Generation of pancreatic progenitors from human pluripotent stem cells by small molecules.

Yuqian Jiang,Chuanxin Chen,Xiaojun Lance Lian,Lauren N Randolph,Xiaoping Bao,Xin Zhang,Songtao Ye

doi:10.1016/j.stemcr.2021.07.021

Yuqian Jiang, Chuanxin Chen + Show 5 more

Open Access

https://doi.org/10.1016/j.stemcr.2021.07.021

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Abstract

SummaryHuman pluripotent stem cell (hPSC)-derived pancreatic progenitors (PPs) provide promising cell therapies for type 1 diabetes. Current PP differentiation requires a high amount of Activin A during the definitive endoderm (DE) stage, making it economically difficult for commercial ventures. Here we identify a dose-dependent role for Wnt signaling in controlling DE differentiation without Activin A. While high-level Wnt activation induces mesodermal formation, low-level Wnt activation by a small-molecule inhibitor of glycogen synthase kinase 3 is sufficient for DE differentiation, yielding SOX17+FOXA2+ DE cells. BMP inhibition further enhances this DE differentiation, generating over 87% DE cells. These DE cells could be further differentiated into PPs and functional β cells. RNA-sequencing analysis of PP differentiation from hPSCs revealed expected transcriptome dynamics and new gene regulators during our small-molecule PP differentiation protocol. Overall, we established a robust growth-factor-free protocol for generating DE and PP cells, facilitating scalable production of pancreatic cells for regenerative applications.

Highlights

Human pluripotent stem cells, including human embryonic stem cells (Thomson et al, 1998) and human induced pluripotent stem cells (Takahashi and Yamanaka, 2006; Yu et al, 2007), can proliferate virtually indefinitely while maintaining the capacity to differentiate to a broad diversity of cell types
We demonstrate that Wnt activation coupled with inhibition of bone morphogenetic protein (BMP) signaling further enhances definitive endoderm (DE) differentiation, yielding over 85% SOX17+FOXA2+ DE cells in 4 days
Differential activation of Wnt pathway leads to distinct cell fates in Human pluripotent stem cell (hPSC) To probe how the differentiation trajectories can be affected by differential activation of the canonical Wnt pathway, we treated a SOX17-mCherry knockin H9 line (Ng et al, 2016) with a glycogen synthase kinase 3 (GSK3) inhibitor, CHIR99021 (CH), at different concentrations in RPMI medium for 24 h (Figure 1A)

Summary

Introduction

Human pluripotent stem cells (hPSCs), including human embryonic stem cells (hESCs) (Thomson et al, 1998) and human induced pluripotent stem cells (iPSCs) (Takahashi and Yamanaka, 2006; Yu et al, 2007), can proliferate virtually indefinitely while maintaining the capacity to differentiate to a broad diversity of cell types Because of these two unique properties, hPSCs are widely used as an in vitro model to study human development (Murry and Keller, 2008) and appreciatory cell sources for cell-based therapies (Randolph et al, 2017). Using a hPSC differentiation model, researchers discovered that definitive endoderm (DE) differentiation of hPSCs is induced by high-dose Activin A treatment for multiple days (D’Amour et al, 2005; Xu et al, 2011)

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Discussion

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