Retinal pigment epithelium cells protect neuroretina from spontaneous degenerative modifications in a porcine co-culture model

Abstract

Purpose: To evaluate the protective role of the retinal pigment epithelium (RPE) cells against neuroretinal degeneration, in a novel co-culture model of central porcine neuroretina supplemented with RPE cells for closely simulating the subretinal space after physical separation of the neural retina and RPE as occurs in retinal detachment and other conditions

Methods: RPE cells were isolated from porcine eyes, expanded, and seeded on the bottom of Transwell^® plates. Neuroretina explants were obtained from the porcine area centralis and cultured alone (controls) on Transwell^® membranes (TM) or supplemented with RPE cells in the same wells but physically separated by the TM. Finally, cellular and tissue specimens were processed for phase contrast, cytological (H&E), and immunochemical evaluation. Antibodies against retinal pigment epithelium-specific 65 kDa (RPE65), cellular retinaldehyde-binding protein (CRALBP), rhodopsin (RHO), calbindin D-28K (CB) and glial fibrillary acidic protein (GFAP) were used. Pigment epithelium-derived factor (PEDF) concentration in the culture medium was also determined by ELISA

Results: RPE cells after 9 days of co-culture were confluent and formed monolayers with polygonal-shaped cells, pigment, and RPE65 immunoexpression. Neuroretinas co-cultured with RPE cells better maintained tissue structure and cellular organization, preserved photoreceptors (CB and RHO), and reduced reactive gliosis (GFAP and CRALBP) compared to controls. PEDF levels were significantly greater in co-cultures (p<0.05)

Conclusions: RPE cells preserved neuroretina from spontaneous degenerative changes in a co-culture model of central porcine neuroretina and RPE cells. These results were probably linked to RPE cell secretion of neuroprotective factors such as PEDF. The ex vivo subretinal space model utilized may be useful to study neuroretina and RPE cell “cross-talk” and to pre-clinically test potential therapies for retinal degenerative diseases