Abstract
Purpose :
The transplantation of retinal pigment epithelium (RPE) is considered to be one of the primary ways to restore RPE functionality in age-related macular degeneration (AMD). However, it is well-established that the local RPE milieu harbour oxidative stress conditions driven by the dying cells and changes in the molecular components. The right scaffold carrying RPE cells should be able to withstand oxidative stress, enabling the cells to potentially restore normal functionality. Ex-vivo models of aged human Bruch’s membrane (BM) could represent a powerful tool for studying the capability of implanted RPE cells to tackle these pathological conditions.
Methods :
BM explants were obtained from cadaver donor eyes within 48 hours of death. Native RPE cells were gently removed to expose the RPE basal lamina. Pluripotent stem cell-derived RPE (PSC-RPE) cells were initially seeded onto BM explant to evaluate their ability to repopulate the explant surface. PSC-RPE were cultured over either the Descemet’s membrane (biological) or the PLGA scaffold (synthetic) prior to explant cultivation. A comprehensive comparison was conducted among the different scaffold-conditions. Electron microscope analysis provided information on the morphology of the RPE cells resurfacing the scaffolds. Morphometric analysis aimed to enhance differences in shape metrics between the cultures. Immunofluorescence was employed to determine the expression and localization of typical RPE markers. RPE monolayer integrity on the investigated scaffolds was assessed by measuring transepithelial resistance.
Results :
PSC-RPE cells exhibited distinct phenotypes when cultured on the selected scaffolds, thereby supporting the hypothesis that the microenvironment in which the cells reside can influence their characteristics, encompassing cell morphology, polarization, and function. PSC-RPE cells cultivated directly on BM explants failed to form an intact monolayer in areas where deep layers of BM were exposed. In contrast, the two tested scaffolds significantly improved the PSC-RPE culture on the BM, irrespective of the initial condition of the explant.
Conclusions :
This assay offers an ex vivo model of aged BM to evaluate the viability and functional characteristics of PSC-RPE cells when seeded on aged/compromised extracellular matrix. These findings suggest that the ability of the RPE cells to repopulate BM in AMD is improved when the cells are cultivated over a scaffold.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.