Soy Protein Nanofiber Scaffolds for Uniform Maturation of Human Induced Pluripotent Stem Cell-Derived Retinal Pigment Epithelium

Retinal pigment epithelium (RPE) differentiated from human induced pluripotent stem cells, called induced retinal pigment epithelium (iRPE), is being explored as a cell-based therapy for the treatment of retinal degenerative diseases, especially age-related macular degeneration. The success of RPE implantation is linked to the use of biomimetic scaffolds that simulate Bruch's membrane and promote RPE maturation and integration as a functional tissue. Due to difficulties associated with animal protein-derived scaffolds, including sterility and pro-inflammatory responses, current practices favor the use of synthetic polymers, such as polycaprolactone (PCL), for generating nanofibrous scaffolds. In this study, we tested the hypothesis that plant protein-derived fibrous scaffolds can provide favorable conditions permissive for the maturation of RPE tissue sheets in vitro . Our natural, soy protein-derived nanofibrous scaffolds exhibited a J-shaped stress–strain curve that more closely resembled the mechanical properties of native tissues than PCL with significantly higher hydrophilicity of the natural scaffolds, favoring in vivo implantation. We then demonstrate that iRPE sheets growing on these soy protein scaffolds are equivalent to iRPE monolayers cultured on synthetic PCL nanofibrous scaffolds. Immunohistochemistry demonstrated RPE-like morphology and functionality with appropriate localization of RPE markers RPE65, PMEL17, Ezrin, and ZO1 and with anticipated histotypic polarization of vascular endothelial growth factor and pigment epithelium-derived growth factor as indicated by enzyme-linked immunosorbent assay. Scanning electron microscopy revealed dense microvilli on the cell surface and homogeneous tight junctional contacts between the cells. Finally, comparative transcriptome analysis in conjunction with principal component analysis demonstrated that iRPE on nanofibrous scaffolds, either natural or synthetic, matured more consistently than on nonfibrous substrates. Taken together, our studies suggest that the maturation of cultured iRPE sheets for subsequent clinical applications might benefit from the use of nanofibrous scaffolds generated from natural proteins.