Biological and synthetic membranes for human induced pluripotent stem cell-based transplantation therapy

Abstract

Purpose: Scaffolds may improve transplantation therapy by providing an organized platform for cell delivery. In this study, we evaluated two membranes for human induced pluripotent stem cell (hiPSC)-based transplantation therapy, an acellular biological membrane derived from porcine small intestine submucosa (SIS), as well as a synthetic micro-patterned membrane (SU-8).

Methods: SIS membrane (Cook Biotech), a porous extracellular matrix sheet roughly 15µm thick, was cultured using Snapwell transwell inserts (Corning). SU-8 membranes were generated with standard photolithography techniques, resulting in free-standing porous microstructures that were 8µm thick with 5µm pores. Both membranes were pretreated with laminin, and then coated with hiPSC-derived RPE or hiPSC-derived neural retina (NR) cells. Cellular gene and protein expression was monitored with RT-PCR and immunocytochemistry. Cell growth, viability, and polarity were analyzed by standard light and confocal microscopy. Subretinal insertion of cell-impregnated membranes (1mm in diameter) into the Long Evans rat retina was performed using a patented transplantation device (Eyevation). Human cells within the rat retina were identified with a human cytoplasm specific antibody (Stem Cell Inc.).

Results: Both membranes promoted cell adhesion and growth in vitro. hiPSC-RPE formed polarized, pigmented monolayers and expressed mature markers of RPE. hiPSC-NR cells, including numerous RCVRN+ photoreceptors, grew in layers on both membranes and expressed characteristic NR markers. Following transplantation, the SIS membrane was generally well tolerated by the host rat retina, although Muller glia became activated. Both hiPSC-NR cells and hiPSC-RPE transplanted on the SIS membrane survived for up to one month in vivo, the latest time point examined. The SU-8 membrane has also been successfully transplanted and analysis is currently underway.

Conclusions: We demonstrate that both synthetic and biological scaffolds permit cell adherence, growth, and differentiation of hiPSC-RPE and NR in vitro. Furthermore, both can be transplanted into the rat retina. These scaffolds provide an organized structure for hPSC-based cellular transplantation and may also improve in vitro modeling.

Keywords: 721 stem cells • 688 retina