VASCULAR-PERMEABILITY IN EXPERIMENTAL DIABETES IS ASSOCIATED WITH REDUCED ENDOTHELIAL OCCLUDIN CONTENT - VASCULAR ENDOTHELIAL GROWTH-FACTORDECREASES OCCLUDIN IN RETINAL ENDOTHELIAL-CELLS

Blood-retinal barrier (BRB) breakdown is a hallmark of diabetic retino pathy, but the molecular changes that cause this pathology are unclear . Occludin is a transmembrane component of interendothelial tight junc tions that may regulate permeability at the BRB. In this study, we exa mined the effects of vascular endothelial growth factor (VEGF) and dia betes on vascular occludin content and barrier function. Sprague-Dawle y rats were made diabetic by intravenous streptozotocin injection, and age-matched animals served as controls. After 3 months, BRB permeabil ity was quantified by intravenous injection of fluorescein isothiocyan ate-bovine serum albumin (FITC-BSA), M-r 66 kDa, and 10-kDa rhodamine- dextran (R-D), followed by digital image analysis of retinal sections. Retinal fluorescence intensity for FITC-BSA increased 62% (P less tha n or equal to 0.05), but R-D fluorescence did not change significantly Occludin localization at interendothelial junctions was confirmed by immunofluorescence, and relative protein content was determined by imm unoblotting of retinal homogenates, Retinal occludin content decreased -35% (P less than or equal to 0.03) in the diabetic versus the contro l animals, whereas the glucose transporter GLUT1 content was unchanged in rat retinas. Additionally, treatment of bovine retinal endothelial cells in culture with 0.12 nmol/l or 12 nmol/l VEGF for 6 h reduced o ccludin content 46 and 54%, respectively. These data show that diabete s selectively reduces retinal occludin protein expression and increase s BRB permeability. Our findings suggest that the elevated VEGF in the vitreous of patients with diabetic retinopathy increases vascular per meability by downregulating occludin content. Decreased tight junction protein expression may be an important means by which diabetes causes increased vascular permeability and contributes to macular edema.