Purification of adult hepatic progenitor cells using green fluorescent protein (GFP)-transgenic mice and fluorescence-activated cell sorting.
OBJECTIVE
Recent advances in stem cell research have revealed that hepatic stem/progenitor cells may play an important role in liver development and regeneration. However, a lack of detectable definitive markers in viable cells has hindered their primary culture from adult livers.


METHODS
Enzymatically dissociated liver cells from green fluorescent protein (GFP)-transgenic mice, which express GFP highly in liver endodermal cells, were sorted by GFP expression using a fluorescence-activated cell sorter. Sorted cells were characterized, and also low-density cultured for extended periods to determine their proliferation and clonal differentiation capacities.


RESULTS
When CD45(-)TER119(-) side-scatter(low) GFP(high) cells were sorted, alpha-fetoprotein-positive immature endoderm-characterized cells, having high growth potential, were present in this population. Clonal analysis and electron microscopic evaluation revealed that each single cell of this population could differentiate not only into hepatocytes, but also into biliary epithelial cells, showing their bilineage differentiation activity. When surface markers were analyzed, they were positive for Integrin-alpha6 and -beta1, but negative for c-Kit and Thy1.1.


CONCLUSIONS
Combination of GFP-transgenic mice and fluorescence-activated cell sorting enabled purification of hepatic progenitor cells from adult mouse liver. Further analysis of this population may lead to purification of their human correspondence that would be an ideal cell-source candidate for regenerative medicine.
Fujikawa
Takahisa
T
Department of Gastroenterological Surgery, Kyoto University Graduate School of Medicine, 54 Kawara, Shogoin, Sakyo-ku, Kyoto 606-8507, Japan.
Hirose
Tetsuro
T
Fujii
Hideaki
H
Oe
Shoshiro
S
Yasuchika
Kentaro
K
Azuma
Hisaya
H
Yamaoka
Yoshio
Y
D000367:Age Factors; D000818:Animals; D015415:Biomarkers; D002454:Cell Differentiation; D002469:Cell Separation / Q000379:methods*; D005434:Flow Cytometry; D015870:Gene Expression; D049452:Green Fluorescent Proteins; D007202:Indicators and Reagents / Q000378:metabolism; D008099:Liver / Q000166:cytology*; D008164:Luminescent Proteins / Q000235:genetics*; D051379:Mice; D008822:Mice, Transgenic; D013234:Stem Cells / Q000166:cytology* / Q000502:physiology
D015415:Biomarkers; D007202:Indicators and Reagents; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins