Effect of Activated Murine FcγRⅡb on the Production of Innate Immune Factors
Objective:To study the effect of FcγRⅡb on innate and antiviral immunity of macrophages in mice.Methods:The recombinant vector pET28a-FcγRⅡb was constructed and transformed into BL21 competent cells for expression induction.The recombinant protein was purified by Nickel affinity chromatography.The purified protein was then used as an antigen to immunize rabbits for polyclonal antibody production.The serum titers were detected by indirect ELISA,and the specificity was determined by Western blotting.Then,the rabbit polyclonal antibody against mouse FcyRⅡb was obtained with saturated ammonium sulfate solution and protein G sapharose.Thus,the antibody was used to activate the FcγRⅡb on mouse peritoneal macrophages,and the mRNA transcript levels of cytokines in MHV-infected cells were detected by qPCR.Results:The activated FcγRⅡb could up-regulate the mRNA transcript levels of TLR3,IRF3,IFN-α,IFN-β,IFN-γ,TNF-α and IL-1β in macrophages at 12-48 hours,and down-regulate the transcript level of TGF-β mRNA.In addition,after MHV infection with FcγRⅡb activated cells for 12-48 hours,the mRNA transcript levels of TLR3,IRF3,IFN-α,IFN-β,IFN-γ,TNF-α and IL-1β were up-regulated and the mRNA transcription levels of TGF-β was down-regulated in the FcγRⅡb activated group,compared with the non-activated group.In addition,the viral copy numbers and viral titers of MHV in the FcγRⅡb activated group were much lower than those in the non-activated group.Conclusion:This study demonstrates that activation of murine FcγRⅡb can effectively enhance the production of the innate immunity related cytokines in macrophages and inhibit virus replication.It provides a basis for further investigation of the role of FcγRⅡb in MHV infection.
FcγRⅡbPolyclonal antibody preparationMouse hepatitis virusImmune related cytokinesInnate immune
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