HMGA2在胃癌细胞上皮-间充质转化中的作用

Role of HMGA2 in epithelial-mesenchymal transition of gastric cancer cells

目的:探讨HMGA2在胃癌细胞上皮-间充质转化(EMT)中的作用及机制。方法:采用Western blot和RT-qPCR实验检测不同分化程度的人胃癌细胞株MKN45、MKN28和SGC7901以及人永生化胃黏膜上皮细胞株GES-1中HMGA2的表达水平;采用脂质体转染法将pcDNA3.0-HMGA2质粒转染至MKN28细胞中,将si-HMGA2干扰片段转染至MKN45细胞中,并采用Western blot和RT-qPCR实验检测转染效率;CCK-8实验检测HMGA2上调对MKN28细胞活力的影响以及HMGA2下调对MKN45细胞活力的影响;采用细胞迁移和侵袭实验检测HMGA2上调对MKN28细胞迁移和侵袭能力的影响;采用Western blot和RT-qPCR实验检测HMGA2过表达对MKN28细胞EMT相关标志蛋白上皮型钙黏蛋白(E-cadherin)、神经型钙黏蛋白(N-cadherin)和波形蛋白(vimentin)表达的影响以及敲减HMGA2表达对MKN45细胞E-cadherin、N-cadherin和vimentin表达的影响;采用RT-qPCR实验检测过表达HMGA2的MKN28细胞Wnt/β-catenin信号通路相关分子表达的变化。结果:HMGA2在不同分化程度的胃癌细胞中的表达水平是不同的(P<0.05)。上调MKN28细胞HMGA2的表达水平能够抑制细胞活力(P<0.05);而在MKN45细胞中下调HMGA2的表达水平能够增强细胞活力(P<0.05)。上调MKN28细胞HMGA2的表达水平能够促进细胞的迁移和侵袭能力(P<0.05),且E-cadherin表达降低,N-cadherin和vimentin表达升高(P<0.05);敲减HMGA2在MKN45细胞的表达水平使E-cadherin表达升高,而N-cadherin和vimentin表达降低(P<0.05)。上调MKN28细胞中HMGA2的表达水平,细胞内Wnt/β-catenin通路的β-catenin及其下游分子c-Myc和cyclin D1的mRNA表达水平显著增加(P<0.05)。结论:HMGA2与胃癌细胞迁移和侵袭能力密切相关,并且能够通过激活细胞内Wnt/β-catenin通路,促进胃癌细胞EMT。

AIM: To investigate the role of HMGA2 in the epithelial-mesenchymal transition(EMT) in gastric cancer cells. METHODS: The expression of HMGA2 in human gastric cancer cell lines with different degrees of differen-tiation(MKN45, MKN28 and SGC7901) and immortalized human gastric epithelial cell line GES-1 was determined by Western blot and RT-qPCR. pcDNA3.0-HMGA2 plasmid was transfected into the MKN28 cells by liposome method. Transfection of si-HMGA2 interference fragments into MKN45 cells was also performed. The transfection efficiency was evaluated by Western blot and RT-qPCR. The effects of HMGA2 over-expression in the MKN28 cells and knock-down in the MKN45 cells on the cell viability were measured by CCK-8 assay. The effects of HMGA2 over-expression in the MKN28 cells on the cell migration and invasion abilities were detected by wound healing and Transwell invasion assays. The effects of HMGA2 over-expression in the MKN28 cells and knock-down in the MKN45 cells on the expression of EMT-related markers E-cadherin, N-cadherin, vimentin at mRNA and protein levels were determined by RT-qPCR and Western blot. The changes of Wnt/β-catenin signaling pathway-related molecules in the MKN28 cells with HMGA2 over-expression were also determined by RT-qPCR. RESULTS: The expression levels of HMGA2 were quite different in different differentiation levels of gastric cancer cells(P<0.05). The increased expression level of HMGA2 in MKN28 cells inhibited the cell viability(P<0.05), while the decreased expression level of HMGA2 in MKN45 cells promoted the cell viability(P<0.05). The increased expression level of HMGA2 in MKN28 cells promoted cell migration and invasion(P<0.05), changed the expression of EMT-related markers(P<0.05), while the decreased expression level of HMGA2 in the MKN45 cells changed the expression of EMT-related markers(P<0.05). The increased expression level of HMGA2 in the MKN28 cells significantly increased the mRNA levels of β-catenin in the Wnt/β-catenin pathway and the downstream molecules c-Myc and cyclin D1(P<0.05). CONCLUSION: HMGA2 is closely related to the migration and invasion abilities of gastric cancer cells. Moreover, it promotes the EMT process of gastric cancer cells by activating Wnt/β-catenin pathway.