lines can inhibit the proliferation and invasion of bladder cancer T24 cells, and its mechanism is related to indirect promotion of EMP3 gene expression by down-regulating the expression of miR-129-5 p.