DJ-1通过促进EMT进程增强辐照诱导下食管鳞癌细胞的侵袭性

DJ-1 enhances the irradiation-induced invasiveness of esophageal squamous cell carcinoma cells by promoting EMT process

目的:探究敲低DJ-1/PARK7表达及联合辐照对食管鳞癌细胞迁移和侵袭能力的影响及分子机制。方法:利用短发夹RNA(short hairpin RNA,shRNA)稳定转染食管鳞癌细胞kyse150和kyse450,敲低DJ-1表达。Western blot检测细胞转染效率。Transwell迁移实验检测梯度辐照(0、4、6、8 Gy)下食管鳞癌细胞的迁移能力。将转染空病毒的对照组(shRNA-Control)与DJ-1敲低组(shRNA-DJ-1)细胞进行6 Gy X线辐照,细胞划痕实验及Transwell迁移侵袭实验检测各组食管鳞癌细胞迁移和侵袭能力。Western blot检测上皮细胞-间充质转化(epithelial-mesenchymal transition,EMT)途径相关蛋白上皮型钙黏蛋白(E-cadherin)、神经型钙黏蛋白(N-cadherin)、基质金属蛋白酶2(matrix metalloproteinase 2,MMP2)的表达变化。结果:敲低DJ-1后,食管鳞癌细胞kyse150和kyse450内DJ-1蛋白表达水平明显降低。Transwell迁移实验显示,6 Gy剂量辐照诱导食管鳞癌细胞kyse150和kyse450的迁移能力最强。细胞划痕实验及Transwell迁移侵袭实验显示,在6 Gy辐照条件下敲低DJ-1后kyse150和kyse450细胞迁移和侵袭能力减弱。Western blot显示,敲低DJ-1联合辐照后E-cadherin表达上升,N-cadherin表达下降,MMP2表达下降。结论:敲低DJ-1表达可以降低辐照诱导的食管鳞癌细胞kyse150和kyse450的迁移和侵袭能力,其机制可能与通过上调E-cadherin、下调N-cadherin和MMP2蛋白表达从而抑制细胞EMT进程有关。

Objective:To investigate the effect and the molecular mechanism of DJ-1/PARK7 on migration and invasion with irradiation of esophageal squamous cell carcinoma.Methods:Short hairpin RNA(shRNA)was used to stably transfect esophageal squamous cell carcinoma cells kyse150 and kyse450 and knockdown the expression of DJ-1 gene.Western blot was used to detect the transfection efficiency of cells.Transwell assay was used to detect the migration ability of esophageal squamous cell carcinoma cells under gradient irradiation(0,4,6,8 Gy).The control group cells transfected with empty virus and the DJ-1 knockdown group cells were irradiated by 6 Gy X-ray.Cell healing assay and Transwell assay were used to analyze the migration and invasion ability of esophageal squamous cell carcinoma cells.Western blot was used to detect the expression changes of epithelial-mesenchymal transition(EMT)pathway related proteins E-cadherin,N-cadherin and matrix metalloproteinase 2(MMP2).Results:After DJ-1 was knocked down,DJ-1 protein expression levels in esophageal squamous cell carcinoma cells kyse150 and kyse450 were significantly decreased.Transwell assay showed that,induced by 6 Gy radiation,esophageal squamous cell carcinoma cells kyse150 and kyse450 had the strongest migration ability.According to the cell healing assay and Transwell assay,the migration and invasion of kyse150 and kyse450 cells were reduced after DJ-1 was knocked down.Western blot analysis showed that E-cadherin expression increased,N-cadherin expression and MMP2 expression decreased after DJ-1 was knocked down.Conclusion:Knockdown of DJ-1 expression can reduce the irradiation-induced migration and invasion ability of esophageal squamous cell carcinoma cells kyse150 and kyse450,and the mechanism may be related to the inhibition of EMT process by up-regulating E-cadherin,down-regulating N-cadherin and MMP2 protein expression.