miR-145通过下调PLCε抑制膀胱癌EMT和迁移及其机制研究

发布时间：2018-03-04 22:14

本文选题：PLCε　切入点：miR-　出处：《中国生物工程杂志》2017年03期 　论文类型：期刊论文

【摘要】：目的:探讨miR-145调控PLCε对膀胱癌细胞T24上皮间质转化(EMT)和迁移的影响及可能的分子机制。方法:(1)腺病毒感染T24细胞,划痕实验和Transwell检测细胞的迁移能力;RTPCR、Western blot分别检测PLCε及EMT相关分子的表达;为探究其分子机制,Western blot检测GSK-3β磷酸化(Ser9位点)和Snail的表达情况。(2)利用生物信息学技术预测可能调控PLCε的miRNA,结合文献报道的膀胱癌microRNA表达谱结果筛选出miR-145;转染miR-145 mimics至T24,q PCR检测miR-145、PLCε的表达,Western blot检测PLCε的表达。(3)转染miR-145 mimics,Western blot检测EMT相关分子及p-GSK-3β、Snail;划痕实验、Transwell检测过表达miR-145后细胞的迁移能力。结果:(1)干扰PLCε表达能显著抑制细胞的迁移,同时,使T24细胞中E-cadherin表达上调,N-cadherin和Vimentin表达下调;干扰PLCε后,GSK-3β磷酸化(Ser9位点)水平下降,Snail表达降低。(2)转染miR-145 mimics可使T24细胞中miR-145表达增高,且明显抑制T24细胞中PLCε的表达。(3)在T24中过表达miR-145,细胞迁移能力显著下降,EMT标志分子的表达情况与沉默PLCε结果一致。同时,与阴性对照组相比,转染miR-145 mimics组p-GSK-3β和Snail表达显著减少。结论:PLCε通过GSK-3β/Snail信号通路促进膀胱癌细胞T24发生EMT及迁移,miR-145可以逆转PLCε诱导膀胱癌EMT的发生,从而阻止膀胱癌细胞的迁移。
[Abstract]:Objective: to investigate the effect and possible molecular mechanism of miR-145 regulating PLC 蔚 on the epithelial mesenchymal transformation and migration of bladder cancer cell line T24. Methods: T24 cells were infected with adenovirus. The migration ability of cells was detected by scratch test and Transwell. The expression of PLC 蔚 and EMT related molecules were detected by RT PCR Western blot. In order to explore its molecular mechanism, the expression of GSK-3 尾 phosphorylation site Ser9) and Snail were detected by Western blot) and the miRNAs that might regulate PLC 蔚 were predicted by bioinformatics. MiR-145 was screened according to the reported results of microRNA expression in bladder cancer and transfected with miR-145 mimics. T24Q PCR was used to detect the expression of miR-145P PLC 蔚. Western blot was used to detect the expression of PLC 蔚.) transfection miR-145 mimicsl Western blot was used to detect EMT related molecules and p-GSK-3 尾 Snail.The scratch test was used to detect the migration ability of the cells after miR-145 expression. At the same time, the expression of E-cadherin in T24 cells was up-regulated and the expression of N-cadherin and Vimentin was down-regulated, and the level of GSK-3 尾 phosphorylated ser9 was decreased after interfering with PLC 蔚. The expression of miR-145 in T24 cells was increased by transfection of miR-145 mimics. The overexpression of miR-145in T24 cells was significantly inhibited, and the expression of PLC markers was significantly decreased in T24 cells, which was consistent with the results of silencing PLC 蔚. At the same time, compared with the negative control group, the expression of miR-145in T24 cells was similar to that in the negative control group. Conclusion the expression of p-GSK-3 尾 and Snail decreased significantly in miR-145 mimics transfection group. Conclusion the expression of p-GSK-3 尾 and Snail in T24 cells induced by GSK-3 尾 / Snail signal pathway can be reversed by PLC 蔚 -induced EMT, thus preventing the migration of T24 cells.
【作者单位】：重庆医科大学检验医学院临床检验诊断学教育部重点实验室重庆市重点实验室;重庆医科大学附属第一医院泌尿外科;
【基金】：国家自然科学基金资助项目(81072086)
【分类号】：R737.14

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