Shen Jingang, Lv Xianbao, Zhang Lei
Department of Urology, Chengwu County People's Hospital, Shandong 274200, People's Republic of China.
Department of Urology, Zoucheng People's Hospital, Shandong 273500, People's Republic of China.
Onco Targets Ther. 2020 Mar 25;13:2511-2522. doi: 10.2147/OTT.S239120. eCollection 2020.
GRHL2 played important roles in different cancers. In this study, we aimed to investigate the roles of GRHL2 in bladder cancer.
The immunohistochemistry assay was performed to detect the expression of GRHL2 in bladder cancer tissues and adjacent noncancerous tissues and the expression levels of GRHL2 and zinc finger E-box binding homeobox (ZEB1) mRNA in tissues were determined by qRT-PCR. In addition, qRT-PCR and Western blotting were applied to detect the expression levels of GRHL2 and ZEB1 in bladder cancer cell lines (RT4, BIU-87, 5637, T24) and immortalized human bladder epithelial cell line (SV-HUC-1). The cell models with up-regulated and down-regulated expression of GRHL2 were constructed using bladder cancer cell lines T24 and 5637 to investigate the underlying roles of GRHL2 on the proliferation, migration, invasion and EMT process of bladder cancer cells. After that, cell proliferation was evaluated by CCK8 assay, cell cycle assay and colony formation assay. Transwell assay and wound healing assay were performed to determine the invasion and migration ability of the bladder cancer cells. The expressions of epithelial-mesenchymal transition (EMT) related proteins (E-cadherin, Vimentin, Slug and Snail) were assessed by Western blot analysis. Moreover, ZEB1 and GRHL2 were co-transfected into T24 and 5637 cells and their effects on EMT process and invasive capacity of cells were examined.
The expression of GRHL2 was down-regulated in bladder cancer tissues and human bladder cancer cell lines compared with the normal bladder tissues and immortalized human bladder epithelial cell line. Besides, down-regulation of GRHL2 improved the proliferation ability of bladder cancer cells and promoted the EMT process through up-regulation of ZEB1. The overexpression of ZEB1 partially reversed the inhibitory effect of GRHL2 on EMT.
GRHL2 acts as an anti-oncogene to regulate bladder cancer cell proliferation and inhibit EMT by targeting ZEB1. This study may provide a theoretical basis for further research.
GRHL2在不同癌症中发挥重要作用。在本研究中，我们旨在探究GRHL2在膀胱癌中的作用。
采用免疫组织化学法检测GRHL2在膀胱癌组织及癌旁非癌组织中的表达，并通过qRT-PCR测定组织中GRHL2和锌指E盒结合同源框蛋白（ZEB1）mRNA的表达水平。此外，运用qRT-PCR和蛋白质免疫印迹法检测GRHL2和ZEB1在膀胱癌细胞系（RT4、BIU-87、5637、T24）及永生化人膀胱上皮细胞系（SV-HUC-1）中的表达水平。利用膀胱癌细胞系T24和5637构建GRHL2表达上调和下调的细胞模型，以研究GRHL2对膀胱癌细胞增殖、迁移、侵袭及上皮-间质转化（EMT）过程的潜在作用。之后，通过CCK8法、细胞周期检测和集落形成实验评估细胞增殖情况。采用Transwell实验和伤口愈合实验测定膀胱癌细胞的侵袭和迁移能力。通过蛋白质免疫印迹分析评估EMT相关蛋白（E-钙黏蛋白、波形蛋白、Slug和Snail）的表达。此外，将ZEB1和GRHL2共转染至T24和5637细胞中，检测它们对细胞EMT过程和侵袭能力的影响。
与正常膀胱组织和永生化人膀胱上皮细胞系相比，GRHL2在膀胱癌组织和人膀胱癌细胞系中的表达下调。此外，GRHL2的下调通过上调ZEB1提高了膀胱癌细胞的增殖能力并促进了EMT过程。ZEB1的过表达部分逆转了GRHL2对EMT的抑制作用。
GRHL2作为一种抑癌基因，通过靶向ZEB1调节膀胱癌细胞增殖并抑制EMT。本研究可能为进一步研究提供理论依据。
