Targeting the tight junction protein, zonula occludens-1, with the connexin43 mimetic peptide, αCT1, reduces VEGF-dependent RPE pathophysiology

Abstract

A critical target tissue in age-related macular degeneration (AMD) is the retinal pigment epithelium (RPE), which forms the outer blood-retina barrier (BRB). RPE-barrier dysfunction might result from attenuation/disruption of intercellular tight junctions. Zonula occludens-1 (ZO-1) is a major structural protein of intercellular junctions. A connexin43-based peptide mimetic, αCT1 was developed to competitively block interactions at the PDZ2 domain of ZO-1, thereby inhibiting ligands that selectively bind to this domain. We hypothesized that targeting ZO-1 signaling using αCT1 would maintain BRB integrity and reduce RPE pathophysiology by stabilizing gap- and/or tight-junctions. RPE-cell barrier dysfunction was generated in mice using laser-photocoagulation triggering choroidal neovascularization (CNV), or bright-light exposure leading to morphological damage. αCT1 was delivered via eyedrops. αCT1 treatment reduced CNV development and fluid leakage as determined by optical coherence tomography, and damage was correlated with disruption in cellular integrity of surrounding RPE cells. Light-damage significantly disrupted RPE cell morphology as determined by ZO-1 and Occludin staining and tiling pattern analysis, which was prevented by αCT1 pre-treatment. In vitro experiments using RPE and MDCK monolayers indicated that αCT1 stabilizes tight junctions, independent of its effects on Cx43. Taken together, stabilization of intercellular junctions by αCT1 was effective in ameliorating RPE dysfunction in models of AMD-like pathology.

Low molecular weight heparin (nadroparin) improves placental permeability in rats with gestational diabetes mellitus via reduction of tight junction factors

“…research into the blood-brain barrier has revealed that the expression of VeGF is upregulated by hypoxia, thereby increasing blood-brain barrier permeability (33). in addition, blood-retinal barrier dysfunction has been recovered by targeting Zo-1 through a reduction in VeGF (38). Hypoxia-induced TJ dysfunction in trophoblasts can be improved by downregulation of VeGF (26).…”

Extracellular vesicle-mediated long-range communication in stressed retinal pigment epithelial cell monolayers

“…Staining was examined by fluorescence microscopy (Zeiss, Thornwood, NY) equipped with a digital black-and-white camera (Spot camera; Diagnostic Instruments, Sterling Heights, MI) [34]. …”

Delivery of CR2-fH Using AAV Vector Therapy as Treatment Strategy in the Mouse Model of Choroidal Neovascularization

“…The CellProfiler v.2.11 software ( http://www.cellprofiler.org/ ) was used to evaluate RPE morphology as previously described. 58 TIF files of images of equal size and exposure time were first imported into the software and analyzed using a customizable script. Cells were compared using the pipeline neighboring cells from which the form factor (in which a perfectly circular object equals 1) and eccentricity (the degree, measured between 0 and 1, to which an object represents an ellipse) were obtained.…”