Investigation Of Impacts Of Yan-Hou-Qing Formula On P38/Erk/NF-?B/COX-2 Pathway In Treatment Of Acute Pharyngitis

Objective:Although the curative effects on pharyngitis have been validated in clinics,the detailed molecular mechanisms of Yan-Hou-Qing?YHQ?formula on pharyngitis was still unclear.This study was designed to explore the anti-inflammation effects and underlying mechanisms of YHQ formula in acute pharyngitis rat model.Methods:?1?48 SD rats?180-220 g?,SPF grade,randomly divided into four groups:normal control group?NC,12 rats?,acute pharyngitis model group?AP,12 rats?,Yan-Hou-Qing formula treatment group?YHQ,12 rats?and dexamethasone treatment group?Dex,12 rats?.?2?Acute pharyngitis rat models were established via spraying with15%ammonia in pharynx for three consecutive days in AP,YHQ and Dex groups.Instead of 15%ammonia,normal saline was used to stimulate the pharynx in NC group.On the fourth day of the experiment,the rats in YHQ group and Dex group were oral administrated with Yan-Hou-Qing formula?0.72 g/kg?and dexamethasone?5 mg/kg?,respectively.NC and AP group were given equal volume of normal saline.?3?After continuous treatment for 5 days,the behavioral changes and pharyngeal apparent index of rats in each group were observed and scored after the last gavage therapy.To observe the permeability of pharyngeal vascular endothelial cells,6 rats in each group were randomly selected for tail vein injection of Evans blue dye solution?30 mg/kg?.After kill the remaining 6 rats,pharyngeal tissues were collected for further hematoxylin-eosin staining?H&E staining?.The morphological structure of pharyngeal tissue and the infiltration of inflammatory cells were observed under light microscope.?4?Pro-inflammatory cytokines and mediators?IL-1?,TNF-?,IL-6 and PGE₂?in pharyngeal tissue were determined by enzyme linked immunosorbent assay?ELISA?.?5?The protein expressions of p38,Erk,JNK,IKK?,I?B?,NF-?B p65 and p-p38,p-Erk,p-JNK,p-IKK?,p-I?B?,p-NF-?B p65,COX-2 protein in pharyngeal tissue were detected by Western blot.Results:?1?Compared to the NC group,the rats in AP group exhibited acute phargitis-related symptoms and signs,and the behaviors scores of acute pharyngitis were apparently higher?P<0.001?in AP group,indicating the AP rat models were established successfully.Atfter administration with YHA and dexamethasone,acute phargitis-related symptoms and signs of YHQ group and Dex group decreased significantly,comparing with AP group?P<0.001?.?2?Based on observsions results via light microscope,the pharyngeal tissue were covered by thickened,keratinized and exfoliated mucosa epithelium,dilated and bleeding blood vessels.Infiltrated inflammatory cells and the hypertrophied grand were also found in pharyngeal mucosa of AP group.After treatment,both Yan-Hou-Qing formula and dexamethasone could improve the mucosa thickening,reduced the inflammatory cells infiltration,and recovered the grand to normal.?3?Compared with NC group,the permeability of pharyngeal vascular endothelial cells in AP group was significantly increased?P<0.001?;the permeability of vascular endothelial cells was alleviated after YHQ treatment?P<0.05?,and the symptom of Dex group was improved?P<0.001?notably.?4?Compared with NC group,the inflammatory cytokines of IL-1?,TNF-?,IL-6 and PGE₂ in AP group were up-regulated significantly?P<0.001?,and these up-regulated inflammatory cytokines were reversed?P<0.05 compared with AP group?after treatment with Yan-Hou-Qing formula and dexamethasone.?5?Compared with NC group,the phosphorylated protein expressions of p-p38,p-Erk,p-JNK,p-IKK?,p-I?B?,p-NF-?B p65 and COX-2 in pharyngeal tissue of AP group rised significantly?P<0.001?,and the higher expression of COX-2 and phosphorylated protein expressions of p-p38,p-Erk,p-IKK?,p-I?B?,p-NF-?B p65 were down-regulated by Yan-Hou-Qing formula?P<0.05?.After treatment of dexamethasone,the expressions of p-p38,p-Erk,p-JNK,p-IKK?,p-I?B?,p-NF-?B p65 and COX-2 also decreased significantly?P<0.01?.Conclusion:?1?Yan-Hou-Qing formula can improve the permeability of pharyngeal vascular endothelial cells,maintain the integrity of pharyngeal mucosa,and improve the symptoms and signs of acute pharyngitis via inhibiting the release of inflammatory cytokines?TNF-?,IL-1?and IL-6?and the production of PGE₂.?2?The therapeutic effects of Yan-Hou-Qing formula in treating acute pharyngitis is associated with inhibition of the p38/Erk/NF-?B/COX-2 inflammatory signal pathway.