title: Directed differentiation of human embryonic stem cells into functional hepatic cells.
abstract: The differentiation capacity of human embryonic stem cells (hESCs) holds great promise for therapeutic applications. We report a novel three-stage method to efficiently direct the differentiation of human embryonic stem cells into hepatic cells in serum-free medium. Human ESCs were first differentiated into definitive endoderm cells by 3 days of Activin A treatment. Next, the presence of fibroblast growth factor-4 and bone morphogenetic protein-2 in the culture medium for 5 days induced efficient hepatic differentiation from definitive endoderm cells. Approximately 70% of the cells expressed the hepatic marker albumin. After 10 days of further in vitro maturation, these cells expressed the adult liver cell markers tyrosine aminotransferase, tryptophan oxygenase 2, phosphoenolpyruvate carboxykinase (PEPCK), Cyp7A1, Cyp3A4 and Cyp2B6. Furthermore, these cells exhibited functions associated with mature hepatocytes including albumin secretion, glycogen storage, indocyanine green, and low-density lipoprotein uptake, and inducible cytochrome P450 activity. When transplanted into CCl4 injured severe combined immunodeficiency mice, these cells integrated into the mouse liver and expressed human alpha-1 antitrypsin for at least 2 months. In addition, we found that the hESC-derived hepatic cells were readily infected by human immunodeficiency virus-hepatitis C virus pseudotype viruses.
  
  
  CONCLUSIONS
  We have developed an efficient way to direct the differentiation of human embryonic stem cells into cells that exhibit characteristics of mature hepatocytes. Our studies should facilitate searching the molecular mechanisms underlying human liver development, and form the basis for hepatocyte transplantation and drug tests.
authors.affiliation: Key Laboratory of Cell Proliferation and Differentiation of the Ministry of Education, College of Life Sciences, Peking University, Beijing, China.
authors.forename: Jun
authors.initials: J
authors.lastname: Cai
authors.forename: Yang
authors.initials: Y
authors.lastname: Zhao
authors.forename: Yanxia
authors.initials: Y
authors.lastname: Liu
authors.forename: Fei
authors.initials: F
authors.lastname: Ye
authors.forename: Zhihua
authors.initials: Z
authors.lastname: Song
authors.forename: Han
authors.initials: H
authors.lastname: Qin
authors.forename: Sha
authors.initials: S
authors.lastname: Meng
authors.forename: Yuezhou
authors.initials: Y
authors.lastname: Chen
authors.forename: Rudan
authors.initials: R
authors.lastname: Zhou
authors.forename: Xijun
authors.initials: X
authors.lastname: Song
authors.forename: Yushan
authors.initials: Y
authors.lastname: Guo
authors.forename: Mingxiao
authors.initials: M
authors.lastname: Ding
authors.forename: Hongkui
authors.initials: H
authors.lastname: Deng
mesh_terms: D028341:Activins / Q000494:pharmacology; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins / Q000494:pharmacology; D002454:Cell Differentiation / Q000502:physiology*; D016895:Culture Media, Serum-Free; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000187:drug effects; D051519:Fibroblast Growth Factor 4 / Q000494:pharmacology; D006678:HIV / Q000502:physiology; D016174:Hepacivirus / Q000502:physiology; D022781:Hepatocytes / Q000166:cytology* / Q000502:physiology / Q000821:virology; D006801:Humans; D016212:Transforming Growth Factor beta / Q000494:pharmacology
chemical_list: C525718:BMP2 protein, human; C525719:Bmp2 protein, mouse; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins; D016895:Culture Media, Serum-Free; C496445:FGF4 protein, human; D051519:Fibroblast Growth Factor 4; D016212:Transforming Growth Factor beta; C422405:activin A; D028341:Activins