Generation of induced pluripotent stem cell-derived anterior foregut endoderms on integrin-binding short peptide-based synthetic substrates

Abstract Anterior foregut endoderms (AFEs) derived from induced pluripotent stem cells (iPSCs) are an important cell source in stem cell technology as they give rise to some important lineages like lung progenitors and thyroid cells. Coating substrates plays a critical role in AFE generation. Currently, conventional large molecule proteins like Matrigel are used in most differentiation protocols. However, the complex components and mechanisms of these coatings have limited both the exploration of cell–extracellular matrix (ECM) interaction and potential clinical applications. In this study, we identified eight pure synthetic integrin-binding short peptides as effective coatings for iPSC growth and AFE generation with an integrin-binding peptide array. Our results showed that integrin α5β1-, αVβ8-, and αIIbβ3-binding peptides supported the adhesion and expansion of iPSCs and AFE generation by guided differentiation via a definitive endoderm (DE) in a full-anchorage-dependent manner. AFE generation was also found on coatings based on integrin α3β1-, α6β1-, αVβ1-, αVβ6-, and αMβ2-binding peptides following a process with temporal suspension growth in the DE-inducing stage, with lower AFE generation efficiency compared to the full-anchorage-dependent peptide groups and Matrigel. According to the results, the integrin α5β1-binding peptide is the most promising defined substrate for inducing AFEs because of its equivalent efficiency with traditional Matrigel coating in supporting iPSC expansion and differentiation toward AFEs. Additionally, the other seven peptide-based coatings also exhibit potential and could be further investigated for developing synthetic-coating strategies in future studies involving AFEs. Our findings provide valuable insights into the role of integrin and ECM function and hold great potential for disease modeling as well as therapeutic exploration of AFE origin organs.