Sequential Hepatogenic Transdifferentiation of Adipose Tissue-Derived Stem Cells: Relevance of Different Extracellular Signaling Molecules, Transcription Factors Involved, and Expression of New Key Marker Genes

Abstract

Adipose tissue contains a mesenchymal stem cell (MSC) population known as adipose-derived stem cells (ASCs) capable of differentiating into different cell types. Our aim was to induce hepatic transdifferentiation of ASCs by sequential exposure to several combinations of cytokines, growth factors, and hormones. The most efficient hepatogenic protocol includes fibroblastic growth factors (FGF) 2 and 4 and epidermal growth factor (EGF) (step 1), hepatocyte growth factor (HGF), FGF2, FGF4, and nicotinamide (Nic) (step 2), and oncostatin M (OSM), dexamethasone (Dex), and insulin-tranferrin-selenium (step 3). This protocol activated transcription factors [GATA6, Hex, CCAAT/enhancer binding protein α and β (CEBPα and β), peroxisome proliferator-activated receptor-γ, coactivator 1 α (PGC1α), and hepatocyte nuclear factor 4 α (HNF4α)], which promoted a characteristic hepatic phenotype, as assessed by new informative markers for the stepby-step hepatic transdifferentiation of hMSC [early markers: albumin (ALB), α-2-macroglobuline (α2M), complement protein C3 (C3), and selenoprotein P1 (SEPP1); late markers: cytochrome P450 3A4 (CYP3A4), apolipoprotein E (APOE), acyl-CoA synthetase long-chain family member 1 (ACSL1), and angiotensin II receptor, type 1 (AGTR1)]. The loss of adipose adult stem cell phenotype was detected by losing expression of Thy1 and inhibitor of DNA binding 3 (Id3). The reexpression of phosphoenolpyruvate corboxykinase (PEPCK), apolipoprotein C3 (APOCIII), aldolase B (ALDOB), and cytochrome P450 1A2 (CYP1A2) was achieved by transduction with a recombinant adenovirus for HNF4α and finally hepatic functionality was also assessed by analyzing specific biochemical markers. We conclude that ASCs could represent an alternative tool in clinical therapy for liver dysfunction and regenerative medicine.

Adipose-Derived Stem Cells in Tissue Regeneration: A Review

“…This low level of HNF4 appears to have functional consequences, apparently restricting ASC-derived hepatocytes to more immature stages, as the HNF4 targets PEPCK, apolipoprotein C3 and aldolase B, are not expressed in their hepatic induced ASCs. However, when these ASCs are transduced with an HNF4 adenovirus, expression of these plasma proteins is detected at both the gene and protein level, confirming that HNF4 -driven terminal hepatogenesis is possible in ASCs [160]. Consistent with this, Lue and coworkers find that induced ASCs exhibit low levels of five key hepatic transcription factors, including HNF4 [161].…”

“…In the field of liver regeneration, Banas and colleagues were the first to show possible in vitro formation of hepatocytes by ASCs by measuring increased albumin expression by the induced ASCs at the protein level [159]. Bonora-Centelles et al 2009 also confirm increased albumin expression, together with increased gene expression of the liver markers 2-macroglobuline (an acute phase protein), complement C3, selenoprotein/SEPP1 (an antioxidant), cytochrome P450 3A4, apolipoprotein E, and HNF4 , albeit at low levels [160]. This low level of HNF4 appears to have functional consequences, apparently restricting ASC-derived hepatocytes to more immature stages, as the HNF4 targets PEPCK, apolipoprotein C3 and aldolase B, are not expressed in their hepatic induced ASCs.…”

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.…”

Implications of the immunoregulatory functions of mesenchymal stem cells in the treatment of human liver diseases

“…2,3 In particular, MSCs can differentiate into hepatocyte-like cells through a specific cell-signaling pathway upon stimulation with a combination of hepatocyte growth factor (HGF), fibroblast growth factor, epidermal growth factor and other cytokines. [4][5][6] However, in a culture of human bone marrowderived MSCs (BM-MSCs) and hepatocytes, BM-MSCs that undergo hepatic differentiation probably account for only a small percentage of the population, 7 and any differentiation may not significantly change the outcome of the bone marrow stromal cell (BMSC) and hepatocyte coculture experiments. 8 MSCs have recently emerged as promising candidates for cell-based immunotherapy because these cells can modulate the immune responses in various ways.…”