Abstract 428: Cytoglobin: A Novel Regulator of the DNA Damage Response Within Cardiomyocytes

Abstract

Rationale: Oxidative stress is a major determinant of DNA damage within cardiomyocytes both at baseline and following cardiac injury. Cytoglobin (Cygb), a stress responsive hemoprotein, serves as a redox modulator and imparts cytoprotection in a variety of cell types. In addition, Cygb is involved in regulation of myocyte proliferation and differentiation due to its ability to scavenge free radicals and maintain redox equilibrium within the myocytes. Objective: The goal of this study was to investigate the role of Cygb in cardiomyocytes and identify the molecular pathways underlying its cytoprotective function. Method and Results: Utilizing transgenic mice with cardiac specific overexpression of Cygb (TgCygb), we demonstrated that Cygb overexpression promoted cardiomyocyte proliferation as compared to the control hearts (NTg) (phospho-Histone 3 IHC: 17 ± 0.7 vs 7 ± 0.5 positive cells/mm 2 TgCygb vs NTg; p<0.05, n=3) . The enhanced cellular proliferation in TgCygb hearts was associated with larger hearts at baseline. However, with exposure to acute cardiac stress (subcutaneous Angiotensin II infusion), TgCygb hearts showed significantly reduced cell death, oxidative stress and an overall lower induction of the DNA damage response pathway as compared to the NTg hearts [TUNEL: 7 ± 0.8 vs 10 ± 1.2 TUNEL positive nuclei/mm 2 , p<0.05, n=3; phospho-ATM IHC: 11 ± 2 vs 22 ± 1.5 phospho-ATM positive cells/mm 2 , p<0.05, n=5, dihydroethidium (DHE) staining: 78 ± 0.6% vs 98 ± 0.7% DHE positive nuclei/mm 2 , p<0.05, n=5]. Using single cell electrophoresis (Comet assay), we demonstrated that Cygb overexpressing cardiomyocytes underwent significantly lower DNA damage due to single and double stranded DNA breaks following exposure to oxidative stress as compared to controls cardiomyocytes (36 ± 2.1 vs 48 ± 3.1; p<0.05, n=6). Finally, chromatin immunoprecipitation, quantitative PCR and immunoblotting indicated that Cygb activates the expression of apurinic/apyrimidinic endonuclease 1 ( Ape1 ) and thus promotes DNA base excision repair following acute cardiac injury. Conclusions: Our data demonstrate that Cygb promotes cellular proliferation and protects cardiomyocytes against oxidative stress-mediated DNA damage by directly activating Ape1 expression.