Highly Efficient Generation of Human Hepatocyte-Like Cells from Induced Pluripotent Stem Cells

Department of Cell Biology, Division of Pediatric Pathology, Medical College of Wisconsin, Milwaukee, WI 53226, USA.
Hepatology (Impact Factor: 11.06). 01/2010; 51(1):297-305. DOI: 10.1002/hep.23354
Source: PubMed


There exists a worldwide shortage of donor livers available for orthotropic liver transplantation and hepatocyte transplantation therapies. In addition to their therapeutic potential, primary human hepatocytes facilitate the study of molecular and genetic aspects of human hepatic disease and development and provide a platform for drug toxicity screens and identification of novel pharmaceuticals with potential to treat a wide array of metabolic diseases. The demand for human hepatocytes, therefore, heavily outweighs their availability. As an alternative to using donor livers as a source of primary hepatocytes, we explored the possibility of generating patient-specific human hepatocytes from induced pluripotent stem (iPS) cells. CONCLUSION: We demonstrate that mouse iPS cells retain full potential for fetal liver development and describe a procedure that facilitates the efficient generation of highly differentiated human hepatocyte-like cells from iPS cells that display key liver functions and can integrate into the hepatic parenchyma in vivo.

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    • "Notable improvements in efficiency and functionality have since been achieved by various groups, which based their protocols on developmental signaling and utilized adherent culture conditions. However, all directed differentiation protocols for hepatocyte-like cells (HLCs) published to date have relied on the use of recombinant growth factors such as activin A, Wnt3a, hepatocyte growth factor (HGF), oncostatin M (OSM), fibroblast growth factor 4 (FGF4), vascular endothelial growth factor (VEGF), epidermal growth factor (EGF), and bone morphogenetic protein 4 (BMP4) (Agarwal et al., 2008; Brolén et al., 2010; Cai et al., 2007; Chen et al., 2012; Hay et al., 2008; Liu et al., 2010; Si-Tayeb et al., 2010b; Song et al., 2009; Sullivan et al., 2010; Touboul et al., 2010). Some recent progress has been made in replacing growth factors for the differentiation of mesoderm and ectoderm (Chambers et al., 2012; Lian et al., 2012), and efforts have been undertaken to find suitable candidates for the production of definitive endoderm (DE), exemplified by the Melton group, who identified IDE1 and 2 (Borowiak et al., 2009). "
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    • "Differentiation of porcine ES in SþSF-ACT media for 3 days induced the highest expression of endoderm markers. Low-serum (2%) or serum-free conditions are usually used for hepatic induction of human and mouse ES because several factors that are contained in serum inhibit hepatic differentiation [10] [24]. However, our porcine ES could not survive long-term in low-serum or serum-free conditions but short-term serum starvation with activin A was beneficial for inducing high expression of the FOXA2 and SOX17 genes. "
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    ABSTRACT: Porcine embryonic stem cells (ES) are considered attractive preclinical research tools for human liver diseases. Although several studies previously reported generation of porcine ES, none of these studies has described hepatic differentiation from porcine ES. The aim of this study was to generate hepatocytes from porcine ES and analyze their characteristics. We optimized conditions for definitive endoderm induction and developed a 4-step hepatic differentiation protocol. A brief serum-free condition with activin A efficiently induced definitive endoderm differentiation from porcine ES. The porcine ES-derived hepatocyte-like cells highly expressed hepatic markers including albumin and α-fetoprotein, and displayed liver characteristics such as glycogen storage, lipid production, and low-density lipoprotein uptake. For the first time, we describe a highly efficient protocol for hepatic differentiation from porcine ES. Our findings provide valuable information for translational liver research using porcine models, including hepatic regeneration and transplant studies, drug screening, and toxicology. Copyright © 2015 Elsevier Inc. All rights reserved.
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    • "A common shortcoming of iHLCs and other iPSCderived cells lies in the fact that they often resemble a developmentally immature state compared to the fully differentiated adult counterpart (Engle and Puppala, 2013). In the field of iHLCs, current in vitro differentiation protocols result in the production of a hepatic cell type that is biologically closer to fetal hepatocytes than adult hepatocytes , due to the incomplete abrogation of the expression of fetal markers such as alpha-fetal protein (AFP) (Si-Tayeb et al., 2010; Song et al., 2009), and the incomplete acquisition of an adult-like levels of key secretory, detoxification, and metabolic activity (Shan et al., 2013). Our data show that further chemical maturation of iHLCs allows the acquisition of primaquine sensitivity, presumably via the acquisition of a drug metabolism enzyme (DME) expression profile that better resembles the adult human hepatocyte . "
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    ABSTRACT: Malaria eradication is a major goal in public health but is challenged by relapsing malaria species, expanding drug resistance, and the influence of host genetics on antimalarial drug efficacy. To overcome these hurdles, it is imperative to establish in vitro assays of liver-stage malaria for drug testing. Induced pluripotent stem cells (iPSC) potentially allow the assessment of donor-specific drug responses, and iPSC-derived hepatocyte-like cells (iHLCs) can facilitate the study of host genetics on host-pathogen interactions and the discovery of novel targets for antimalarial drug development. We establish in vitro liver-stage malaria infections in iHLCs using P. berghei, P. yoelii, P. falciparum, and P. vivax and show that differentiating cells acquire permissiveness to malaria infection at the hepatoblast stage. We also characterize antimalarial drug metabolism capabilities of iHLCs using prototypical antimalarial drugs and demonstrate that chemical maturation of iHLCs can improve their potential for antimalarial drug testing applications. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.
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