Optimal Hypoxia Regulates Human iPSC-Derived Liver Bud Differentiation through Intercellular TGFB Signaling.

Hiroaki Ayabe,Tomoya Sato,Autumn Ferguson,Barbara Treutlein,J Gray Camp,Takuo Kamoya,Takanori Takebe,Masaki Kimura,Emi Yoshizawa,Takahisa Anada,Hideki Taniguchi,Keisuke Sekine,Shunyuu Kikuchi,Osamu Suzuki,Yasuharu Ueno

doi:10.1016/j.stemcr.2018.06.015

Abstract

SummaryTimely controlled oxygen (O2) delivery is crucial for the developing liver. However, the influence of O2 on intercellular communication during hepatogenesis is unclear. Using a human induced pluripotent stem cell-derived liver bud (hiPSC-LB) model, we found hypoxia induced with an O2-permeable plate promoted hepatic differentiation accompanied by TGFB1 and TGFB3 suppression. Conversely, extensive hypoxia generated with an O2-non-permeable plate elevated TGFBs and cholangiocyte marker expression. Single-cell RNA sequencing revealed that TGFB1 and TGFB3 are primarily expressed in the human liver mesenchyme and endothelium similar to in the hiPSC-LBs. Stromal cell-specific RNA interferences indicated the importance of TGFB signaling for hepatocytic differentiation in hiPSC-LB. Consistently, during mouse liver development, the Hif1a-mediated developmental hypoxic response is positively correlated with TGFB1 expression. These data provide insights into the mechanism that hypoxia-stimulated signals in mesenchyme and endothelium, likely through TGFB1, promote hepatoblast differentiation prior to fetal circulation establishment.

Highlights

Organoid technology using human pluripotent stem cells or isolated organ progenitors is rapidly evolving to model the elaborate spatiotemporal processes of development and regeneration
All of the hypoxic culture conditions assessed in this study are summarized in Figure S1A.The most important feature of this plate is that it provides a direct O2 supply to the cells because the plate is made of PDMS that is only permeable to gas (Anada et al, 2012), promoting the differentiation and survival of 3-D cultured mesenchymal stem cells (MSCs) (Anada et al, 2016; Kamoya et al, 2016)
Dissolved O2 modulated with Excess-hypoxia condition (20% O2, O2-non-permeable plate) had the lowest (10%) O2 levels under all conditions during the first 3 to 4 days and the O2 tension gradually increased

Summary

Introduction

Organoid technology using human pluripotent stem cells or isolated organ progenitors is rapidly evolving to model the elaborate spatiotemporal processes of development and regeneration. Progenitors are directed to self-organize into an organoid comprising multiple cell types, including epithelial and mesenchymal cell lineages (Lancaster and Knoblich, 2014). A human induced pluripotent stem cell-derived liver bud (hiPSC-LB) can be generated by co-culture with hiPSCderived liver progenitor cells, human umbilical vein endothelial cells (HUVECs), and human mesenchymal stem cells (MSCs) (Takebe et al, 2013 and 2017). The complex self-organized hiPSC-LB is an organ-like 3D tissue with cell-cell interactions that recapitulate early LB structures and transcriptomic signatures (Camp et al, 2017). In vitro organoids generally remain developmentally immature without transplantation into animals (Dye et al, 2016; Xinaris et al, 2012; Watson et al, 2014), highlighting the need for filling the maturation gaps between in vitro and in vivo systems

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