Abstract
Purpose::
Breakdown of the blood-retinal barrier (BRB) is a central event in early diabetic retinopathy. The role of extracellular proteases in angiogenesis and retinal neovascularization occurring in late proliferative diabetic retinopathy, has been well-documented. Their role in early retinal vascular permeability changes however remains unexplored. The aim of this study was to determine the role of the urokinase plasminogen activator (uPA) and its receptor system (uPAR) in mediating breakdown of the BRB in diabetes.
Methods::
The mRNA levels of both uPA and uPAR were assessed in the retinal vasculature, following 2 weeks of diabetes in the rat. To examine the function of these proteins in retinal microvascular permeability, diabetic animals were treated with a peptide inhibitor derived from the non-receptor-binding region of urokinase, that blocks the interaction of uPA with uPAR (Å6, Angstrom Pharmaceuticals, San Diego). Diabetic rats received intraperitoneal injections of Å6 (100mg/kg) daily for 2-weeks after induction of diabetes. Retinal vascular permeability was assayed using the Evans Blue technique. To elucidate how inhibition by Å6 affects cell junctions, endothelial cells were treated with AGE-BSA, a known inducer of permeability, in the presence or absence of Å6. Under these conditions, VE-cadherin surface expression and induction of MMPs were examined by immunochemistry and zymography respectively. Monolayer permeability to FTIC-dextran was also evaluated in a modified Boyden chamber assay.
Results::
Diabetic rats treated with Å6 showed a 71% decrease in vascular permeability as compared to untreated diabetic rats. AGE-BSA treatment of cells increased monolayer permeability, caused a loss of cell surface VE-cadherin expression, decreased cell-cell contact and significantly upregulated the expression of MMP-2 and 9. All of these effects were effectively reversed by simultaneous treatment with Å6.
Conclusions::
The uPA/uPAR system might be crucial in mediating the breakdown of the blood-retinal-barrier during diabetes. Inhibition of this uPA and uPAR interaction might be a promising target for early therapeutic intervention in diabetic retinopathy.
Keywords: diabetic retinopathy • edema • proteolysis