Ultrathin micro-molded biodegradable scaffolds for outer retina reconstruction

Abstract

Purpose : Cell-based therapies have the potential to treat blinding retinal disorders caused by photoreceptor (PR) and/or retinal pigment epithelium (RPE) cell death. Recent advances in biomaterials and tissue engineering have produced scaffold-based strategies for PR or RPE replacement, but organized approaches supporting both cell types—which many later-stage outer retinal disease patients will likely require—are lacking. We hypothesized that biodegradable scaffolds could meet this need. Herein, we describe an ultrathin biodegradable scaffold capable of supporting organized, high-density capture of human pluripotent stem cell (hPSC)-derived PRs and/or RPE.

Methods : Micro-fabricated polyglycerol sebacate (PGS) “ice cube tray” scaffolds were generated, mounted in transwells, and gas sterilized for outer retinal cell culture. hPSC-derived retinal organoids and/or RPE were generated, dissociated, and plated onto laminin-coated scaffolds to determine optimum cell seeding density and culture conditions for PRs, RPE, or PRs+RPE. Scaffolds were fixed, immunostained, and imaged on a Nikon A1R confocal microscope. Images were analyzed in NIS-Elements and statistical analyses were performed in GraphPad Prism.

Results : The “ice cube tray” PGS scaffold design maximized cell-to-biomaterial payload, supporting 15-20 PRs per scaffold well, or approximately 18,000 cells/mm² for PR scaffolds. At an optimal seeding density (5 million cells per transwell), rod and cone PRs on scaffolds matured to express outer segment markers and presynaptic proteins organized near the top of the scaffold. The micro-patterned surface also supported RPE growth, with a polarized monolayer forming at higher (300K-400K RPE/transwell) but not at lower seeding densities. Scaffold degradation in vitro only occurred when seeded with RPE, peaking at 2 months in culture. RPE-seeded scaffolds were filled almost entirely with RPE, however, small pockets within wells remained available for seeding a sequential PR layer.

Conclusions : PGS “ice cube tray” scaffolds facilitate robust high-density human stem cell-derived PR or RPE growth. Scaffolds degrade in the presence of RPE, highlighting a need for balance between cell maturity and scaffold integrity for biodegradable cell replacement constructs. This study presents proof-of-concept for scaffold-supported growth of organized PRs and RPE, critical to future applications in transplantation for outer retinal diseases.

This is a 2020 ARVO Annual Meeting abstract.