Submitted on June 2, 2006
Accepted on September 12, 2006
Two populations of Thy1-positive mesenchymal cells regulate the in vitro maturation of hepatic progenitor cells
Naoko Kamo1, Kentaro Yasuchika2, Hideaki Fujii2, Toshitaka Hoppo2, Takafumi Machimoto2, Takamichi Ishii2, Naoya Fujita3, Takashi Tsuruo3, Jun k Yamashita4, Hajime Kubo5, and Iwao Ikai1*
1 Department of Surgery, Graduate School of Medicine Kyoto University, Kyoto, Japan
2 Kyoto, Japan; Department of Surgery, Graduate School of Medicine Kyoto University, Kyoto, Japan
3 Institute of Molecular and Cellular Biosciences, The University of Tokyo, Tokyo, Japan
4 Laboratory of Stem Cell Differentiation, Institute for Frontier Medical Sciences, Kyoto University, Kyoto, Japan
5 Molecular & Cancer Research Unit, HMRO, Kyoto University Graduate School of Medicine, Kyoto, Kyoto, Japan
* To whom correspondence should be addressed. E-mail: ikai{at}kuhp.kyoto-u.ac.jp.
We previously reported that the in vitro maturation of CD49f+Thy1-CD45- (CD49f-positive) fetal hepatic progenitor cells (HPCs) is supported by Thy1-positive mesenchymal cells derived from fetal liver. These mesenchymal cell preparations contain two populations, one of a cuboidal shape and the other spindle shaped in morphology. In this study, we determined that the mucin-type transmembrane glycoprotein gp38 could distinguish the cuboidal cells from the spindle cells by immunocytochemistry. RT-PCR analysis revealed the differences between the isolated CD49f±Thy1+gp38+CD45- (gp38-positive) cells and CD49f±Thy1+gp38-CD45- (gp38-negative) cells, while both cells expressed mesenchymal cell markers. The coculture with gp38-positive cells promoted the maturation of CD49f-positive HPCs, which was estimated by the positivity for Periodic Acid Schiff (PAS) staining, whereas the coculture with gp38-negative cells maintained CD49f-positive HPCs negative for PAS staining. Expression of mature hepatocyte markers, such as tyrosine amino transferase (TAT), tryptophan-2, 3-dioxygenase (TO) and glucose-6-phosphatase (G6P), were upregulated on HPCs by coculture with gp38-positive cells. Furthermore, transmission electron microscopy revealed the acquisition of mature hepatocyte features by HPCs cocultured with gp38-positive cells. This effect on maturation of HPCs was inhibited by the addition of conditioned medium derived from gp38-negative cells. By contrast, the upregulation of BrdU incorporation by HPCs demonstrated the proliferative effect of coculture with gp38-negative cells. In conclusion, these results suggest that in vitro maturation of HPCs promoted by gp38-positive cells may be opposed by an inhibitory effect of gp38-negative cells, which likely maintain the immature, proliferative state of HPCs.
