Development of 3D retinal grafts for the treatment of retinal degenerative blindness

Abstract

Purpose : Induced pluripotent stem cell (iPSC)-derived retinal cells hold great promise for restoring vision in patients with retinal degenerative blindness. However, traditional bolus transplantation approaches result in poor cell survival and limited synaptic integration. Tissue engineering scaffold-based strategies, aimed at reducing post-transplant cell death, have potential to overcome these issues. The purpose of this study was to evaluate local and systemic toxicity following transplantation of human iPSC-derived retinal cell grafts in an immune suppressed rat model.

Methods : Polycaprolactone (PCL) scaffolds were generated by injecting a PCL prepolymer solution into a polymerization chamber containing a photomask designed to induce pore formation. Scaffolds were polymerized under UV light, washed and sterilized via gamma irradiation. IPSC derived retinal organoids were generated from 6 patients using a 3D differentiation protocol. Organoids were dissociated and seeded in sterilized PCL scaffolds. Retinal cell grafts were analyzed via immunocytochemistry and 1mm punches were transplanted into the subretinal space of RNU-/- rats (N=210). Full necropsy with complete histopathology, clinical chemistry and hematology were performed at 1-, 3-, and 6-months post-transplantation.

Results : Nine photomasks with 25 to 75μm diameter spots, separated from each other by 25 to 75μm, were designed and fabricated. The photomask with 50μm spots separated by 25μm was ideal for production of consistent PCL scaffolds containing full thickness pores with stable interpore structure. Two weeks after scaffold seeding, iPSC derived photoreceptor precursor cells expressing recoverin were polarized and densely packed in columns throughout the pores, recapitulating the retina. Following subretinal transplantation in RNU-/- rats, retinal reattachment was noted in all eyes, and slow degradation of the biodegradable PCL scaffold was detected via fundoscopy over the duration of the study. No evidence of treatment induced retinal or systemic toxicity/tumorgenicity was detected by gross or histopathologic evaluation, hematology and clinical chemistry.

Conclusions : By using photomasks designed to generate PCL cell delivery scaffolds during photopolymerization, we were able to successfully develop a high-throughput platform for testing of local and systemic toxicity following subretinal transplantation of patient derived retinal cell grafts.

This is a 2020 ARVO Annual Meeting abstract.