Expression Of Thansciption FOXP3 In HCC Cell Lines And Its Significance

Objective :We firstly evaluated the Foxp3 mRNA and protein expression in HCC tumor cell lines. Then, analyzed the down-stream regulatory gene by Foxp3 plasmid and Foxp3 shRNA transfected to up/down-regulating the Foxp3 expression in HCC tumor cells.Thirdly, co-cultured the Foxp3+ HCC tumor cells with CD4+ and CD8+ T response cells and evaluated immune-related inhibitory functions of Foxp3+ HCC cells in co-culture systems.Methods:Conventional RT-PCR and cDNA expression profile were used to detect the expression of Foxp3 mRNA in 10 HCC cell lines while protein expression was assessed by western blots, immunocytochemistry and flow cytometry. Down-regulation of Foxp3 was used by Foxp3 shRNA in hepatocellular carcinoma cells. Meanwhile,up-regulation of Foxp3 was used by Foxp3 span-RNA in hepatocellular carcinoma cells. Down stream genes regulated by Foxp3 in hepatocellular carcinoma cell lines were analyzed using microarray and then verified by quantitative realtime PCR and FlowCytomix Pro. The immune-related inhibitory functions in Foxp3-expressing HCC cells were evaluated by co-culturing tumor cells with native CD4+ T cells and CD8+T cells respectively.Results :Foxp3 mRNA and protein was detected in all hepatocellular carcinoma cell lines . In addition, the foxp3-expressing hepatoma cells can significantly inhibit of CD4 + /CD8+T cell proliferation and activation in the co-culture system, Foxp3 expression correlated with the expression levels of IL-1a, IL-1b, IL-6, IL-8, TGFb2, TGFb3, HLA-A, HLA-B, SMAD6 , B2M and Jun. Silencing of Foxp3 by shRNA could up-regulate the expression of IL-1a, IL-1b, IL-6, IL-8, HLA-A, HLA-B, B2M and Jun. The expression of SMAD6, TGFb2, TGFb3 in hepatocellular carcinoma cell lines down-regulated after Foxp3 silenced by shRNA. Silencing of Foxp3 by shRNA could up-regulate the expression of IL-17,IFN-γin effector T cells.Conclusions:These findings offer evidence that Foxp3 express in all hepatocellular carcinoma cells. The biological significance of these findings indicates Foxp3 expression and down stream genes regulated by Foxp3 maybe involved in tumor cell mediated immune-suppression.