AP-4基因表达下调抑制子宫内膜癌细胞侵袭和迁移能力

目的:观察靶向抑制激活增强子结合蛋白-4(AP-4)基因的表达对子宫内膜癌细胞侵袭和迁移能力的影响。方法:用荧光定量PCR(qRT-PCR)检测人子宫内膜癌细胞株HEC-1A、RL-952、HEC-1B、ishikawa中AP-4基因的表达水平。采用脂质体介导的细胞转染法将AP-4 siRNA转染至HEC-1A细胞,同时设置转染对照。转染48 h后,采用qRT-PCR和蛋白印迹法(Western blot)检测转染后各组HEC-1A细胞中AP-4的表达水平,通过细胞划痕实验观察转染后各组细胞迁移情况,采用Transwell实验检测转染后各组细胞侵袭能力。采用Western blot检测转染后各组细胞中上皮细胞标志分子E-钙粘蛋白(E-cadherin)和间充细胞标志分子N-钙粘素(N-cadherin)、波形蛋白(Vimentin)表达水平。结果:AP-1基因在四株子宫内膜癌细胞中均有表达,在HEC-1A细胞中相对表达水平最高(P<0.05),用于后续转染实验。qRT-PCR和Western blot结果均显示转染AP-4 siRNA能够成功抑制HEC-1A细胞中AP-1 mRNA和蛋白的表达(P<0.05)。Transwell实验显示转染AP-4 siRNA后HEC-1A细胞侵袭能力明显受到抑制(P<0.05)。划痕实验显示转染AP-4 siRNA后HEC-1A细胞迁移能力明显受到抑制(P<0.05)。Western blot结果显示转染AP-4 siRNA后HEC-1A细胞中N-cadherin和Vimentin蛋白水平降低,E-cadherin蛋白水平升高,抑制了上皮间质转化(EMT)过程(P<0.05)。结论:AP-4基因表达下调能够有效抑制子宫内膜癌细胞的侵袭和迁移能力,该过程可能与逆转细胞EMT过程有关。

Down-regulation of AP-4 gene expression inbibits invasion and migration of endometrial cancer cells

Objective:To observe the effect of targeting inhibitory activation enhancer binding protein-4(AP-4)gene expression on the invasion and migration of endometrial cancer cells.Methods:The expression levels of AP-4 gene in human endometrial cancer cell lines HEC-1A，RL-952，HEC-1B and ishikawa were detected by real-time PCR(qRT-PCR).AP-4 siRNA was transfected into HEC-1A cells by liposome-mediated cell transfection and a transfection control was set up.After transfection for 48 h，the expression levels of AP-4 in HEC-1A cells were detected by qRT-PCR and Western blot.The cell migration was observed by cell scratch test.Transwell assay was used to detect the invasive ability of each group after transfection.Western blot was used to detect the expression levels of E-cadherin and N-cadherin and Vimentin in the epithelial cells of each group after transfection.Results:AP-1 gene was expressed in all four endometrial cancer cells，and the expression level was highest in HEC-1A cells(P<0.05)，which was used for subsequent transfection experiments.Both qRT-PCR and Western blot showed that AP-4 siRNA could successfully inhibit the expression of AP-1 mRNA and protein in HEC-1A cells(P<0.05).Transwell experiments showed that the invasive ability of HEC-1A cells was significantly inhibited after transfection with AP-4 siRNA(P<0.05).Scratch experiments showed that the migration ability of HEC-1A cells was significantly inhibited after transfection of AP-4 siRNA(P<0.05).Western blot results showed that the levels of N-cadherin and Vimentin protein in HEC-1A cells were decreased after transfection with AP-4 siRNA，and the level of E-cadherin protein was increased，which inhibited the process of epithelial-mesenchymal transition(EMT)(P<0.05).Conclusion:The down-regulation of AP-4 gene expression can effectively inhibit the invasion and migration of endometrial cancer cells，which may be related to the reversal of cell EMT.