Vascular permeability in experimental diabetes is associated with reduced endothelial occludin content: vascular endothelial growth factor decreases occludin in retinal endothelial cells. Penn State Retina Research Group.

Abstract

Blood-retinal barrier (BRB) breakdown is a hallmark of diabetic retinopathy, but the molecular changes that cause this pathology are unclear. Occludin is a transmembrane component of interendothelial tight junctions that may regulate permeability at the BRB. In this study, we examined the effects of vascular endothelial growth factor (VEGF) and diabetes on vascular occludin content and barrier function. Sprague-Dawley rats were made diabetic by intravenous streptozotocin injection, and age-matched animals served as controls. After 3 months, BRB permeability was quantified by intravenous injection of fluorescein isothiocyanate-bovine serum albumin (FITC-BSA), Mr 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 < or = 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 immunoblotting of retinal homogenates. Retinal occludin content decreased approximately 35% (P < or = 0.03) in the diabetic versus the control 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 occludin content 46 and 54%, respectively. These data show that diabetes selectively reduces retinal occludin protein expression and increases BRB permeability. Our findings suggest that the elevated VEGF in the vitreous of patients with diabetic retinopathy increases vascular permeability 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.

PKC-βExacerbatesin vitroBrain Barrier Damage in Hyperglycemic Settings via Regulation of RhoA/Rho-kinase/MLC2 Pathway

“…The reduced expression of TJ protein occludin may shed some light on the HG-induced barrier hyperpermeability and support the previous in vivo studies reporting a direct correlation between the levels of TJ proteins and the extent of endothelial barrier permeability. [26][27][28] The discontinuous appearances of occludin and ZO-1 on cell periphery provide further explanation for the HG-mediated increases in paracellular flux. Consistent with the recent data showing an insulin-mediated elevation in the cerebral contents of occludin and ZO-1, normalization of glucose levels in our study also significantly improved occludin levels and effectively restored occludin and ZO-1 staining to the cell periphery, a set of findings replicated by inhibitions of RhoA or Rho-kinase.…”

“…Consistent with the recent data showing an insulin-mediated elevation in the cerebral contents of occludin and ZO-1, normalization of glucose levels in our study also significantly improved occludin levels and effectively restored occludin and ZO-1 staining to the cell periphery, a set of findings replicated by inhibitions of RhoA or Rho-kinase. [26][27][28] Subsequent studies focusing on the specific impact of PKC-b on cerebral endothelial cell content of TJ proteins have shown that knockdown of PKC-b significantly increased occludin and ZO-1 protein levels under hyperglycemic conditions and selectively that of occludin also under normoglycemic conditions. Decreases in phosphorylation, ubiquitination, and trafficking of occludin may account for the PKC-b-inhibition-mediated improvements observed in occludin protein levels.…”

Vascular endothelial growth factor in eye disease

“…The role of an alteration in endothelial cell junction adhesion in diabetes-induced permeability has been supported by studies showing a decreased level of the tight junction protein occludin in rats rendered diabetic with streptozotocin or treated with intraocular injections of VEGF (Antonetti et al, 1998;Barber et al, 2000). The VEGF-induced hyperpermeability response appears to be associated with phosphorylation of occludin and another junction protein, ZO-1 (Antonetti et al, 1999a).…”

Combination therapy of intravitreal bevacizumab with single simultaneous posterior subtenon triamcinolone acetonide for macular edema due to branch retinal vein occlusion

“…1,2 Although the exact mechanism of ME in BRVO is not fully understood, ME is thought to be caused by the flux of fluid from the blood vessels to the tissue, because of breakdown of the blood-retinal barrier as a result of damage to the tight junctions of capillary endothelial cells, enhanced expression of inflammatory cytokines such as prostaglandins and interleukin 6, and increased secretion into the vitreous of vasopermeability factors such as vascular endothelial growth factor (VEGF) produced in the retina. [3][4][5][6][7] Several recent studies have reported that VEGF has an important role in the pathophysiology of ME by inducing blood-retinal barrier breakdown and increasing vascular permeability. [6][7][8][9] Elevated VEGF level in vitreous has been reported in patients with BRVO, and this elevation was correlated with the severity of ME and area of non-perfusion.…”