Hyperglycemia-induced ubiquitination and degradation of β-catenin with the loss of platelet endothelial cell adhesion molecule-1 in retinal endothelial cells.

Abstract

Increased retinal vascular permeability is one of the earliest manifestations of diabetic retinopathy. The aim of this study was to investigate the role of hyperglycemia-induced platelet endothelial cell adhesion molecule-1 loss on retinal vascular permeability via the β-catenin pathway. Type I diabetes was induced in male Wistar rats using streptozotocin injections, with age-matched non-diabetic rats as controls. Rat retinal microvascular endothelial cells were grown under normal or high glucose conditions for 6days. Small interfering Ribonucleic Acid was used to knock down platelet endothelial cell adhesion molecule-1 in rat retinal microvascular endothelial cells for loss-of-function studies. Retinas and rat retinal microvascular endothelial cells were subjected to Western blot, immunofluorescence labeling, and co-immunoprecipitation analyses to assess protein levels and interactions. A biotinylated gelatin and fluorescein isothiocyanate-avidin assay was used for retinal endothelial cell permeability studies. β-catenin, β-catenin/platelet endothelial cell adhesion molecule-1 interaction, active Src homology 2 domain-containing protein tyrosine phosphatasewere significantly decreased, while β-catenin ubiquitination levels and endothelial permeability were significantly increased, in hyperglycemic retinal endothelial cells. Similar results were observed with platelet endothelial cell adhesion molecule-1 partial knockdown, where β-catenin and active Src homology 2 domain-containing protein tyrosine phosphatase levels were decreased, while phospho-β-catenin and retinal endothelial cell permeability were increased. Platelet endothelial cell adhesion molecule-1 loss may contribute to increased retinal endothelial cell permeability by attenuating β-catenin levels under hyperglycemic conditions.