Autologous iris pigment epithelium cultured on cellulose acetate membrane transplanted into the subretinal space of rabbits

Abstract

Abstract: : Purpose:Pilot clinical trials have shown encouraging results transplanting cell suspensions of autologous retinal pigment epithelium (RPE) or iris pigment epithelium (IPE) after removal of choroidal neovascularization in patients with age–related macular degeneration. Limitations include permanent damage in Bruch’s membrane and the requirement that the transplanted cells form a monolayer in the subretinal space covering the defect in the native RPE. We have investigated the use of porous cellulose acetate membranes as a substrate for transplanted cells and a replacement for Bruch’s membrane. Methods: Cellulose acetate with a molecular weight cut–off of 100 kD was selected due to its superior support of cultured cells in vitro. The thickness of the membrane was altered with the use of a 193 nanometer excimer laser, resulting in a final thickness of approximately 30 microns. Autologous IPE cells were harvested by a simple iridectomy from New Zealand red rabbits and seeded on the cellulose acetate membranes. The seeded membranes were cultured in IPE medium until a confluent monolayer was achieved. In order to implant the membrane, an implantation instrument was designed to allow insertion through a sclerostomy and eliminate intraoperative handling. Implantation was achieved through a standard 3–port pars plana vitrectomy. A retinal bleb was raised and the membrane was inserted through a 1.5 mm retinotomy. The retina was then flattened with the use of perflurocarbon heavy fluid. A perflurocarbon–silicon oil exchange was then performed to ensure postoperative retinal tamponade. The rabbits were euthanased at one week and one month and the eyes were processed for light and electron microscope analysis. Results: The cellulose acetate membranes seeded with IPE cells were easily inserted into the subretinal space with the use of the implantation device. Silicon oil resulted in consistent successful retinal attachment. A uniform confluent monolayer was achieved on the cellulose acetate membrane prior to implantation and this was maintained at one week after implantation into the subretinal space.The membrane was well tolerated in the subretinal space with preservation of the overlying retinal morphology. Conclusion: We have successfully cultured autologous IPE on 30 micron–thick cellulose acetate membranes and developed an implantation instrument to implant the substrate and cells into the subretinal space of rabbits. We have demonstrated biocompatibility and stability of these implants, which raises the possibility of clinical applications.

Keywords: age–related macular degeneration • retinal pigment epithelium • transplantation