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Few studies on the long‐term culture of postnatal mouse hepatic stem/progenitor cells have been reported. We successfully adapted a serum‐free culture system that we employed previously to expand fetal mouse hepatic stem/progenitor cells and maintained them in culture over long periods. The expanded postnatal cells contained immature α‐fetoprotein‐positive cells along with hepatocytic and cholangiocytic lineage‐committed cells. These cells expressed CD49f but not CD45, CD34, Thy‐1, c‐kit, CD31, or flk‐1, and oncostatin M induced their differentiation. This heterogeneous population contained side population (SP) cells, which express the ATP‐binding cassette transporter ABCG2, and sca‐1+ cells. As mice aged, the frequency of SP and sca‐1+ cells decreased along with the ability of cultured cells to expand. Approximately 20%–40% of the SP cells expressed sca‐1, but only a few sca‐1+ cells were also SP cells. Analysis of colonies derived from single SP or sca‐1+ cells revealed that, although both cells had dual differentiation potential and self‐renewal ability, SP cells formed colonies more efficiently and gave rise to SP and sca‐1+ cells, whereas sca‐1+ cells generated only sca‐1+ progeny. Thus, SP cells are more characteristic of stem cells than are sca‐1+ cells. In regenerating livers, ABCG2+ cells and sca‐1+ cells were detected around or in the portal area (the putative hepatic stem cell niche). The expanded cells share many features of fetal hepatic stem/progenitor cells or oval cells and may be useful in determining the mechanisms whereby hepatic stem cells self‐renew and differentiate.  Disclosure of potential conflicts of interest is found at the end of this article.

Long‐Term Culture of Postnatal Mouse Hepatic Stem/Progenitor Cells and Their Relative Developmental Hierarchy