Differentiation and transplantation of human embryonic stem cell-derived hepatocytes.
OBJECTIVE
The ability to obtain unlimited numbers of human hepatocytes would improve the development of cell-based therapies for liver diseases, facilitate the study of liver biology, and improve the early stages of drug discovery. Embryonic stem cells are pluripotent, potentially can differentiate into any cell type, and therefore could be developed as a source of human hepatocytes.


METHODS
To generate human hepatocytes, human embryonic stem cells were differentiated by sequential culture in fibroblast growth factor 2 and human activin-A, hepatocyte growth factor, and dexamethasone. Functional hepatocytes were isolated by sorting for surface asialoglycoprotein-receptor expression. Characterization was performed by real-time polymerase chain reaction, immunohistochemistry, immunoblot, functional assays, and transplantation.


RESULTS
Embryonic stem cell-derived hepatocytes expressed liver-specific genes, but not genes representing other lineages, secreted functional human liver-specific proteins similar to those of primary human hepatocytes, and showed human hepatocyte cytochrome P450 metabolic activity. Serum from rodents given injections of embryonic stem cell-derived hepatocytes contained significant amounts of human albumin and alpha1-antitrypsin. Colonies of cytokeratin-18 and human albumin-expressing cells were present in the livers of recipient animals.


CONCLUSIONS
Human embryonic stem cells can be differentiated into cells with many characteristics of primary human hepatocytes. Hepatocyte-like cells can be enriched and recovered based on asialoglycoprotein-receptor expression and potentially could be used in drug discovery research and developed as therapeutics.
Basma
Hesham
H
Department of Surgery, University of Nebraska Medical Center, Omaha, Nebraska, USA.
Soto-Gutiérrez
Alejandro
A
Yannam
Govardhana Rao
GR
Liu
Liping
L
Ito
Ryotaro
R
Yamamoto
Toshiyuki
T
Ellis
Ewa
E
Carson
Steven D
SD
Sato
Shintaro
S
Chen
Yong
Y
Muirhead
David
D
Navarro-Alvarez
Nalu
N
Wong
Ronald J
RJ
Roy-Chowdhury
Jayanta
J
Platt
Jeffrey L
JL
Mercer
David F
DF
Miller
John D
JD
Strom
Stephen C
SC
Kobayashi
Naoya
N
Fox
Ira J
IJ
D028341:Activins / Q000494:pharmacology; D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D002460:Cell Line; D003907:Dexamethasone / Q000494:pharmacology; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000637:transplantation* / Q000648:ultrastructure; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D015870:Gene Expression; D053450:Glucocorticoid-Induced TNFR-Related Protein / Q000494:pharmacology; D017228:Hepatocyte Growth Factor / Q000494:pharmacology; D022781:Hepatocytes / Q000166:cytology*; D006801:Humans; D008297:Male; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D008854:Microscopy, Electron; D010641:Phenotype; D033581:Stem Cell Transplantation
D053450:Glucocorticoid-Induced TNFR-Related Protein; C422405:activin A; D016222:Fibroblast Growth Factor 2; D028341:Activins; D017228:Hepatocyte Growth Factor; D003907:Dexamethasone