Traditional Korean Medicine Mahuang Dingchuan Decoction Relieving Cough Variant Asthma,An Airway Inflammation,In Guinea Pigs By Supressing NLRP3 Inflammasome

Objective:To observe effects of traditional Chinese Korean medicine Mahuang Dingchuan Decoction on airway inflammation in guinea pig models with cough variant asthma（CVA）,and to determine whether the mechanism related to the signal pathway of NLRP3.Methods:selecting 50 male guinea pigs,randomly putting them into 5 groups,being Controls,Models,Low Dosage（5mg）Mahuang Dingchuan Decoction,High Dosage（20mg）Mahuang Dingchuan Decoction and Dexamethasone（0.5mg）,with10 guinea pigs in each group.After one week’s care,expose all groups to smoke except for the Controls for the first 29 days,and perform muscle injection of 4%OVA and abdominal injection of 2%AL（OH）3 to cause allergic reactions on the 15^thand 22^ndday,and starting from the 29^thday,create CVA models by surrounding guinea pigs with vaporised 1%OVA solution.Use drugs to perform intragastric treatment for 30 minutes prior to each vapour session,and for Control,substitute the vapour with normal saline of the same amount.After the last session,use capsaicin aerosol to induce coughing,recording amount of coughs and delay time of coughs,examine high-level reaction in guinea pigs’airway;the following day,terminate the guinea pigs’lives,followed by sampling bronchial alveolar lavage fluid（BALF）and performing Diff-Quik stain in order to detect and tally up the amount of inflamed cells;perform HE,PAS,Masson stain and caspase-1 biopsy on lung tissues and observe pathological changes and post-biopsy changes in caspase-1;use ELISA test to determine the amount of relevant cytokine in BALF;use Western blot test to determine the NLRP3、caspase-1、ASC、IL-18、IL-1βprotein levels in the lung tissues;use immunohistochemical method and immunofluorescence method to observe the distribution of NLRP3 in airway’s surrounding areas.Results:1.Effects of Mahuang Dingchuan Decoction on the amount of coughs and delay time of coughs from the guinea pigs:compared to the Controls,the Models had earlier onset of coughing which is at incremental frequency,compared to the Models,both Low and High Dosage Mahuang Dingchuan Decoction Groups as well as Dexamethasone group exhibited various levels of improvement in how early coughing started and how frequently it occurred.2.Effects of Mahuang Dingchuan Decoction on the amount of inflamed cells and relevant cytokine in BALF:compared to the Controls,the Models showed different levels of increase in the amount of inflamed cells and cytokines（IL-1β、IL-18、IL-17A、IL-22、IL-23）,compared to the Models,there was a notable decline in the amount of inflamed cytokines（IL-1β、IL-18、IL-17A、IL-22、IL-23）in the BALF in both Low and High Dosage Mahuang Dingchuan Decoction Groups as well as Dexamethasone group.3.Findings in HE stain:in the Models group,the dominance of a significant amount of inflamed cells is seen in the bronchial mucosa,some alveoli collapsed or enlarged;compared to the models,both Low and High Dosage Mahuang Dingchuan Decoction Groups as well as Dexamethasone group demonstrated a noticeable ability to cure the above-mentioned symptoms.4.Findings in PAS stain:in the Models group,goblet cell overgrowth was seen on the airway accompanied with high levels of mucus secretion;the amount of positive cells and mucus secretion dropped more or less in both Low and High Dosage Mahuang Dingchuan Decoction Groups as well as Dexamethasone group.5.Findings in Masson stain:in the Models group,smooth muscles in the airway thickened,there was a considerable amount of deposit of collagen;compared to the Models,both Low and High Dosage Mahuang Dingchuan Decoction Groups as well as Dexamethasone group lessened collagen deposit in the airway and reduced the thickening of smooth muscles in the airway.6.Results of Western blot test:compared to the Controls,higher levels of NLRP3、caspase-1、ASC、IL-18、IL-1βproteins in the lung tissues was seen in the Models;compared to the Models,both Low and High Dosage Mahuang Dingchuan Decoction Groups as well as Dexamethasone group lowered the NLRP3、caspase-1、ASC、IL-18、IL-1βprotein levels in the lung tissues.7.Results of immunohistochemical test:compared to the Controls,a large amount of cells in the surrounding areas of the airway turned yellow-brown in the Models;compared to the Models,both Low and High Dosage Mahuang Dingchuan Decoction Groups as well as Dexamethasone group had less cells turning yellow-brown.8.Results of immunofluorescence:compared to the Controls,there was a noticeable increase in fluorescence intensity of NLRP3 in the Models;compared to the Models,both Low and High Dosage Mahuang Dingchuan Decoction Groups as well as Dexamethasone group exhibited a clear decrease in fluorescence intensity.9.Findings in caspase-1 biopsy:compared to the Controls,there was a clear increase in caspase-1 activity;compared to the Models,both Low and High Dosage Mahuang Dingchuan Decoction Groups as well as Dexamethasone group showed a clear downward trend in caspase-1 activity.Conclusion:1.Guinea pigs with cough variant asthma have noticeable symptoms of airway inflammation and airway remodeling,compared to normal guinea pig,higher levels of NLRP3 proteins in the lung tissues was seen in CVA Models.2.Mahuang Dingchuan Decoction is effective in reducing cough count and frequency in guinea pigs with cough variant asthma,its mechanism of action may be to inhibit the activation of NLRPS inflammasome which in turn reduces airway inflammation in guinea pigs with cough variant asthma.3.High-dosage Mahuang Dingchuan Decoction group generated better treatment outcomes compared to low-dosage group,it worked best in inhibiting the release of NLRP3 proteins and relevant factor of Th17 cells.