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Title	Targeting the tight junction protein, zonula occludens-1, with the connexin 43 mimetic peptide, αCT-1 reduces VEGF-dependent RPE pathophysiology
Author	Obert, Elizabeth Christa
Date	2015
Affiliation/College	College of Graduate Studies
Department	Neurosciences
Subject.MeSH	Academic Dissertations Academic Dissertations--South Carolina Macular Degeneration Developed Countries Aged Retinal Pigment Epithelium Zonula Occludens-1 Protein Eye Diseases
Keywords	Age-Related Macular Degeneration, Industrialized Country, Elderly, Blood-Retina Barrier, Alpha Connexin carboxy-Terminal 1
Collection	MUSC Theses and Dissertations
Contributing Institution	Waring Historical Library (MUSC)
Rights	Copyright is held by the author.
Language	English
Type	Text
Format	application/pdf
Media Type	Manuscripts
Resource Identifier	mtd2015_obert_ec
Abstract	Age-related macular degeneration (AMD) is a multifactorial disease and is regarded as the predominant cause of central vision loss in the elderly in industrialized countries. A critical target tissue in AMD is the retinal pigment epithelium (RPE), which together with Bruch’s membrane forms the outer blood-retina barrier (BRB). RPE-barrier dysfunction in AMD might result from attenuation and disruption of intercellular tight junctions. Zonula occludens-1 (ZO-1) is a major structural protein of intercellular junctions. A connexin-based peptide mimetic, αCT-1 (Alpha Connexin carboxy-Terminal 1), was developed which competitively inhibits ZO-1 interaction with its binding partners. We hypothesized that targeting ZO-1 signaling using αCT-1 would maintain BRB integrity and reduce RPE pathophysiology by stabilizing gap- and/or tight-junctions. Choroidal neovascularization (CNV) was induced using laser-photocoagulation; RPE-cell barrier loss was triggered by bright light exposure (3000 lux for 3 hours). Both models lead to VEGF- dependent loss of cell junctions. The αCT-1 peptide was delivered via daily eyedrops. CNV lesion sizes were determined using optical coherence tomography (OCT). RPE flatmounts were stained for cell-junction proteins ZO-1 and occludin. Cell profiler software was used to examine the RPE tiling pattern. In vitro experiments using ARPE-19 cells showed that αCT-1 stabilizes intercellular tight junctions. αCT1 treatment reduced CNV development and fluid leakage, and damage was correlated with disruption in cellular integrity of the surrounding RPE cells. Light-damage significantly disrupted RPE cell morphology, which was prevented by αCT-1 pre-treatment. In vitro experiments using ARPE-19 cell monolayers suggest that αCT-1 stabilizes intercellular tight junctions. Taken together, stabilization of cellular junctions with αCT-1 was effective in ameliorating RPE dysfunction in AMD models of photo-coagulation-induced CNV and bright-light exposure RPE-cell barrier loss. Future research will include additional investigation into the peptide’s mechanism of action.

Targeting the tight junction protein, zonula occludens-1, with the connexin 43 mimetic peptide, αCT-1 reduces VEGF-dependent RPE pathophysiology

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