Protective Effect And Mechanism Of Imperatorin On Airway Inflammation In Mice With Allergic Asthma

Objective1.To investigate the effects of imperatorin on mast cell-mediated allergic responses in vitro and in vivo.2.To investigate the effects and mechanism of action of imperatorin on airway inflammation and airway remodeling in asthmatic models in vitro and in vivo.3.To explore the antioxidant effects and related mechanisms of imperatorin on asthmatic models in vitro and in vivo.Methods1.A mouse model of passive skin allergic reaction(PCA)was established and histological examinations were performed.The degranulation,histamine release and calcium influx levels of imperatorin in mast cells were observed by in vitro experiments.The levels of cytokines and the key proteins of Fc?RI-mediated signaling pathway and NF-?B signaling pathway were detected by ELISA and Western blot.2.Asthmatic mouse model was established and the pathological changes of mice lung tissue were examined by HE,PAS and Masson staining.Diff-Quick staining counts total cells and eosinophilia(EOS)in mouse BALF;The levels of IL-4,IL-5,IL-13 and IFN-y were detected by ELISA.Airway hyperresponsiveness was detected by inhalation of acetylcholine.The positive area of a-smooth muscle actin(a-SMA),matrix metalloproteinase-9(MMP-9),p-STAT3,and NF-?B p65 in lung tissue were detected by immunohistochemistry.Western blot was used to quantitatively analyze the contents of a-SMA,MMP-9,and STAT3 and NF-?B signaling pathways key proteins.3.The content of reactive oxygen species(ROS)in BALF of asthmatic mice was detected by dihydrorhodamine(DHR)-123 method.The content of protein carbonyl and the expression level of malondialdehyde(MDA)in the lung tissue were detected.Antioxidant enzyme kit detected superoxide dismutase(SOD)activity and glutathione(GSH)and total antioxidant capacity(TAOC)in BALF.Immunohistochemistry and Western blotting were used to detect the expression of key proteins in MAPK/Nrf2/HO-1 signaling pathway in lung tissue and human bronchial epithelial(16HBE)cells.Results1.In the PCA model,the amount extravasation of Evans blue dye and the thickness of ear swelling were significantly decreased.Imperatorin could also reduce the degranulation,histamine release,and calcium influx in mast cells.In addition,the levels of TNF-?,IL-1?,IL-4,IL-8,and IL-13 in mast cells were significantly reduced.The expression levels of key proteins in the Fc?RI-mediated signaling pathway and the downstream NF-?B signaling pathway were significantly inhibited by imperatorin.2.Imperatorin reduced inflammatory cell infiltration,goblet cell proliferation,and collagen deposition.Imperatorin reduced the amounts of total cells and EOS in BALF,the content of IL-4,IL-5,IL-13,IFN-y and the airway hyperresponsiveness.Imperatorin obviously decreased the positive areas of a-SMA,MMP-9,p-STAT3 and NF-?B p65.Imperatorin obviously decreased the expression levels of a-SMA,MMP-9,and the key proteins in STAT3 and NF-?B signaling pathways.3.Imperatorin could reduce the amount of ROS,protein carbonyl and MDA and increase the amount of SOD,GSH and TAOC.Imperatorin obviously decreased the positive area of key proteins in the MAPK/Nrf2/HO-1 signaling pathway of lung tissue.Imperatorin obviously decreased the expression level of key proteins in the MAPK/Nrf2/HO-1 signaling pathway of lung tissues and 16HBE cells.Conclusions1.The antiallergic effect of imperatorin may be partly through inhibition of Fc?RI-mediated signaling pathway and downstream of NF-?B signaling pathway,and then it inhibits the degranulation of mast cells,releases histamine,calcium influx,and reduces the levels of inflammatory factors such as TNF-?,IL-1?,IL-4,IL-8,IL-13.2.Imperatorin reduces airway inflammation and airway remodeling in both in vitro and in vivo asthma models through STAT3 and NF-?B signaling pathways.3.Imperatorin maintains the balance of the oxidative/antioxidant system through the MAPK/Nrf2/HO-l signaling pathway,thereby playing a role in improving asthma.