Extracellular matrix proteins refine microenvironments for pancreatic organogenesis from induced pluripotent stem cell differentiation

Abstract

Rationale: The current understanding on manipulating signaling pathways to generate mature human islet organoids with all major hormone-secreting endocrine cell types (i.e., α, β, δ, and γ cells) from induced pluripotent stem cells (iPSCs) is insufficient. However, donor islet shortage necessitates that we produce functional islets in vitro. In this study, we aimed to find decellularized pancreatic extracellular matrix (dpECM) proteins that leverage signaling pathways and promote functional iPSC islet organogenesis. Methods: We performed proteomic analysis to identify key islet promoting factors from porcine and rat dpECM. With this, we identified collagen type II (COL2) as a potential biomaterial cue that endorses islet development from iPSCs. Using global transcriptome profiling, gene set enrichment analysis, immunofluorescence microscopy, flow cytometry, Western blot, and glucose-stimulated hormonal secretion analysis, we examined COL2’s role in regulating iPSC pancreatic lineage specification and signaling pathways, critical to islet organogenesis and morphogenesis. Results: We discovered COL2 acts as a functional biomaterial that augments islet development from iPSCs, similar to collagen type V (COL5) as reported in our earlier study. COL2 substantially stimulates the formation of endocrine progenitors and subsequent islet organoids with significantly elevated expressions of pancreatic signature genes and proteins. Furthermore, it enhances islets’ glucose sensitivity for hormonal secretion. A cluster of gene expressions associated with various signaling pathways, including but not limited to oxidative phosphorylation, insulin secretion, cell cycle, the canonical WNT, hypoxia, and interferon-γ response, were considerably affected by COL2 and COL5 cues. Conclusion: We demonstrated dpECM’s crucial role in refining stem cell differentiation microenvironments for organoid development and maturation. Our findings on biomaterial-stimulated signaling for stem cell specification, organogenesis, and maturation open up a new way to increase the differentiation efficacy of endocrine tissues that can contribute to the production of biologically functional islets.