Carboxymethylpachymaran enhances immunologic function of dendritic cells cultured in two kinds of hepatoma carcinoma cell line’s supernatant via nuclear factor κB/Rel pathway

Objective:To study the immunologic function of dendritic cells(DCs) cultured in two kinds of hepatoma cell line’s supernatant and the enhancing effects of carboxymethylpachymaran(CMP) on DCs.Methods:DCs were harvested after stimulation by granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin(IL)-4 from umbilical cord blood using density-gradient centrifugation method. Cultured supernatant of two hepatoma cell lines(HepG2 and HepG2.2.15) were collected for condition medium(CM) according to a volume ratio of supernatant to incomplete RPMI-1640 medium,which was 3:1.CMP was dissolved in incomplete RPMI-1640 medium.Experimental groups were divided according to the culture medium,either CM or with CMP in it.DCs subsets CD83,CD86,CD1a,and d-related human leukocyte antigens(HLA-DR) were analyzed by flow cytometry.The proliferation ability of allogeneic T cells in mixed lymphocyte reaction(MLR) stimulated by DCs was examined by 3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide(MTT) analysis.IL-12p70,interferon-γ(IFN-γ),and nuclear factor k B(NF- k B) were detected by enzyme-linked immunosorbent assay analysis.Results:The proliferation of lymphocytes and secreting level of IL-12 and expression of phenotype of DCs cultured in two kinds of CM were lower than those of normal group(P<0.01).Compared with the normal group,groups treated with CMP showed a higher expression level of DCs subsets,lymphocyte reproductive activity,as well as IL-12 and IFN-γsecretion levels.Groups treated with CMP also demonstrated higher levels of DCs phenotype expression and IL-12 and IFN-γsecretion in supernatant of MLR and higher lymphocyte reproductive activity compared with CM group(P<0.05).Compared with the normal group,the expression level of NF-k B in DCs nuclear was higher in CMP groups but lower in two CM groups(P<0.05).After CMP was added,the NF-κB expression levels of two CM groups were increased compared with levels before CMP was added(P<0.05).However,there was no significant difference between the two CM groups(P>0.05).Conclusions:Two kinds of hepatoma cell line’s supernatant can inhibit the immunologic function of DCs.This suppressive effect may be related to the inhibition of NF-κB/Rel pathway.CMP may up-regulate the DCs function by activating the NF-κB/Rel pathway.