Generation of Hepatocyte-Like Cells from Human Embryonic Stem Cells

Abstract

Use of human hepatocytes for therapeutic and drug discovery applications is hampered by limited tissue source and the inability of hepatocytes to proliferate and maintain function long term in vitro. Human embryonic stem (hES) cells are immortal and pluripotent and may provide a cell source for functional human hepatocytes. We report here that hES cells can be induced to differentiate into hepatocyte-like cells. Treatment with sodium butyrate induced hepatic differentiation as well as significant cell death, resulting in approximately 10-15% yield of a homogeneous population of cells. The differentiated cells have morphological features similar to that of primary hepatocytes and 70-80% of the cells express liver-associated proteins (albumin, alpha-1-antitrypsin, cytokeratin 8 and 18), accumulate glycogen, have inducible cytochrome P450 activity, and do not express alpha-fetoprotein. Because of the inherent proliferative capacity of hES cells, these cells may provide a reliable source of normal human hepatocytes for research and transplantation.

In vitro differentiation of hepatic progenitor cells from mouse embryonic stem cells induced by sodium butyrate

“…2D-F), but not AFP ( Fig. 2A-C), which was consistent with the process previously described [Rambhatla et al, 2003]. After removal of sodium butyrate, the granular-rich large cells changed into ES-derived hepatic progenitor cells accompanied with the increased expression of AFP and decreased expression of ALB until undetectable with immunofluorescence ( Fig.…”

“…The treated cells exhibited morphological features of primary hepatocytes and expressed liver-associated genes and proteins. In addition, the majority of the treated cells remained in cell cycle for 3 days after the addition of sodium butyrate, but the cycling fraction decreased to 17% 7 days later [Rambhatla et al, 2003]. It was now established that treatment of cells both in vitro and in vivo with sodium butyrate, a potential histone deacetylase (HDAC) inhibitor, could result in specific functional outcomes such as proliferation, cell cycle arrest, apoptosis, or differentiation [Janson et al, 1997;Sambucetti et al, 1999;Wang et al, 1999;Travers et al, 2002;Camphausen et al, 2004;Hsieh et al, 2004;Bug et al, 2005;Cho et al, 2005;Jiang et al, 2005;Rossig et al, 2005].…”

Differentiation of mouse embryonic stem cells to hepatocyte-like cells by co-culture with human liver nonparenchymal cell lines

“…The recent report of a technique for efficient generation of definitive endoderm from ES cells by culture in lower concentrations of serum and Activin A [7][8][9] has been an important advance for subsequent production of hepatocyte-like cells. We have found that induction of definitive endoderm from ES cells was a necessary step for subsequent hepatic differentiation in vitro.…”

Study of hepatocyte differentiation using embryonic stem cells

“…In addition, the hepatic-like cells were suggested to develop in a niche next to cardiac mesodermal cells and that aFGF may play a role in this differentiation. The differentiation towards hepatic-like cells were also demonstrated by using other factors [Rambhatla et al, 2003;Shirahashi et al, 2004]. Shirahashi et al [2004], added insulin and dexamethasone to EBs cultured on collagen type I and showed that the cells express various endodermal genes.…”

“…Shirahashi et al [2004], added insulin and dexamethasone to EBs cultured on collagen type I and showed that the cells express various endodermal genes. Rambhatla et al [2003], added sodium butyrate to the culture media inducing hepatic differentiation as well as significant cell death. The resultant cells had morphological features similar to that of primary hepatocytes and most of the cells expressed liver associated proteins.…”