Comparative Analysis of Human Embryonic Stem Cell and Induced Pluripotent Stem Cell-Derived Hepatocyte-Like Cells Reveals Current Drawbacks and Possible Strategies for Improved Differentiation

Abstract

Hepatocytes derived from human embryonic stem cells (hESCs) or induced pluripotent stem cells (iPSCs) could provide a defined and renewable source of human cells relevant for cell replacement therapies and toxicology studies. However, before patient-specific iPSCs can be routinely used for these purposes, there is a dire need to critically compare these cells to the golden standard-hESCs. In this study, we aimed at investigating the differences and similarities at the transcriptional level between hepatocyte-like cells (HLCs) derived from both hESCs and iPSCs. Two independent protocols for deriving HLCs from hESCs and iPSCs were adopted and further characterization included immunocytochemistry, real-time (RT)-polymerase chain reaction, and in vitro functional assays. Comparative microarray-based gene expression profiling was conducted on these cells and compared to the transcriptomes of human fetal liver and adult liver progenitors. HLCs derived from hESCs and human iPSCs showed significant functional similarities, similar expression of genes important for liver physiology and common pathways. However, specific differences between the 2 cell types could be observed. For example, among the cytochrome P450 gene family, CYP19A1, CYP1A1, and CYP11A1 were enriched in hESC-derived HLCs, and CYP46A1 and CYP26A1 in iPSC-derived HLCs. HLCs derived from hESCs and human iPSCs exhibited broad similarities but as well meaningful differences. We identified common upregulated transcription factors, which might serve as a source for generating a cocktail of factors able to directly transdifferentiate somatic cells into HLCs. The findings may be vital to the refinement of protocols for the efficient derivation of functional patientspecific HLCs for regenerative and toxicology studies.

Molecular mechanisms regulating the establishment of hepatocyte polarity during human hepatic progenitor cell differentiation into a functional hepatocyte-like phenotype

“…Selected differential express genes involved in the HLCs polarity formation differentiation (Wang and Boyer, 2004;Takashi et al, 2007). Although information has recently been accumulated suggesting specific routes and mechanisms for this process (Matsui et al, 2002;Michalopoulos et al, 2003;Imamura et al, 2007;Chiao et al, 2008;Bonora-Centelles et al, 2009;Kidambi et al, 2009;Saulnier et al, 2010;Synnergren et al, 2010;Jozefczuk et al, 2011;Fu et al, 2011), the exact mechanism for hepatocyte polarity generation is unclear partly because only a few in vitro models are available for developing complex human hepatocyte polarity (Decaens et al, 2008). The goal of this study was to generate functional polarized hepatocytes from the hFHPCs in vitro and to explore the molecular mechanisms involved in hepatocyte polarization.…”

“…Using microarray analysis, several studies have examined the key genes and pathways of potential importance for generating hepatocyte-like cells (HLCs). In these reports, the liver-specific gene expression was identified within the total, heterogeneous population of cells that differentiated from human embryonic stem cells (Chiao et al, 2008;Synnergren et al, 2010;Jozefczuk et al, 2011) and induced pluripotent stem cells (Jozefczuk et al, 2011), HepaRG liver progenitor cells (Parent and Beretta, 2008) or human adipose tissue-derived stromal cells (Bonora-Centelles et al, 2009;Saulnier et al, 2010). Although these systems have limitations inherent to their respective origins, they represent human models of hepatocyte differentiation.…”

Differentiation and Application of Induced Pluripotent Stem Cell–Derived Vascular Smooth Muscle Cells

“…For instance, the embryoniclike nature of iPSCs makes modeling diseases usually seen in the elderly or in patients in the later stages of disease more difficult. 98 Thus, alternative cell types, such as skin-derived precursors, ought to be recognized for their potential as VSMC sources as they readily differentiate into VSMCs using the same growth factors required to generate VSMCs from neural crest precursors. 99 Moreover, they have been used to model type II diabetes mellitus in elderly patients, without the need for reprogramming required in iPSC generation, often resulting in erasure of disease-relevant epigenetic modifications.…”

New Models for the In Vitro Study of Liver Toxicity: 3D Culture Systems and the Role of Bioreactors

“…Derived from the inner cell mass of preimplantationstage blastocysts  Totipotent stem cells  Self-renewal  Huge proliferative potential  High differentiation potential and plasticity  Expression of early markers of hepatic differentiation  Some mature hepatic functions are maintained  Difficulty to be regulated and maintained under controlled conditions  Risk of oncogenicity  Ethical issues  Limited availability Agarwal et al, 2008Baharvand et al, 2006& 2008Ishii, et al 2010Jozefczuk et al, 2011Liu T et al, 2010Rambhatla , et al, 2003Soto-Gutierrez et al, 2007 Extra-hepatic adult stem cells Bone marrow  Pluripotent stem cells  Inducible to express a number of liver-specific functions  Difficulty to reach full adult hepatocyte phenotype  Less differentiated than primary hepatocytes Avital et al, 2001Chen et al, 2006Chivu et al, 2009Petersen et al, 1999 Adipose tissue  Pluripotent stem cells  Express to some extent specific hepatic functions  Quantitatively less differentiated than HEPG2 cell line  Less differentiated than primary hepatocytes Okura et al, 2010 Umbilical cord blood Takahashi et al, 2007Espejel et al, 2010Hu et al, 2010Q. Feng et al, 2010Sullivan et al, 2010H.…”