Mast Cell Derived Exosomes Suppress Allergic Reaction By Binding To Free IgE

Objective: To analyze biological characteristics of mast cell derived exosomes,explore the influence of mast cell derived exosomes on mast cell activation by binding to free IgE,and study the anti-allergic effect in mouse models of allergic asthma.Methods:(1)Bone marrow-derived mast cells(BMMC)were generated from BALB/c mice femurs by incubation with IL-3 and SCF for 4 weeks and exosomes were isolated from the supernatant by differential centrifugation.The biological characteristics of exosomes were analyzed with transmission electron microscope,Western blot,flow cytometry and confocal laser scanning microscopy.(2)Flow cytometry were used to detect the combination of IgE to BMMCs after incubation of IgE with exosomes from mast cells.The suppression effect of BMMC-exosomes on MCs activation was examined by degranulation test,and activated-associated pathways were determined with Western blot analysis.(3)To explore the anti-allergic effect in mouse model of allergic asthma,mice were executed after theatment with BMMC-exosomes for 1month,2 months and 3 months,respectively.We assessed the AHR in mice,examined the OVA-sIgE in serum and histamine,Th1/Th2 cytokines in BALF using ELISA or Luminex,as well as inflammatory cells.Furthermore,the pathological changes in lung tissue of mice were observed.Results:(1)Mature BMMCs were obtained by culturing the progenitor cells from the bone marrow for 4 weeks and exosomes could be isolated from the supernatant of BMMCs.The electron micrographs of exosome-like vesicles revealed rounded structures with a size of approximately 50-80 nm.Exosomes derived from BMMCs were positive for the receptor protein FcεRI.We found that exosomes derived from either BMMCs or P815 cells were rapidly taken up by corresponding cells,a process that rised abruptly after 1hour and peaked after approximately 12 hours.(2)We demonstrated that exosomes from BMMCs expressed FcεRI and could bind to free IgE,which reduced the free IgE level in the supernatant,which then inhibited the binding of IgE to FcεRI on BMMC.We also observed the competitive effect of IgE binding between exosomes and BMMCs.As exosomes derived from P815 cells failed to show the ability of binding to IgE,it is believed that BMMC-exosomes bind to IgE via the receptor FcεRI.BMMC-exosomes could inhibit the release of pro-inflammatory mediators,such as β-hexosaminidase,and histamine from activated mast cells.Accordingly,binding of BMMC-exosomes to IgE in turn reduced their phosphorylation of PLCγ1 and PKC.(3)In mouse model of allergic asthma,treatment with BMMC esosomes significantly reduced the levels of serum OVA-sIgE and histamine,interleukin(IL)-4,IL-5,IL-13 and inflammatory cells in BALF,although the levels of IL-10 and IFN-γ rose at the same time.Moreover,we found that BMMCexosomes modulates not only airway inflammation and AHR,but also some parts of remodeling in chronic asthma,and positively correlated with treatment time.Conclusion: IgE is the central macromolecular mediator responsible for the progression of allergic reactions and has long been known as a therapeutic target for allergic disease.Several anti-IgE agents have been developed or are in development to target IgE directly.In this study,we demonstrate that exosomes derived from BMMCs can bind to free IgE via FcεRI.Thus,BMMC-exosomes possess anti-IgE effect,which decrease IgE levels and inhibit MC activation.In mouse model of allergic asthma,our results show that BMMC-exosomes modulates not only airway inflammation and AHR,but also some parts of remodeling in chronic asthma.It is concluded that BMMC-exosomes have the potential to be developed as a novel anti-IgE agent.