RESUMO
OBJECTIVE: The purpose of this study was to observe the apoptosis of A2780 cells transfected with the recombinant plasmid of pcDNA-Bax and 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-Bax as 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-9 and 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.
Assuntos
Apoptose , Neoplasias Ovarianas/patologia , Proteína X Associada a bcl-2/metabolismo , Caspase 3/metabolismo , Caspase 9/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular , Citocromos c/metabolismo , Feminino , Humanos , Mitocôndrias/metabolismo , TransfecçãoRESUMO
OBJECTIVE: To clone the Noxa gene and to observe the apoptosis of A549 cells transfected with the recombinant plasmid of pcDNA-Noxa. METHODS: The Noxa gene was obtained by PCR, and was cloned into pcDNA3. 1(-). A549 cells were transfected with the recombinant plasmid of pcDNA-Noxa. Western blot analysis was performed to determine the overexpression of Noxa. A549 cells were stained with Hoechst 33258 to observe the apoptosis. RESULTS: The recombinant plasmid of pcDNA-Noxa was successfully constructed evidenced by endonuclease digestion and sequence analysis. The overexpression of Noxa was identified using Western blot analysis. The recombinant plasmid of pcDNA-Noxa induced apoptosis of A549 cells. CONCLUSION: Nora has exhibited potential pro-apoptotic activity against A549 cells. This study is a foundation for further research into pro-apoptotic activity of Noxa gene.
