Recombinant adenovirus overexpressing nkx2.5 protects H9c2 cells against H2O2-induced apoptosis.
- Author:
Tao LI
1
;
Kesheng JIANG
;
Qin RUAN
;
Zhiqiang LIU
Author Information
1. Chemistry and Life Science College, Zhejiang Normal University, Jinhua 321004, Zhejiang, China. litao@zjnu.cn
- Publication Type:Journal Article
- MeSH:
Adenoviridae;
genetics;
metabolism;
Animals;
Apoptosis;
drug effects;
Caspase 3;
metabolism;
Cell Line;
Genetic Vectors;
genetics;
Homeobox Protein Nkx-2.5;
Homeodomain Proteins;
biosynthesis;
genetics;
Hydrogen Peroxide;
pharmacology;
Myocytes, Cardiac;
cytology;
Oxidative Stress;
drug effects;
Proto-Oncogene Proteins c-bcl-2;
metabolism;
Rats;
Recombinant Proteins;
biosynthesis;
genetics;
Transcription Factors;
biosynthesis;
genetics;
bcl-2-Associated X Protein;
metabolism
- From:
Chinese Journal of Biotechnology
2012;28(10):1253-1264
- CountryChina
- Language:Chinese
-
Abstract:
To study the function and potential application of nkx2.5, a critical gene for heart development, we constructed a recombinant adenovirus overexpressing nkx2.5 gene (Ad-Nkx2.5) with the AdEasy system. To evaluate the effect and mechanism of Ad-Nkx2.5 against oxidative injury, the H9c2 myocardial cells were infected with the recombinant adenoviruses Ad-Nkx2.5 or Ad-EGFP, and subsequently exposed to H2O2 to induce apoptosis. The anti-apoptotic potential of Ad-Nkx2.5 was validated by MTT assay for cell viability, Hoechst33342 staining for cellular morphology, and immunoblotting for caspase-3 activity. Ad-Nkx2.5 infection led to an increased survival rate of H9c2 cells and decreased the amount of caspase-3 in an active form. Additionally, overexpression of Nkx2.5 inhibited the release of cytochrome C from the mitochondria into the cytosol. Mechanismic studies showed that Nkx2.5 upregulated bcl-2 gene expression and significantly repressed H2O2-induced expression of bax detected by Real-time PCR. Additionally, H2O2 treatment did not affect the nuclear localization of Nkx2.5. These findings indicate that adenovirus-mediated nkx2.5 gene transfer exerted a protective effect on H9c2 cells against H2O2-induced apoptosis via mitochondrial pathway, and the Nkx2.5-mediated expression modulation of apoptosis-associated genes could be involved in this event.
