A novel rabbit model for studying RPE transplantation

¹ Department of Ophthalmology, Second Affiliated Hospital of Harbin Medical University, Harbin, China
² Ophthalmology, Second Affiliated Hospital of Harbin Medical University, Harbin, China
³ Dept. Surgery/Anatomy, Yale University School of Medicine, New Haven, Connecticut, United States
⁴ Ophthalmology, Second Affiliated Hospital of Harbin Medical University, Harbin, China; Surgery and Ophthalmology, Yale University School of Medicine, New Haven, Connecticut, United States

	Abstract

Purpose. The goal of this project is to develop a model of retinal pigment epithelium (RPE) transplantation that permits extensive and reliable analysis of the transplants. Methods. Cultures of newborn rabbit RPE were evaluated by morphology, electrophysiology and the expression of zonula occludens-1, cytokeratin and a melanocyte marker (S-100). Cells labeled with 5,6-carboxyfluorescein diacetate succinimidyl ester (CFDA-SE) were transplanted into the subretinal space of rabbits using a 30 gauge needle without making a conjunctival flap or sclerotomy. The transplants were examined by fundus photography, confocal scanning laser ophthalmoscopy (cSLO), optical coherence tomography (OCT) and angiography. At two months the retina was examined histochemically. Results. A one minute incubation at 37°C with 20µM CFDA-SE did not affect morphology or the expression of marker proteins. In co-culture, the labeled cells integrated into monolayers that developed a normal transepithelial electrical resistance of 400-450 xcm². Dye was not transferred from labeled to non-labeled RPE cells. Transplanted RPE was detectable for at least 2 months. Angiography demonstrated an intact blood retinal barrier. The normal morphology of the retina and lack of debris in the subretinal space, suggested the transplanted RPE was functional. Conclusions. Primary cultures of newborn rabbit RPE were highly differentiated even when labeled with CFDA-SE. Labeled cells could be followed long-term in vitro and in vivo. This model can examine how culture and transplantation protocols affect the reformation of a functional RPE monolayer. The similar size of rabbit and human eyes will facilitate the translation of these protocols to the bedside.

Key Words: retinal pigment epithelium, retinal transplantation, cell culture, blood-retinal barrier, CFDA_SE, cell label

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