A Roadmap for Human Liver Differentiation from Pluripotent Stem Cells

Ang, Lay Teng; Tan, Antson Kiat Yee; Autio, Matias; Goh, Su Hua; Choo, Siew Hua; Lee, Kian Leong; Tan, Ji; Pan, Bangfen; Lee, Jane Jia Hui; Lum, Jen Jen; Lim, Christina Ying Yan; Yeo, Isabelle Kai Xin; Wong, Chloe Jin Yee; Liu, Min; Oh, Jueween Ling Li; Chia, Cheryl Pei Lynn; Loh, Chet Hong; Chen, Angela; Chen, Yi‐Ping Phoebe; Weissman, Irving L.; Loh, Kyle M.; Lim, Bing

doi:10.1016/j.celrep.2018.01.087

Cited by 174 publications

(187 citation statements)

References 86 publications

(162 reference statements)

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“…Is there only one "posterior foregut" route to generate pancreas, or are there multiple such routes, and if so, does the choice of intermediate route impact downstream pancreatic differentiation? Indeed, hPSC differentiation studies have suggested the existence of at least two types of posterior foregut, one poised for pancreatic, and the other primed for liver, differentiation in vitro (Ang et al, 2018). Third, while hPSC differentiation protocols endeavor to generate early Pdx1 + pancreatic progenitors, there is no singular "pancreatic progenitor" at this stage in vivo.…”

Section: Endodermmentioning

confidence: 99%

A critical look: Challenges in differentiating human pluripotent stem cells into desired cell types and organoids

Fowler

Ang

Loh

2019

WIREs Developmental Biology

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Too many choices can be problematic. This is certainly the case for human pluripotent stem cells (hPSCs): they harbor the potential to differentiate into hundreds of cell types; yet it is highly challenging to exclusively differentiate hPSCs into a single desired cell type. This review focuses on unresolved and fundamental questions regarding hPSC differentiation and critiquing the identity and purity of the resultant cell populations. These are timely issues in view of the fact that hPSC‐derived cell populations have or are being transplanted into patients in over 30 ongoing clinical trials. While many in vitro differentiation protocols purport to “mimic development,” the exact number and identity of intermediate steps that a pluripotent cell takes to differentiate into a given cell type in vivo remains largely unknown. Consequently, most differentiation efforts inevitably generate a heterogeneous cellular population, as revealed by single‐cell RNA‐sequencing and other analyses. The presence of unwanted cell types in differentiated hPSC populations does not portend well for transplantation therapies. This provides an impetus to precisely control differentiation to desired ends—for instance, by logically blocking the formation of unwanted cell types or by overexpressing lineage‐specifying transcription factors—or by harnessing technologies to selectively purify desired cell types. Conversely, approaches to differentiate three‐dimensional “organoids” from hPSCs intentionally generate heterogeneous cell populations. While this is intended to mimic the rich cellular diversity of developing tissues, whether all such organoids are spatially organized in a manner akin to native organs (and thus, whether they fully qualify as organoids) remains to be fully resolved. This article is categorized under: Adult Stem Cells > Tissue Renewal > Regeneration: Stem Cell Differentiation and Reversion Gene Expression > Transcriptional Hierarchies: Cellular Differentiation Early Embryonic Development: Gastrulation and Neurulation

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Section: Endodermmentioning

confidence: 99%

A critical look: Challenges in differentiating human pluripotent stem cells into desired cell types and organoids

Fowler

Ang

Loh

2019

WIREs Developmental Biology

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“…With our interest in characterizing multiple different iPSC-Heps, we tested and found significant differentiation variability and cell death during endoderm induction using popular published protocols (Ang et al, 2018;Ma et al, 2013;Si-Tayeb et al, 2010). In order to better optimize our system, we selected one Matrigel lot for all experiments based on its ability to somewhat support our induction of our weakest iPSC lines; however, even with this Matrigel lot, we did not see efficient differentiation in such lines ( Figure 1C).…”

Section: Induction Of Multiple Ipsc Lines From Different Patients Usimentioning

confidence: 99%

“…Our protocol, for convenience designated ML1 (Figure 1D), showed efficient induction of endoderm. More importantly the ML1 protocol allowed induction of complete monolayers of endoderm from nearly all iPSC lines tested, even those that failed other protocols recommended for poorly differentiating iPSC (Ang et al, 2018;Si-Tayeb et al, 2010).…”

