550. Adenoviral-Mediated Antioxidant Overexpression: Possible Protective Role Against Hyperglycemic Conditions in Human Aortic Endothelial Cells

Abstract

Intoduction: Increased oxidative stress is a well documented characteristic of diabetes mellitus, which plays a key role in the pathogenesis of diabetic vascular dysfunction. An increase in oxygen free radicals (O2-) is believed to be associated with diabetes mellitus. Efficient free radical scavenging is imperative for normal endothelial cell function and survival in diabetes mellitus. As superoxide dismutase (SOD) is responsible for scavenging O2- in eukaryotic cells, the aim of this study was to assess if transducing Human Aortic Enothelial Cells (HAEC) with recombinant adenovirus encoding MnSOD, CuZnSOD and Catalase, could reverse high glucose induced decrease in cell viability. Methods: HAECs exposed to adenoviral vectors encoding MnSOD (AdMnSOD), CuZnSOD (AdCuZnSOD), Catalase (AdCat) or Green Fluorescent Protein (AdGFP) or diluent (control) were cultured in normal glucose (NG)(5.5 mmol/L) or high glucose (HG) (30 mmol/L) medium. Cell viability was assessed by Methlythiazolydiphenyl-tetrazolium bromide (MTT) Assay. Transgene expression was detected by confocal microscopy in AdGFP transduced cells and by Western blot analysis for AdMnSOD/ AdCuZnSOD/ AdCat-transduced cells 48 hours post transduction. Results: From initial non-transduced cell viability assays, HAECs in HG displayed decrease in viability of 14% (n=11, P|[le]|0.01) compared to NG. Under HG conditions, exposure to CuZnSOD and MnSOD resulted in increased cell viability (versus non-transduced controls). Under NG conditions, exposure to MnSOD and CuZnSOD resulted in decreased cell viability (versus non-transduced controls). Catalase reduced viability in NG and HG conditions, but the latter to a lesser extent. Control vector AdGFP had no effect on viability. Conclusion: Decreased viability observed in HAECs under HG conditions can be reversed by overexpression of MnSOD and CuZnSOD. As such, antioxidant overexpression may hold promise in protecting endothelial cells against hyperglycemic conditions.