Abstract
Purpose: :
Porcine-derived SIS (small intestine submucosa) membrane is an acellular, biocompatible, extracellular matrix sheet that is FDA approved for use in humans. In this study, we examined the ability to grow human induced pluripotent stem cell (hiPSC)-derived retinal cell types on SIS membrane as a platform for future transplantation studies.
Methods: :
hiPSCs were differentiated toward a neural retina or RPE cell fate using our previously described protocols. RPE or optic vesicle-like structures (OVs) were dissociated and plated onto laminin-coated SIS membrane. Human fetal RPE (hfRPE) was also grown on SIS membrane for comparison. RPE and neural retina marker expression was monitored by RT-PCR and immunocytochemistry, and cellular structure and organization was examined by light and electron microscopy (EM).
Results: :
Both dissociated hiPSC-OVs and hiPSC-RPE adhered to laminin-coated SIS membrane. Over the course of several weeks, hfRPE and hiPSC-derived RPE established a pigmented monolayer of cells. RT-PCR confirmed the expression of mature RPE markers, and immunocytochemistry showed the spatially appropriate distribution of RPE proteins. EM analysis suggested the formation of a basement membrane and revealed the presence of many structures typical of RPE including numerous microvilli. EM also demonstrated the porous nature of the SIS membrane and ordered arrangement of collagen fibrils. Dissociated hiPSC-OVs expressed neural retina genes over time. At later time points in culture, the majority of cells were recoverin-positive, indicating the differentiation of photoreceptor-like cells on the SIS membrane.
Conclusions: :
We demonstrate that retinal cell types derived from hiPSCs adhere to SIS membrane, proliferate, and express markers typical of RPE and photoreceptors. Therefore, SIS membrane provides a promising platform for future transplantation and modeling studies.
Keywords: retinal culture • photoreceptors • retinal pigment epithelium