【摘要】 目的观察Bax对人卵巢癌A2780细胞的凋亡诱导作用,并探讨其作用机制。方法构建重组质粒pcDNA-Bax,将重组质粒pcDNA-Bax转染A2780细胞,采用Hoechst 33258染色观察细胞凋亡、MTT法检测细胞活力,采用Western blot法检测Bax基因的过表达、线粒体细胞色素C的释放及Caspase-9和Caspase-3的激活。结果经酶切和测序鉴定,重组质粒pcDNA-Bax构建成功。转染质粒pcDNA-Bax能明显诱导A2780细胞凋亡,Hoechst染色显示凋亡的细胞出现细胞核形态学改变;MTT结果显示转染质粒pcDNA3.1-Bax后,细胞活力明显下降;Western blot结果表明转染质粒pcDNA-Bax后的A2780细胞线粒体释放细胞色素C,激活了Caspase-9和Caspase-3。结论 Bax能促进卵巢癌细胞线粒体细胞色素C释放,从而诱导细胞凋亡。更多还原

【Abstract】 Objective The purpose of this study was to observe the apoptosis of A2780 cells transfected with the recombinant plasmid of pcDNA-Baxand to observe the release of cytochrome C from the mitochondria.Methods The recombinant plasmid of pcDNA-Bax was constructed and transfected into A2784 cells.The Hoechst 33258 stain method was applied to evaluate the apoptosis of the transfected cells and MTT mothod was used to test the cell viability.Western blot analysis was performed to determine the overexpression of Bax and the release of cytochrome C from the mitochondria.Results The recombinant plasmid of pcDNA-Bax was successfully constructed by using endonuclease digestion and the sequence analysis.The apoptosis of A2780 cells was induced after transfected with pcDNA3.1-Baxas demonstrated with Hoechst staining.The cell viability were decreased in the pcDNA3.1-Bax transfected group by MTT assay.The release of cytochrome C from the mitochondria was observed when using Western blotting analysis.And the caspase-9and the caspase-3 were activated.Conclusion Our data suggestted that Bax exhibited potent pro-apoptotic activity against the ovarian cancer cells.This study is a foundation for the further research in the pro-apoptotic activity of Bax.更多还原