Dataset Information

A Roadmap for Human Liver Differentiation from Pluripotent Stem Cells.

ABSTRACT: How are closely related lineages, including liver, pancreas, and intestines, diversified from a common endodermal origin? Here, we apply principles learned from developmental biology to rapidly reconstitute liver progenitors from human pluripotent stem cells (hPSCs). Mapping the formation of multiple endodermal lineages revealed how alternate endodermal fates (e.g., pancreas and intestines) are restricted during liver commitment. Human liver fate was encoded by combinations of inductive and repressive extracellular signals at different doses. However, these signaling combinations were temporally re-interpreted: cellular competence to respond to retinoid, WNT, TGF-?, and other signals sharply changed within 24 hr. Consequently, temporally dynamic manipulation of extracellular signals was imperative to suppress the production of unwanted cell fates across six consecutive developmental junctures. This efficiently generated 94.1% ± 7.35% TBX3+HNF4A+ human liver bud progenitors and 81.5% ± 3.2% FAH+ hepatocyte-like cells by days 6 and 18 of hPSC differentiation, respectively; the latter improved short-term survival in the Fah-/-Rag2-/-Il2rg-/- mouse model of liver failure.

SUBMITTER: Ang LT

PROVIDER: S-EPMC5854481 | biostudies-literature | 2018 Feb

REPOSITORIES: biostudies-literature

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A Roadmap for Human Liver Differentiation from Pluripotent Stem Cells.

Ang Lay Teng LT Tan Antson Kiat Yee AKY Autio Matias I MI Goh Su Hua SH Choo Siew Hua SH Lee Kian Leong KL Tan Jianmin J Pan Bangfen B Lee Jane Jia Hui JJH Lum Jen Jen JJ Lim Christina Ying Yan CYY Yeo Isabelle Kai Xin IKX Wong Chloe Jin Yee CJY Liu Min M Oh Jueween Ling Li JLL Chia Cheryl Pei Lynn CPL Loh Chet Hong CH Chen Angela A Chen Qingfeng Q Weissman Irving L IL Loh Kyle M KM Lim Bing B

Cell reports 20180201 8

How are closely related lineages, including liver, pancreas, and intestines, diversified from a common endodermal origin? Here, we apply principles learned from developmental biology to rapidly reconstitute liver progenitors from human pluripotent stem cells (hPSCs). Mapping the formation of multiple endodermal lineages revealed how alternate endodermal fates (e.g., pancreas and intestines) are restricted during liver commitment. Human liver fate was encoded by combinations of inductive and repr ...[more]

PMID: 29466743

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Project description:Efficient and effective methods for converting human induced pluripotent stem cells into differentiated derivatives are critical for performing robust, large-scale studies of development and disease modelling, and for providing a source of cells for regenerative medicine. Here, we describe a 14-day neural differentiation protocol which allows for the scalable, simultaneous differentiation of multiple iPSC lines into cortical neural stem cells We currently employ this protocol to differentiate and compare sets of engineered iPSC lines carrying loss of function alleles in developmental disorder associated genes, alongside isogenic wildtype controls. Using RNA sequencing (RNA-Seq), we can examine the changes in gene expression brought about by each disease gene knockout, to determine its impact on neural development and explore mechanisms of disease. The 10-day Neural Induction period uses the well established dual-SMAD inhibition approach combined with Wnt/β-Catenin inhibition to selectively induce formation of cortical NSCs. This is followed by a 4-day Neural Maintenance period facilitating NSC expansion and rosette formation, and NSC cryopreservation. We also describe methods for thawing and passaging the cryopreserved NSCs, which are useful in confirming their viability for further culture. Routine implementation of immunocytochemistry Quality Control confirms the presence of PAX6-positive and/or FOXG1-positive NSCs and the absence of OCT4-positive iPSCs after differentiation. RNA-Seq, flow cytometry, immunocytochemistry (ICC) and RT-qPCR provide additional confirmation of robust presence of NSC markers in the differentiated cells. The broader utility and application of our protocol is demonstrated by the successful differentiation of wildtype iPSC lines from five additional independent donors. This paper thereby describes an efficient method for the production of large numbers of high purity cortical NSCs, which are widely applicable for downstream research into developmental mechanisms, further differentiation into postmitotic cortical neurons, or other applications such as large-scale drug screening experiments.

Dataset Information

A Roadmap for Human Liver Differentiation from Pluripotent Stem Cells.

Publications

A Roadmap for Human Liver Differentiation from Pluripotent Stem Cells.

Similar Datasets

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