关键词: 肝前体样细胞, 永生化, 生物人工肝, 急性肝衰竭

Abstract: Objective To verify the function of hepatocyte-derived liver progenitor-like cells (HepLPCs), and to find new seed cells for bioartificial liver application.Methods Human primary hepatocytes were isolated by 2-step separation method, and hepatocytes were purified by fluorescence activated cell sorting to exclude HepLPCs from cluster of differentiation 24 + or epithelial cell adhesion molecule + progenitor cells. HepLPCs were immortalized by large T antigen. Then, cell morphology, proliferation states, functional gene levels, detoxification and synthesis levels, and responses to acute hepatic failure (AHF) toxic plasma of immortalized HepLPCs (iHepLPCs) were comprehensively evaluated. Results After immortalization, iHepLPCs could proliferate rapidly and stably in different passages. There was no significant difference in the proliferation rates of passage 0, passage 10 and passage 20 at the same time point (P>0.05), and the cell morphology remained unchanged. The cell detoxification and synthesis functions were stable during the proliferation process, and there was no significant difference among passages (P>0.05). Besides, the differentiation of iHepLPCs cells significantly enhanced expression of the functional genes. The urea production of differentiated iHepLPCs cells was higher than that of undifferentiated ones (13.683±1.238 vs 7.769±0.323, P<0.005). Also, albumin synthesis of differentiated iHepLPCs cells was higher than that of undifferentiated ones (11.270±0.63 vs 5.565±0.025, P<0.005). Furthermore, differentiated iHepLPCs cells had certain tolerance to AHF plasma with detoxification and synthesis abilities. After 6-hour culture in AHF plasma, urea synthesis reached 8.25±0.31, and albumin synthesis reached 12±0.41.Conclusion The iHepLPCs have certain tolerance to AHF plasma, whose cell morphology, detoxification and synthesis functions maintain stable in the process of rapid proliferation. Therefore, iHepLPCs could be a new source of bioartificial hepatocytes.

Key words: Hepatocyte-derived liver progenitor-like cells, Immortalization, Bioartificial liver, Acute liver failure