Abstract 18789: BRCA2 is a Novel Regulator of Endothelial Cell Function and Apoptosis Following Hyperglycemia

Abstract

Background: Germ-line mutations in the tumor suppressor gene BRCA2 (breast cancer 2, early onset) predispose carriers mostly to breast cancer, and BRCA2 mutation carriers also face a 2-fold increase in the risk of developing diabetes.Hyperglycemia, the hallmark of diabetes, is a major risk factor for endothelial dysfunction leading to vascular complications. The relationship linking role of BRCA2 in the development of diabetes and endothelial dysfunction remains mainly unexplored. Methods: To elucidate the role of BRCA2 in diabetes and endothelial apoptosis/dysfunction in vitro, we silenced BRCA2 in human umbilical vein endothelial cells (ECs) and evaluated the markers of EC function and apoptosis following hyperglycemia. Results: We first confirmed the basal expression of BRCA2 in ECs at transcript and protein levels by qPCR and immunoblotting, respectively. Interestingly, hyperglycemia significantly induced BRCA2 expression after 48 hours of treatment. We then silenced BRCA2 in ECs and confirmed successful silencing at transcript and protein levels. Hyperglycemia significantly increased the reactive oxygen species (ROS) production in the BRCA2-deficient ECs in comparison to control ECs. Increased ROS production was associated with exacerbated DNA-damage in BRCA2-silenced ECs as evidenced by increased expression and activation of DNA double-stranded breaks (DSBs) marker H2A.X and reduced RAD51-foci formation, an essential regulator of DSB repair. Increased DSBs were associated with significantly increased activation of p53. Elevated levels of DNA-damage and activated-p53 were further associated with significantly increased hyperglycemia-induced apoptosis in BRCA2-silenced ECs as measured by immunoblotting for cleaved-caspase-3, Bax and by TUNEL-staining.Key indices of endothelial function, including tube formation and NO production, were significantly reduced following hyperglycemia-treatment in BRCA2-deficient ECs. Conclusion: Our data for the first time, show an entirely novel role of BRCA2 as a regulator of endothelium in the setting of hyperglycemia, and provide important clues regarding the potential susceptibility of BRCA2 mutation carriers to hyperglycemia-induced cardiovascular complications.