Simvastatin Inhibits Diabetes–Induced Retinal Vascular Hyperpermeability, Suppresses NAD(P)H Oxidase and Preserves the Balance Between VEGF and PEDF

Abstract

Abstract: : Purpose: In diabetic retinopathy, vascular hyperpermeability and vitreo–retinal neovascularization are associated with increased oxidative stress, elevated expression of the angiogenic growth factor VEGF and reduced expression of the angiostatic and suggested anti–permeability factor PEDF. Statins have potent vasoprotective actions independent of their cholesterol lowering function, including inhibition of NAD(P)H oxidase. Statins also prevent diabetes–induced VEGF over–expression and breakdown of the blood–retinal barrier by an unknown mechanism. We have shown before that NAD(P)H oxidase–derived oxidative stress has a key role in causing VEGF over–expression and vitreoretinal neovascularization in a model of ischemic retinopathy. The purpose of this work was to study statins' effect on diabetes–induced retinal vascular hyperpermeability and oxidative stress in relation to expression of VEGF, PEDF and the NAD(P)H oxidase catalytic subunit gp91phox. Methods:Diabetes was induced in rats with streptozotocin (45 mg/kg IV). One group of STZ injected rats was treated with simvastatin (5 mg/kg/day) delivered by osmotic pump for 30 days. Retinal permeability was assessed by extravasation of FITC–conjugated albumin. The average fluorescence intensity was calculated and normalized to a non–injected control retina and to plasma fluorescence intensity for each animal. Oxidative stress was analyzed by measuring levels of lipid peroxidation via malondialdehyde (MDA) production. Expression of the gp91phox subunit of NADPH oxidase, VEGF and PEDF was assessed by western blotting and immunohistochemistry. Results:Diabetes increased retinal vascular permeability (OD=11 + 2.2 vs 4 + 0.7 in control), and induced over–expression of gp91phox (OD=0.99 + 0.12 vs 0.6 + 0.04) and VEGF (OD=847 + 46 vs 350 + 15). These effects were accompanied by a decrease in the expression of PEDF to undetectable levels. Diabetes–induced oxidative stress was indicated by lipid peroxidation (MDA, nMol/mg protein=161 + 12 vs 102 + 15.4). Treatment with simvastatin prevented each of these effects. Conclusions: Simvastatin treatment prevents diabetes–induced retinal hyperpermeability. This action may occur through blocking VEGF over–expression and maintaining normal levels of PEDF. Suppression of NAD(P)H oxidase activity may be critical for preserving the balance between VEGF and PEDF.

Keywords: retina • diabetes • oxidation/oxidative or free radical damage