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目的:探讨有丝分裂检查点蛋白着丝粒蛋白-E(CENP-E)基因在肿瘤发生发展中的作用。方法:利用shRNA下调CENP-E基因的表达,分别用巢式PCR和Western blot检测CENP-E mRNA和蛋白的表达;MTT检测CENP-E下调后MCF-7细胞的增殖变化;流式细胞术检测CENP-E下调后对MCF-7细胞凋亡的影响;Transwell试验检测MCF-7细胞的迁移和侵袭能力变化;间接免疫荧光检测细胞内CENP-E蛋白和有丝分裂情况。结果:shRNA能有效抑制CENP-E mRNA和蛋白的表达。MTT结果显示CENP-E下调后MCF-7细胞的增殖能力减弱(P<0.05);流式细胞术显示下调CENP-E后能促进MCF-7细胞的凋亡;间接荧光结果显示CENP-E干扰后MCF-7细胞内CENP-E蛋白减少并伴有核分裂异常;Transwell试验显示CENP-E干扰组细胞的迁移和侵袭能力增强(P<0.05)。结论:下调部分CENP-E的表达能抑制MCF-7细胞的增殖,促进MCF-7细胞的凋亡,增强MCF-7细胞的迁移和侵袭能力。
Objective: To investigate the role of mitotic checkpoint protein CENP-E in tumorigenesis. Methods: The expression of CENP-E gene was down-regulated by shRNA. The expression of CENP-E mRNA and protein was detected by nested PCR and Western blot respectively. The proliferation of MCF-7 cells was detected by MTT assay. The effect of CENP-E on the apoptosis of MCF-7 cells was detected by flow cytometry. The changes of migration and invasion of MCF-7 cells were detected by Transwell assay. CENP-E protein and mitosis were detected by indirect immunofluorescence. Results: shRNA can effectively inhibit CENP-E mRNA and protein expression. The results of MTT showed that the proliferation of MCF-7 cells was decreased after CENP-E down-regulation (P <0.05). Flow cytometry showed that downregulation of CENP-E promoted the apoptosis of MCF-7 cells. Indirect fluorescence showed CENP-E interference The CENP-E protein was decreased and accompanied by mitotic abnormality in MCF-7 cells. Transwell assay showed that CENP-E knockdown cells had more migration and invasion ability (P <0.05). Conclusion: The down-regulation of CENP-E expression can inhibit the proliferation of MCF-7 cells, promote the apoptosis of MCF-7 cells and enhance the migration and invasion ability of MCF-7 cells.