1. Characterization of Human iPSC-RPE on a Prosthetic Bruch's Membrane Manufactured From Silk Fibroin.
- Author
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Galloway CA, Dalvi S, Shadforth AMA, Suzuki S, Wilson M, Kuai D, Hashim A, MacDonald LA, Gamm DM, Harkin DG, and Singh R
- Subjects
- Blotting, Western, Cell Differentiation, Cells, Cultured, Coated Materials, Biocompatible, Extracellular Matrix Proteins metabolism, Gene Expression Regulation physiology, Humans, Immunohistochemistry, Induced Pluripotent Stem Cells metabolism, Real-Time Polymerase Chain Reaction, Retinal Pigment Epithelium metabolism, Tissue Scaffolds, Bruch Membrane, Fibroins chemistry, Induced Pluripotent Stem Cells cytology, Membranes, Artificial, Retinal Pigment Epithelium cytology
- Abstract
Purpose: RPE cell transplantation as a potential treatment for AMD has been extensively investigated; however, in AMD, ultrastructural damage affects both the RPE and its underlying matrix support, the Bruch's membrane (BrM). An RPE monolayer supported by a surrogate scaffold could thus provide a more effective approach to cell-based therapy for AMD. Toward this goal, we aimed to establish a functional human induced pluripotent stem cell-derived (hiPSC)-RPE monolayer on a Bombyx mori silk fibroin (BMSF) scaffold., Methods: RPE differentiated from five distinct hiPSC lines were cultured on BMSF membrane coated with extracellular matrix (ECM, COL1), and either regular tissue culture plastic or Transwell coated with ECM (LAM-TCP). Morphologic, gene and protein expression, and functional characteristics of the hiPSC-RPE cultured on different membranes were compared in longitudinal experiments spanning 1 day to ≥3 months., Results: The hiPSC-RPE monolayers on ECM-coated BMSF and TCP could be maintained in culture for ≥3 months and displayed RPE-characteristic morphology, pigmentation, polarity, and expression of RPE signature genes and proteins. Furthermore, hiPSC-RPE on both ECM-coated BMSF and TCP displayed robust expression and secretion of several basement membrane proteins. Importantly, hiPSC-RPE cells on COL1-BMSF and LAM-TCP showed similar efficacy in the phagocytosis and degradation of photoreceptor outer segments., Conclusions: A biomaterial scaffold manufactured from silk fibroin supports the maturation and long-term survival of a functional hiPSC-RPE monolayer. This has significant implications for both in vitro disease modeling and in vivo cell replacement therapy.
- Published
- 2018
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