Differentiation and isolation of hepatic-like cells from human embryonic stem cells

Neta Lavon, Ofra Yanuka, Nissim Benvenisty*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

226 Scopus citations

Abstract

Human embryonic stem cells are pluripotent cells that can serve as a cell source for transplantation medicine, and as a tool to study human embryogenesis. We investigate here the potential of human embryonic stem cells to differentiate into hepatic cells. We have characterized the expression level of liver-enriched genes in undifferentiated and differentiated human embryonic stem cells by DNA microarrays. Our analysis revealed a subset of fetal hepatic enriched genes that are expressed in human embryonic stem cells upon differentiation into embryoid bodies. In order to isolate the hepatic-like cells, we introduced a reporter gene regulated by a hepatocyte-specific promoter into human embryonic stem cells. We isolated clones of human embryonic stem cells that express enhanced green fluorescent protein upon in vitro differentiation. Through immunostaining, we showed that most of these cells express albumin, while some cells still express the earlier expressed protein α-fetoprotein. Using fluorescence activated cell sorter, we were able to sort out the fluorescent differentiated cells and expand them for a few more weeks. This is the first report to demonstrate the possibility of purifying differentiated derivatives of human embryonic stem cells and culturing them further. Through confocal microscopy, we detected clusters of hepatic-like cells in 20-day-old embryoid bodies and in teratomas. As observed during embryonic development, we showed that in teratomas, the hepatic-like endodermal cells develop next to cardiac mesodermal cells. In order to examine the secreted factors involved in the induction of hepatic differentiation, human embryonic stem cells were grown in the presence of various growth factors, demonstrating the potential involvement of acidic fibroblast growth factor in the differentiation. In conclusion, given certain growth conditions and genetic manipulation, we can now differentiate and isolate hepatic-like cells from human embryonic stem cells.

Original languageAmerican English
Pages (from-to)230-238
Number of pages9
JournalDifferentiation
Volume72
Issue number5
DOIs
StatePublished - Jun 2004

Bibliographical note

Funding Information:
Acknowledgments We are grateful to Dr. Joseph Itskovitz-Eldor at the Rambam Medical Center for kindly providing us with the human ES cells as a collaboration. We are indebted to Drs. Karen Kotkow and Lee Rubin from Curis, Inc. for their valuable assistance. We thank Dr. Neomi Melamed-Book and Nurit Ilan for excellent technical assistance. This research was partially supported by funds to N.B. from the Herbert Cohn Chair (Hebrew University), and by funds from the United States-Israel Binational Science Foundation (grant no. 2001021).

Keywords

  • Development
  • Endoderm
  • Genetic manipulation
  • Hepatocytes
  • Liver
  • Stem cells

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