Tie2在口腔鳞状细胞癌中的表达及其对细胞增殖、迁移及上皮-间充质转化过程的影响

目的 研究酪氨酸激酶受体2(tyrosine kinase receptor 2, Tie2)在口腔鳞状细胞癌（oral squamous cell carcinoma, OSCC）中的表达情况及其对细胞增殖、迁移及上皮-间充质转化（epithelial-mesenchymal transition, EMT）过程的影响。方法 采用免疫组织化学（immunohistochemistry, IHC）检测Tie2在OSCC组织与正常口腔黏膜组织中的表达。Western blot检测DOK细胞及OSCC细胞株中Tie2的表达，选取Tie2高表达细胞株作为实验株。将沉默Tie2的慢病毒载体成功转染至实验株用于后续实验。CCK-8及克隆形成实验检测细胞增殖、克隆能力；细胞划痕和Transwell实验检测细胞迁移能力；激光共聚焦显微镜检测细胞骨架纤丝状肌动蛋白（F-actin）重塑能力及上皮钙黏素（E-cadherin）、神经钙黏素（Ncadherin）的表达；Western blot检测EMT相关标志性蛋白E-cadherin、N-cadherin、波形蛋白（vimentin）的表达以及蛋白激酶...

Expression of Tyrosine Kinase Receptor 2 in Oral Squamous Cell Carcinoma and the Effect on Cell Proliferation and Migration and Epithelial-Mesenchymal Transition Process

  Objective  To study the expression of tyrosine kinase receptor 2 (Tie2) in oral squamous cell carcinoma (OSCC) and its effect on cell proliferation and migration and the epithelial-mesenchymal transition (EMT) process.   Methods  Immunohistochemistry (IHC) tests were conducted to examine the expression of Tie2 in OSCC tissues and normal oral mucosa tissues. Western blot was performed to examine the expression of Tie2 in dysplastic oral keratinocyte (DOK) cell line and OSCC cell lines, and the cell line with high Tie2 expression was selected as the experimental cell line. The Tie2-silenced lentiviral vector was successfully transfected onto the experimental cell line for subsequent experiments. Cell proliferation and cloning abilities were examined with CCK-8 and clone formation assays. Cell migration ability was examined with scratch and Transwell assays. The remodeling ability of cytoskeletal F-actin and the expressions of E-cadherin and N-cadherin were examined with confocal laser scanning microscope. Western blot was performed to examine the expression of EMT-related signature proteins, including E-cadherin, N-cadherin, and vimentin, and the expression of the protein kinase B (AKT) and extracellular signal-regulated kinase (ERK).  Results   IHC results showed that the Tie2-positive rate of the OSCC group (74.5%) was significantly higher than that of the control group (19.4%) (P<0.0001). The expression of Tie2 was higher in HSC-4 and SCC-9 cell lines compared to that in DOK cells. The lentiviral shRNA-162 group showed the best silencing effect, which was used as the experimental group and applied in subsequent experiments. Compared with those of the control group, the proliferation, cloning and migration capacities of the cells of the experimental group were significantly reduced. Furthermore, the green fluorescence intensity of the cytoskeleton F-actin was reduced, the number of filamentous pseudopods at the leading edge of the cells decreased and their length was shortened, and the expression of E-cadherin was significantly increased, while the expression of N-cadherin and vimentin was significantly reduced in the experimental group in comparison with those of the control group. The expression of p-AKT and p-ERK proteins decreased, while AKT and ERK protein expression increased.   Conclusion  Tie2 was highly expressed in most OSCC cells. Silencing Tie2 can inhibit the proliferation, cloning, and migration ability of OSCC cells, inhibit F-actin remodeling, and alter the expression of its EMT-related signature proteins by regulating AKT and ERK signaling pathway, which suggests that Tie2 may be involved in the growth, metastasis and EMT process of OSCC.