The Effect And Molecular Mechanism Research Of Daphnetin In Immunoglobulin(Ig) E-mediated Type I Anaphylaxis

Objective:In this research, the effects and possible molecular mechanism of Daphnetin on Ig E-stimulated Type I Anaphylaxis were detected by Passive Cutaneous Anaphylaxis of Balb/c mice in vivo and RBL-2H3 cells in vitro.Methods:MTT assay was performed to establish the effect of DAP on RBL-2H3 cells viability. The inhibitory effect of DAP on the release of β-HEX in Ig E-stimulated RBL-2H3 cells was detected. ELISA assay was ran to evaluate the effects of DAP on the product of histamine. The fluorescence microscope was used to measure the effect of DAP on Ig E-induced intracellular Ca2+ mobilization in RBL-2H3 cells. P-Lyn/Lyn, P-Syk/Syk and phosphor-MAPK protein expression of Ig E-induced RBL-2H3 after treated with Dap were assayed by Western blot analysis. According to the absorbance of Evans Blue leakage of Balb/c mice ears to determine the anti-hypersensitive effect of DAP.Results:1. Daphnetin(25 ~ 200 mg/kg) got an affective inhibition on the absorbance of Evans Blue leakage of Balb/c mice Passive Cutaneous Anaphylaxis Model.2. Daphnetin(1.68 ~ 56 μM) got no signi?cant influence on cell viability compared to control group.3. Daphnetin(1.68 ~ 56 μM) significantly decreased the release of β-HEX in Ig E-stimulated RBL-2H3 cells with a dose dependent manner.4. Daphnetin(5.6 ~ 56 μM) remarkably reduced the release of histamine in Ig E-stimulated RBL-2H3 cells.5. Daphnetin(5.6 ~ 56 μM) significantly reduced the mobilization of intracellular Ca2+ in Ig E-stimulated RBL-2H3 cells with a dose dependent manner.6. Daphnetin(5.6 ~ 56 μM) significantly decreased the quantities of Lyn, Syk, JNK, ERK, p38 phosphorylated protein in Ig E-stimulated RBL-2H3 cells with a concentration-dependent manner.Conclusion:Daphnetin(25 ~ 200 mg/kg) could regular the blood leakage in chronic allergic response of Balb/c mice Passive Cutaneous Anaphylaxis model. Daphnetin(1.68 ~ 56 μM) significantly inhibited the release of β-HEX and histamine in Ig E-stimulated RBL-2H3 cells. The molecular mechanism may be related to the regulation of intracellular Ca2+ mobilization and Ig E/FcεR I mediated P-Lyn/Lyn, P-Syk/Syk, MAPK signalling transduction pathway. Meanwhile,...