PUTATIVE LIVER PROGENITOR CELLS - CONDITIONS FOR LONG-TERM SURVIVAL IN CULTURE

Oval cells proliferate extensively in the livers of animals exposed to oncogenic insults, are bipotent and are believed to be related to the so far unidentified liver stem cell. In normal liver, cells antigenic ally related to oval cells and expressing Liver and epithelial markers are considered to be liver progenitor cells. We isolated, by fluoresc ence-activated cell sorting or magnetic bead sorting, cells expressing the oval cell antigens OC.2 or OC.3 from the liver of normal newborn or day 12 embryonal age rats. Magnetic bead sorting of positive cells was as efficient as fluorescence-activated cell sorting. A two-chamber culture system was devised in which cells were plated onto transwell filters coated with type IV collagen and cultured in a serum-free Ham' s F12 medium supplemented with free fatty acids and bovine serum album in. Under these conditions, cells remained viable for up to 6 weeks an d their antigenic phenotype was unchanged throughout. Approximately 30 % of sorted cells expressed epithelial and/or liver-specific markers. Growth factors mitogenic for epithelial cells and hepatocytes did not elicit cell proliferation. These results provide an important backgrou nd for further studies designed to determine the biological significan ce of OC.2(+) and OC.3(+) cells in normal liver, to test the liver ste m cell hypothesis and to develop protocols for the expansion in vitro of normal liver progenitors.