Subretinal integration of a vascular endothelial growth factor slow-release gelatin functionalized cell carrier for retinal pigment epithelium cell therapy

PURPOSE: Replacement of retinal pigment epithelium (RPE) with a cell-based therapeutic represents a potential treatment strategy for age-related macular degeneration (AMD). To improve subretinal integration via angiogenesis, RPE cell carrier transplants were equipped with gelatin for vascular endothelial growth factor (VEGF) release after implantation. METHODS: Gelatin coatings were applied to the basal side of polyethylene terephthalate (PET) membranes using doctor blading, dip coating, or pipetting to optimize film thickness and reproducibility. Human fetal RPE (hfRPE) cells were cultured on treated PET and monitored for morphology, pigmentation, transepithelial resistance, polarization, and expression of RPE-relevant proteins by immunofluorescence. PET/gelatin loaded w/or w/o VEGF, or PET/gelatin/VEGF carriers with hfRPE were subretinally implanted for 6 weeks into pigmented rabbits onto RPE wounds. Subretinal integration was assessed using optical coherence tomography (OCT), fluorescein angiography (FA), indocyanine green angiography (ICGA), and histological and immunofluorescence staining. RESULTS: Reproducible gelatin films were formed by pipetting a defined volume of solution, depending on concentration and gelatin type. In vitro, gelatin coating promoted hfRPE cell growth, monolayer formation, pigmentation, and tight junction development. Polarized secretion of VEGF and pigment epithelium-derived factor (PEDF) was maintained and bestrophin, ezrin and RPE65 showed a qualitatively more homogeneous distribution in gelatin treated hfRPE cells compared to controls. Six weeks after implantation, the hfRPE layer appeared largely intact as judged by a double layered OCT hyperreflectivity, and vascular channel-like structures were observed in the subimplant space. CONCLUSIONS: Integration of RPE grafts into the subretinal space is promoted by PET carriers coated with VEGF-loaded gelatin.