Investigation Of The Anti-tumor Mechanism Of Ficolin-2/A By Triggering Immune Cells In Vivo

Background:Human serum ficolins are a group of complement lectins with an overall structure similar to that of mannose-binding lectin(MBL)and Clq.These oligomeric lectins play a key role in first-line host defense.Three human ficolins,namely,L-ficolin/P35(FCN-2 or ficolin-2),M-ficolin(FCN-1 or ficolin-1),and H-ficolin/Hakata antigen(FCN-3 or ficolin-3),and two mouse ficolins(ficolins-A and-B)have been identified.Mouse ficolin-A is closely related to human ficolin-2,and they are mainly synthesized in the liver and then secreted into blood circulation.Ficolin-2 is characterized by the presence of a collagen-like and a fibrinogen(FBG)-likedomain.The FBG-like domain of ficolin-2 forms a globular structure and binds to sugar structures.Increasing studies have shown that ficolin-2 can bind to viral and bacterial glycoproteins to inhibit viral and bacterial infections.Furthermore,abnormal ficolin-2 expression plays a crucial role in various infectious diseases,including viral diseases and tuberculosis.A recent report showed that ficolin-3 directly attacked cancer cells via a novel complement pathway.The valuable insight of ficolin-2 in association with immune cells in the context of cancer remains elusive.Objective:We investigated the immune role and mechanism of ficolin-2 in several common human cancers,including colon cancer,lung cancer and hepatocellular carcinoma,in vitro and in vivo.Method:Ficolin-2 concentrations of serum samples from cancer patients and healty donors(HDs)were tested using a sandwich ELISA.To determine whether ficolin-2/A exerts protective effects against tumor cells growth in vivo and the immune cells involed in,tumor-bearing mice models and immune cell-depleted tumor-bearing mice models were builded.The binding abilities of the ficolin-2 to cells were detected by FCM.The release of cytokines from macrophages stimulated by ficolin-2 was detected by ELISA kits.The influences of cytokines released by macrophages stimulated by ficolin-2 on tumor cells were detected by Transwell.The influence of ficolin-2 on macrophage polarization was detected by RT-PCR.The effects of ficolin-2 treated APCs(macrophages and dendritic cells)on CD8+ T cells.Were detected using MLR(mixed lymphocyte reaction)assay.The expressions of Wnt 9a and the downstream moleculars of Wnt signaling pathway in ficolin-A knockout mice were tested by western blotting assay.Results:Serum ficolin-2 levels of cancer patients were significantly lower than healthy donors.Both ficolin-2 and ficolin-A inhibit CT26 cells and Lewis cells growth in vivo dependely on macrophages and CD8+T cells.Ficolin-2 protein bound to macrophages,CD8+ T cells and tumor cells in vitro.Ficolin-2 can tilt the balance from MO or M2 macrophages to M1 macrophages,increase the secretions of inflammatory cytokines.IL-6 and TNF-α are crucial cytokines secreted by macrophages that exert important inhibitory effects.Ficolin-2-activated APCs(macrophage and dendritic cell)facilitated enhanced antigen-specific cytotoxicity and proliferation of CD8+ T cell.TLR4 modified ficolin-2-stumilated macrophage releasing cytokines,Ml-biased polarization and ficolin-2 antitumor effects in vivo.The levels of the tumor-related molecules Wnt 9a and cyclin D1 were increased most notably in the liver and colon in FCN-AKO mice compared to WT C57BL/6 mice.Cornclusion:Serum ficolin-2 levels of cancer patients were significantly lower than healthy donors.Ficolin-2 inhibited tumor cell proliferation in vitro via enhancing the macrophage Ml-biased polarization response and pro-inflammatory cytokines production.Ficolin-2 enhanced the antigen-specific activation of CD8+ T cells via macrophages and dendritic cells.Ficolin-2 exerts antitumor effects by interacting with TLR4 on the surface of macrophages.FCN-A knockout mice exhibited increased Wnt 9a signaling pathway activation.Our findings provide a new therapeutic strategy for tumors based on the triggering of immune-mediated antitumor effect by ficolin-2.