Preservation of the Blood Retinal Barrier (BRB) in Diabetes by Deletion of gp91phox or Inhibition of NAD(P)H Oxidase

Abstract

Purpose: : To study the role of NAD(P)H oxidase in diabetes–induced breakdown of the BRB. Vision loss in diabetic retinopathy is strongly correlated with increased vascular permeability. Increased formation of reactive oxygen species (ROS) is thought to be one of the key events in the pathogenesis of endothelial dysfunction in diabetes. We have recently demonstrated that ROS derived from NAD(P)H oxidase are important for hypoxia–induced VEGF expression and angiogenesis in a model of ischemic retinopathy. We have also shown that the protective effects of statin treatment in blocking diabetes–induced breakdown of BRB are accompanied by decreased gp91phox expression and reduced superoxide generation in retinal vessels. Thus, we hypothesized that NAD(P)H oxidase plays a key role in diabetes–induced VEGF expression and vascular dysfunction in retina.

Methods: : Diabetes was induced in wild–type and gp91phox–/– mice with streptozotocin (STZ). One group of STZ injected wild–type mice was treated with the NAD(P)H oxidase inhibitor apocynin (10mg/kg/day in drinking water). 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. Expression of gp91phox and VEGF was analyzed by immunofluorescence and Western blot.

Results: : Expression of gp91phox and formation of ROS were increased in the retinal vasculature of diabetic mice. Permeability assay showed that the BRB was preserved in diabetic mice treated with apocynin or lacking gp91phox. Moreover apocynin treatment or deletion of gp91phox gene reduced VEGF expression in the diabetic retina.

Conclusions: : These data indicate that NAD(P)H oxidase activity has a key role in diabetes–induced VEGF expression and breakdown of BRB and could be a therapeutic target to treat retinal vascular dysfunction associated with diabetes.

Keywords: diabetic retinopathy • oxidation/oxidative or free radical damage • growth factors/growth factor receptors