Yan Hongchao, Sun Yuping
Department of Oncology, Shandong University School Of Medicine, Jinan, Shandong 250012, P.R. China.
Department of Medical Oncology, Jinan Central Hospital Affiliated to Shandong University, Jinan, Shandong 250013, P.R. China.
Oncol Lett. 2014 Jul;8(1):426-430. doi: 10.3892/ol.2014.2063. Epub 2014 Apr 14.
The aim of the present study was to investigate the impact of the WW domain-containing oxidoreductase () gene on the mechanisms underlying epithelial-mesenchymal transition (EMT) in human ovarian cancer stem cells. Western blot analysis was performed to detect the differences in the expression of the EMT markers, E-cadherin, β-catenin, N-cadherin, vimentin and fibronectin, between human ovarian cancer stem cells and the human epithelial ovarian carcinoma cell line, HO-8910. A pcDNA3.1-WWOX eukaryotic expression vector was subsequently transfected into the ovarian cancer stem cells (recombinant plasmid group) or an empty plasmid (empty plasmid group) and non-transfected ovarian cancer stem cells (blank control group) served as the controls. Following the transfection of the gene, methyl thiazolyl tetrazolium cell viability and Transwell invasion assays, and western blot analysis were performed to detect changes in the proliferative capability and invasive capacity of ovarian cancer stem cells, as well as the expression of EMT markers and regulatory factors, Elf5 and Snail. The expression levels of E-cadherin and β-catenin in the ovarian cancer stem cells were identified to be significantly lower than those in the HO-8910 cells, whereas the expression levels of N-cadherin, vimentin and fibronectin in the ovarian cancer stem cells were found to be significantly higher than those in the HO-8910 cells. At each time point, the cellular proliferative capacity of the recombinant plasmid group was observed to be significantly lower than that of the empty plasmid or blank control groups (P<0.05 vs. the controls). The number of penetrating cells in the recombinant plasmid, empty plasmid and the blank control groups were 105.5±3.1, 199.7±3.4 and 191.4±4.1, respectively (mean ± standard error of the mean; P<0.05 vs. the controls). In addition, the protein expression of E-cadherin, β-catenin and Elf5 in the recombinant plasmid group was found to be significantly higher than that in the other two groups, whereas the protein expression of N-cadherin, vimentin, fibronectin and Snail in the recombinant plasmid group was significantly lower than that in the other two groups. An EMT exists in ovarian cancer stem cells, and the gene inhibits the cellular proliferation of ovarian cancer stem cells and reduces their invasive capability. Therefore, the gene may reverse the EMT in ovarian cancer stem cells by regulating the expression of the EMT regulatory factors, Elf5 and Snail.
本研究旨在探讨含WW结构域的氧化还原酶（WWOX）基因对人卵巢癌干细胞上皮-间质转化（EMT）相关机制的影响。采用蛋白质免疫印迹法分析人卵巢癌干细胞与人上皮性卵巢癌细胞系HO-8910中EMT标志物E-钙黏蛋白、β-连环蛋白、N-钙黏蛋白、波形蛋白和纤连蛋白表达的差异。随后将pcDNA3.1-WWOX真核表达载体转染至卵巢癌干细胞（重组质粒组），以空质粒转染的卵巢癌干细胞（空质粒组）和未转染的卵巢癌干细胞（空白对照组）作为对照。转染WWOX基因后，通过甲基噻唑基四氮唑盐细胞活力检测、Transwell侵袭实验及蛋白质免疫印迹法，检测卵巢癌干细胞增殖能力、侵袭能力的变化，以及EMT标志物和调控因子Elf5、Snail的表达情况。结果显示，卵巢癌干细胞中E-钙黏蛋白和β-连环蛋白的表达水平显著低于HO-8910细胞，而N-钙黏蛋白、波形蛋白和纤连蛋白的表达水平显著高于HO-8910细胞。在各个时间点，重组质粒组细胞的增殖能力均显著低于空质粒组和空白对照组（与对照组相比，P<0.05）。重组质粒组、空质粒组和空白对照组穿膜细胞数分别为105.5±3.1、199.7±3.4和191.4±4.1（平均值±平均标准误；与对照组相比，P<0.05）。此外，重组质粒组中E-钙黏蛋白、β-连环蛋白和Elf5的蛋白表达显著高于其他两组，而重组质粒组中N-钙黏蛋白、波形蛋白、纤连蛋白和Snail的蛋白表达显著低于其他两组。卵巢癌干细胞中存在EMT，WWOX基因可抑制卵巢癌干细胞的细胞增殖并降低其侵袭能力。因此，WWOX基因可能通过调控EMT调控因子Elf5和Snail的表达来逆转卵巢癌干细胞中的EMT。
