miR-186-5p通过靶向调控PTTG1抑制肺腺癌细胞的上皮-间质转化

发布时间：2018-07-09 23:05

本文选题：肺腺癌 + miR--p　；参考：《中国生物化学与分子生物学报》2017年04期

【摘要】：先前研究表明,miR-186-5p在人类许多恶性肿瘤中扮演抑癌基因的作用,但其在肺腺癌上皮-间质转化(epithelial-mesenchymal transition,EMT)中的作用并不明确。本研究旨在证明,miR-186-5p可通过靶向调控PTTG1抑制肺腺癌细胞的上皮-间质转化。我们首先分析了miR-186-5p在人肺癌细胞中的表达。荧光定量PCR(QRT-PCR)结果显示,与人正常肺上皮细胞BEAS-2B相比,肺腺癌细胞SPC-A1、A549中的miR-186-5p表达量明显降低。为研究miR-186-5p在肺腺癌细胞中的功能,利用GV369-miR-186-5p表达载体,实现了在A549细胞中的过表达。基因转染结合Transwell侵袭结果显示,与对照质粒转染的A549细胞相比,过表达miR-186-5p的A549细胞的体外侵袭能力明显下降。Western印迹检测细胞中EMT相关标志物揭示,GV369-miR-186-5p转染的A549细胞中的上皮-钙黏着蛋白(E-cadherin)表达明显上调,而神经-钙黏着蛋白(N-cadherin)和波形蛋白(vimentin)表达明显下调。同时,GV369-miR-186-5p转染引起其靶基因——垂体肿瘤转化基因1(pituitary tumor-transforming gene 1,PTTG1)编码蛋白在A549细胞中明显降低。重要的是,过表达miR-186-5p与敲减PTTG1均可导致上皮-钙黏着蛋白表达上调,而神经-钙黏着蛋白和波形蛋白下调;而miR-186-5p和PTTG1表达载体共转染后,3种EMT相关标志物在A549的表达与对照细胞的表达无明显差异,提示过表达PTTG1可抵消miR-186-5p对EMT相关标志物表达的影响。综上所述,miR-186-5p可通过靶向调控PTTG1抑制EMT的发生,进而抑制肺腺癌细胞的侵袭转移。
[Abstract]:Previous studies have shown that miR-186-5p plays a role as a tumor suppressor gene in many human malignancies, but its role in epithelial-mesenchymal transition is not clear. The aim of this study was to demonstrate that miR-186-5p can inhibit the epithelial-interstitial transformation of lung adenocarcinoma cells by targeting PTTG1. We first analyzed the expression of miR-186-5p in human lung cancer cells. The results of fluorescence quantitative PCR (QRT-PCR) showed that the expression of miR-186-5p in human lung adenocarcinoma cell line SPC-A1 / A549 was significantly lower than that in normal human lung epithelial cell line BEAS-2B. In order to study the function of miR-186-5p in lung adenocarcinoma cells, overexpression of miR-186-5p in A549 cells was achieved by using GV369-miR-186-5p expression vector. The results of gene transfection combined with Transwell invasion showed that compared with A549 cells transfected with control plasmid, The invasion ability of A549 cells overexpressed by miR-186-5p was significantly decreased. Western blot analysis revealed that the expression of E-cadherin in A549 cells transfected with GV369-miR-186-5p was up-regulated, and the expression of E-cadherin was up-regulated by Western blotting. The expression of N-cadherin and vimentin (vimentin) was down-regulated. At the same time, GV369-miR-186-5p transfection resulted in the decrease of pituitary tumor transforming gene 1 (pituitary tumor-transforming gene 1 / PTTG1) coding protein in A549 cells. Importantly, overexpression of miR-186-5p and knockdown of PTTG1 could result in up-regulation of epithelial-cadherin expression, but down-regulation of neuron-cadherin and vimentin. However, after co-transfection of miR-186-5p and PTTG1, the expression of three EMT related markers in A549 cells was not significantly different from that in control cells, suggesting that over-expression of PTTG1 could counteract the effect of miR-186-5p on the expression of EMT related markers. In conclusion, miR-186-5p can inhibit the development of EMT by targeting PTTG1, and then inhibit the invasion and metastasis of lung adenocarcinoma cells.
【作者单位】：山东省潍坊医学院生物科学与技术学院;山东省潍坊医学院病理学教研室;山东省潍坊医学院医学研究实验中心;山东省潍坊医学院护理学院;
【基金】：国家自然科学基金青年基金项目(No.81402389,No.81641111) 山东省自然科学基金(No.ZR2015HL065) 山东省高等学校科技计划(No.J12LK03,J13LK03) 潍坊医学院大学生科技创新基金项目(No.KX2016016,KX2016022)资助~~
【分类号】：R734.2

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