| Objective: Tumor malignancy is one of the major diseases that threaten human life and health.There is an urgent need to develop new strategies to treat human cancer.Dendritic cell(DC)-based immunotherapy is a promising approach for cancer therapy.DCs are the most effective antigen-presenting cells(APCs)and play an important role in the induction of anti-tumor immunity.We and other researchers have found that Th9 cells secreting IL-9 have a strong antitumor activity in mice.Therefore,development of DC vaccines that specifically induce and amplify Th9 cells will strongly enhance the antitumor effect of DC vaccines.Our recent study shows that Dectin-1 signal activated dendritic cells(Dectin-1-DCs)can potently promote Th9 cells in vitro but their mechanisms need to be further elucidated.We found that Dectin-1-DCs express abundant IL-33,but the role of IL-33 in Dectin-1-DCs in the induction of Th9 and antitumor immune responses is unclear.IL-33 is a multifunctional cytokine.Studies have shown that IL-33 promotes CD8+T,ILC2,Th2 and Th1 cell responses which are beneficial for antitumor immunity and the generation of Tregs which are not conducive to antitumor immunity.In this study,we will further explore the role of IL-33 in Dectin-1-DCs inducing Th9 and anti-tumor immunity and its intrinsic mechanism.Methods: First,we used gene expression profile(GEP)to analyze up regulated genes in Dectin-1-DCs and IL-33 was found to be significantly up regulated.To verify that dectin-1 signaling promotes DCs to express IL-33,we generated BMDCs and Dectin-1-DCs: Cur DCs and Scl DCs in vitro from C57BL/6 bone marrow-derived DCs,the m RNA and protein levels of IL-33 were detected by QPCR,ELISA and FCM.We used the same method to verify whether the levels of IL-33 were decreased after knockdown the gene dectin-1.In order to study whether or not the primary anti-tumor effector cells induced by Dectin-1-DCs were Th9 cells,we detected the expression of Th9,Th1 and Th17 cytokines and transcription factors in CD4+T cells from spleen of OT2 mice which immunized with DCs by FCM,ELISA and QPCR.To study how Dectin-1-DCs induce Th9 cells,we detected the expression of ST2 in CD4+T cells from spleen of OT2 mice or C57BL/6 which immunized with DCs by FCM and QPCR.To further verify that Dectin-DCs promote Th9 differentiation through ST2,ST2 blocking antibody was added and detected the effect of blocking ST2 on the production of Th9 cells by Dectin-1-DCs.Since the secretion of IL-33 by Dectin-1-DCs is very low,in order to study the effect of IL-33 on the induction of Th9 by Dectin-1-DCs,we detected the m RNA and protein levels of ST2 and Th cytokines in CD4+T cells from spleen of OT2 mice which immunized with DCs with exogenous IL-33 by FCM,ELISA and QPCR.Then,to study whether IL-33 could promote the anti-tumor immunity of Dectin-1-DCs,we used B16-OVA melanoma-OT2 mouse as the model,and immunized with DCs.Finally,as a control,we also examined whether IL-33 combined with BMDCs contributes to the induction of Th9 and anti-tumor immune responses.Results: Dectin-1 signaling promotes the expression of IL-33 by DCs.Dectin-1-DCs can induce a large number of Th9 cells and a certain amount of Th1 cells in vivo,in which Th9 cells predominate.Dectin1-DCs promoted the production of ST2 by CD4+ T cells,and blocked ST2 significantly inhibited the induction of Th9 cells by Dectin1-DCs.Exogenous IL-33 significantly enhanced the production of Th9 by Dectin-1-DCs but had no effect on the induction of Th9 and its anti-tumor effect by BMDCs.Addition of exogenous IL-33 significantly increased the antitumor effect of Dectin-1-DCs.However,exogenous IL-33 promoted BMDCs to induce Treg cells, inhibited the expression of IL-2 in CD4+ T cells induced by BMDCs and promoted the expression of immunosuppressive molecule PD-1 in T cells,thereby inhibiting the anti-tumor effect of BMDCs.Conclusion:IL-33 promotes Dectin-1-DCs to induce Th9 cells in vivo and further enhances the antitumor immune response of Dectin-1-DCs in vivo. |
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