Section: Figure 4)mentioning

confidence: 99%

Doxycycline Significantly Enhances Induction of iPSCs to Endoderm by Enhancing survival via AKT Phosphorylation

Esteva‐Font

Peaslee

et al. 2020

Preprint

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Induced pluripotent stem cells (iPSCs) provide an important tool for the generation patientderived cells including hepatocyte-like cells via developmental cues through an endoderm intermediate. However, most iPSCs fail to differentiate into endoderm, with induction resulting in apoptosis. To address this issue, we built upon published methods to develop an improved protocol with our discovery that doxycycline dramatically enhances the iPSC to endoderm differentiation efficiency by inhibiting apoptosis and promoting proliferation via the AKT pathway.We tested this new protocol in more than 70 iPSC lines with consistent formation of complete sheets of endoderm in 90%. Endoderm generated by our method achieves similar transcriptomic profiles, including FOXA2, HNF1, CXCR4, and SOX17 positive cells, and the ability to be further differentiated. Furthermore this method achieves a four-fold increase in endoderm cell number and will accelerate studies of human diseases in vitro and facilitate the expansion of iPSC-derived cells for transplantation studies.

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“…Human pluripotent stem cells (hPSCs), embryonic stem cells (ESCs) [5], and induced pluripotent stem cells (iPSCs) [6] all can differentiate into almost any kind of cell and possess the ability to self-renew indefinitely. Previous researchers have successfully used growth factors and other chemicals to induce differentiation of hPSCs to hPSC-HLCs [7][8][9][10][11], but these studies did not achieve mature hepatic function of hPSC-HLCs in vitro. The results of these studies clearly demonstrate the need to develop new methods to obtain in vitro hepatic function of hPSC-HLCs.…”

Section: Hepatocyte-like Cells Derived From Human Pluripotent Stem Cementioning

confidence: 99%

Heat treatment functionalizes hepatocyte-like cells derived from human embryonic stem cells

Imamura

Yoshimoto

Terada

et al. 2020

Preprint

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Hepatocyte-like cells derived from human pluripotent stem cells (hPSC-HLCs) may offer an alternative to primary hepatocytes that are commonly used for drug screenings and toxicity tests. Although tremendous efforts have been made to facilitate hepatic functions of hPSC-HLCs using growth factors and chemicals, these cells have not yet reached hepatic functions that are comparable to primary hepatocytes. Therefore, there exists a critical need to use an alternative trigger to facilitate hepatic functions in hPSC-HLCs. We noted that human liver temperature (around 39°C) is higher than normal human body temperature (around 36.5°C), yet hepatocytes are generally cultured at 37°C. Here, we tested the hypothesis that hepatic functions of hPSC-HLCs would be facilitated under physiologically-relevant hepatic temperatures. We identified the optimal temperature by treating HLCs derived from H9 human embryonic stem cells (hESC-HLCs) at 39°C and 42°C. As expected, 42°C caused significantly greater cell death compared to 39°C. Next, we measured the activities of cytochrome P450 3A4 (CYP3A4), which is one of the most important enzymes for hepatic detoxification. The 39°C-treated hESC-HLCs had CYP3A4 activities that were greater than the 37°C-treated hESC-HLCs. Albumin secretion significantly increased in the 39°C-treated hESC-HLCs. In combination with existing hepatic differentiation protocols, the method proposed here may further improve hepatic functions for hPSCs and thereby aid drug discovery efforts and improve drug toxicity tests.

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A Roadmap for Human Liver Differentiation from Pluripotent Stem Cells

Cited by 174 publications

References 86 publications

A critical look: Challenges in differentiating human pluripotent stem cells into desired cell types and organoids

A critical look: Challenges in differentiating human pluripotent stem cells into desired cell types and organoids

Doxycycline Significantly Enhances Induction of iPSCs to Endoderm by Enhancing survival via AKT Phosphorylation

Heat treatment functionalizes hepatocyte-like cells derived from human embryonic stem cells

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