title: A combining method to enhance the in vitro differentiation of hepatic precursor cells.
abstract: The ideal bioartificial liver should be designed to reproduce as nearly as possible in vitro the habitat that hepatic cells find in vivo. In the present work, we investigated the in vitro perfusion condition with a view to improving the hepatic differentiation of pluripotent human liver stem cells (HLSCs) from adult liver. Tissue engineering strategies based on the cocultivation of HLSCs with hepatic stellate cells (ITO) and with several combinations of medium were applied to improve viability and differentiation. A mathematical model estimated the best flow rate for perfused cultures lasting up to 7 days. Morphological and functional assays were performed. Morphological analyses confirmed that a flow of perfusion medium (assured by the bioreactor system) enabled the in vitro organization of the cells into liver clusters even in the deeper levels of the sponge. Our results showed that, when cocultured with ITO using stem cell medium, HLSCs synthesized a large amount of albumin and the MTT test confirmed an improvement in cell proliferation. In conclusion, this study shows that our in vitro cell conditions promote the formation of clusters of HLSCs and enhance the functional differentiation into a mature hepatic population.
authors.affiliation: Hepatobiliary and Liver Transplant Unit, Department of General Surgery and Organ Transplantation, University of Padova, Padova, Italy.
authors.forename: Amedeo
authors.initials: A
authors.lastname: Carraro
authors.forename: Marina
authors.initials: M
authors.lastname: Flaibani
authors.forename: Umberto
authors.initials: U
authors.lastname: Cillo
authors.forename: Lisa
authors.initials: L
authors.lastname: Michelotto
authors.forename: Enrico
authors.initials: E
authors.lastname: Magrofuoco
authors.forename: Maurizio
authors.initials: M
authors.lastname: Buggio
authors.forename: Giovanni
authors.initials: G
authors.lastname: Abatangelo
authors.forename: Roberta
authors.initials: R
authors.lastname: Cortivo
authors.forename: Maria Beatriz
authors.initials: MB
authors.lastname: Herrera
authors.forename: Ciro
authors.initials: C
authors.lastname: Tetta
authors.forename: Nicola
authors.initials: N
authors.lastname: Elvassore
authors.forename: Barbara
authors.initials: B
authors.lastname: Zavan
mesh_terms: D000328:Adult; D053687:Adult Stem Cells / Q000166:cytology* / Q000502:physiology*; D018929:Cell Culture Techniques / Q000379:methods; D002454:Cell Differentiation / Q000502:physiology*; D049109:Cell Proliferation; D002470:Cell Survival; D002478:Cells, Cultured; D018920:Coculture Techniques / Q000379:methods; D022781:Hepatocytes / Q000166:cytology / Q000502:physiology*; D006801:Humans; D008099:Liver / Q000166:cytology; D008954:Models, Biological; D008962:Models, Theoretical; D023822:Tissue Engineering / Q000379:methods*