Effects Of Imidacloprid On IgE-mediated And TLR2 Mediated Mast Cell Activation

Imidacloprid,1-[?6-chloro-3-pyridinyl?methyl]-N-nitro-2-imidazolidinimine,HS A been used as an pesticide for crop protection worldwide over the last decade.The residue of it continue to be found in the environment,foods,and in human tissues.There are studies focusing on the effects of it's residues on human health,but few information about the relationship between imidacloprid and allergy such as IgE-mediated food allergy,is available;Mast cell express toll-like receptor 2?TLR2?and play an important role in immune defense against bacteria and viruses.Recent advances showed that imidacloprid can suppress adaptive and inflammatory immune responses in BALB/c or rats,with them being particularly susceptible to pathogens.However,there is no information about the possible effects of imidacloprid on TLR2-dependent activation of mast cells;It is now increasingly recognized that simultaneous exposure of humans to different chemicals including pesticides from various sources and with different but specific immunological effects become a reality,and that there are more attention being given in health care science.However,little information about the effects of both pesticides together,such as imidacloprid and endosulfan on mast cell degranulation is available.In this proposal,the IgE-activated mast cell in vitro,such as IgE-mediated mast cell activation challenged by?-milk globulin,and passive systemic anaphylaxis in mice were treated with imidacloprid Then,the preformed granule-associated chemical mediators will be determined.These results provide fundamental knowledge about the effects of imidacloprid on the IgE-mediated activation of mast cells and further explore the mechanisms by which imidacoprid affect IgE-mediated activation;On the other hand,the effects of imidacloprid on the TLR2-mediated activation of mast cell was determined in this proposal,in order to provide data about the possibility that imidacloprid affect the host mmune response via affecting the TLR2-mediated activation of mast cells.Finally,the combined effects of equi-molar mixtures of endosulfan and imidacloprid on mast cell was tested in IgE-activated bone marrow-derived mast cells?BMMCs?,in order to demonstrate the combined effects of multiple pesticides on allergic diseases such as food allergy.The approaches,results and conclusions were depicted as follows.1.The model of the IgE-mediated activation of mast cell in vitro and the model of passive systemic anaphylaxis in mice were established and treated with imidacloprid.Then,the allergic mediators,such as histamine,?-hexosaminidase??-hex?,leukotrienes C4?LTC4?,TNF-?,IL-6,Ca²⁺influx in mast cells and vascular extravasation in IgE-induced PCA of mice were tested.It was shown that 10^-3-10^-99 M imidacoprid suppressed the production of histamine,?-hex,LTC4,IL-6 and TNF-?,Ca²⁺mobilization in IgE-activated RBL-2H3 cells and decreased vascular extravasation in IgE-induced PCA and this effect was dose-dependent.It is the first time to show that imidacloprid suppressed the IgE-mediated activation of mast cell.2.Based on the above the results,the model was furtherly treated?7 nAchR antagonists and imidacloprid in sequence.Then,the allergic mediators,such as histamine,?-hex,LTC4,TNF-?,IL-6,Ca²⁺influx,the phosphorylation antibodies such as p-PLC-?and p-NF-?B in mast cell and vascular extravasation in IgE-induced PCA of mice were tested.The result showed that 10^-33 M MLA alleviated the imidacloprid-induced inhibition on the production of allergic mediators,Ca²⁺mobilization,cPLA2 activity and the expression of the phosphorylated antibodies of PLC-?and NF-?B in IgE-activated degranulation of RBL-2H3 and KU812 cells and IgE-activated PCA in mice.It is the first time to demonstrate that?7-nAChRs involve the inhibitory effects of imidacloprid on the IgE-mediated activation of mast cells.3.The model of the TLR2-mediated activation of mast cell in vitro and in vivo were established and treated with imidacloprid.Then,the allergic mediators,such as histamine,?-hex,LTC4,TNF-?,IL-6,Ca²⁺influx,the phosphorylation antibodies such as p-PLC-?and p-NF-?B in mast cell and TLR2-induced skin inflammation in BALB/c mice were tested.The result showed that 10^-3-10^-5 M imidacloprid markedly suppressed the production of inflammatory cytokines TNF-?and IL-6 in TLR2/TLR1-dependent activation of RBL-2H3 cells and skin inflammation of BALB/c mice,but it had no effects on the TLR2-dependent RBL-2H3 cell degranulaion.Our results implicated that imidacloprid inhibited the late stage of TLR2-mediated mast cell activation.4.The IgE-mediated mast cell in vitro and passive systemic anaphylaxis?PSA?in mice were treated with equi-molar mixtures of endosulfan and imidacloprid.Then,the allergic mediators,such as histamine,?-hex,LTC4,IL-6,Ca²⁺influx,the phosphorylation antibody p-NF-?B in mast cell,vascular extravasation in IgE-induced PSA of mice and the serum levels of histamine,?-hex,LTC4,IL-6 were tested.It was shown that the application of equi-molar mixtures of both pesticides with 10^-4-10^-55 M significantly inhibited the IgE-mediated mouse bone marrow-derived mast cells degranulation in vitro.Morever,10^-44 M of them decreased vascular extravasation and the serum levels of histamine,?-hex,LTC4,IL-6 in IgE-induced PSA,as the application of imidacloprid at the same concentration alone did.In conclusion,simultaneous application of equi-molar concentrations of both pesticides generally showed highly similar responses compared to the responses to imidacloprid alone,suggesting that the effects of the mixture could be solely attributed to the effects of imidacloprid.5.This study mainly clarified the effects and mechanism of imidacloprid on degranulation of mast cell,providing a scientific basis for evaluating the effects of imidacloprid on the allergy.and host defense mediated by mast cell degranulation,furtherly providing more theoretical guidance for assessing the residues of pesticides on allergic diseases such as food allergy.