Adult human hepatocytes induce Interleukin-10 producing allogeneic dendritic cells with tolerogenic properties

One of the major limitations of liver cell transplantation (LCT) is the fading of the metabolic effect. To shed some light on the potential involvement of the immune response in this loss of effect, we investigated the immunogenic properties of isolated human hepatocytes (hHep) and compared it to those of adult derived human liver progenitor cells, an alternative cell candidate for LCT. Using flow cytometry, we observed that both hHep and progenitor cells constitutively expressed major histocompatibility complex (MHC) class I and FAS but did not express human leukocyte antigen (HLA)-DR, FAS-ligand and co-stimulatory molecules CD80 and CD40. In a model of co-culture with allogeneic human adult peripheral blood mononuclear cells (PBMC), hHep and progenitor cells did not induce early T cell activation or proliferation, as demonstrated by flow cytometry and the tritiated thymidine incorporation method respectively. Furthermore, transwell experiments demonstrated that hHep induced a cell contact-dependent production of interleukin (IL)-10 that was not observed with progenitor cells, even after in vitro hepatogenic differentiation. We demonstrated using flow cytometry that IL-10 was produced by a myeloid dendritic cell (DC) subset characterized by an incomplete mature state with upregulated CD40 but not HLA-DR expression. To further characterize the consequence of IL-10 production, monocyte-derived DC were co-cultured for 48 hours with allogeneic hHep then harvested and co-cultured with allogeneic naive CD4+ T lymphocytes in the presence or absence of a blocking antibody directed against IL-10. Using tritiated thymidine incorporation, we demonstrated that DC previously co-cultured with hHep, induced less proliferation of naive CD4+ T lymphocytes than DC cultured alone. Blocking experiment demonstrated that IL-10 contributes to the naïve CD4+ T lymphocyte hyporesponsiveness. Finally, in order to determine whether CD4+ T cell hyporesponsiveness was antigen dependent or not, DC-primed CD4+ T lymphocytes were restimulated by DC from the same donor as the primary co-culture or from an unrelated donor. We observed that CD4+ T lymphocyte were less responsive to restimulation when initially stimulated by hepatocyte-primed DC than DC previously cultured in medium alone, regardless of the stimulating DC donor used for the restimulation. Our results suggest that both hHep and progenitor cells present a low immunogenic phenotype in vitro. We further demonstrated that allogeneic hHep but not progenitor cells, promote a cell contact-dependent production of IL-10 by myeloid DC, which consequently induced naïve CD4+ T lymphocyte antigen-independent anergy.