Professor Miao Qing's Experience In Treating Cough Variant Asthma From Cold And Mechanism Research Of Jinfei Zhisou Powder Regulates TRPA1

Background:Cough variant asthma is mainly manifested as chronic cough,cold and pungent smell could induce cough,even make cough aggravated.Modern medicine believes that the pathogenesis of the disease is similar to typical asthma,the treatment is the same as asthma,such as the control of airway inflammation,the dilatation of bronchus,the antagonism of leukotriene.However,some patients after anti-asthma treatment,cough is still poorly controlled,for CVA patients,chronic cough influences their working,learning,life and psychological activities.Professor Miao Qing,who has long been engaged in clinical,teaching and scientific research in traditional Chinese medicine,using TCM to treat CVA,has rich experience,and attaches importance to the role of cold in the development and evolution of CVA,and puts forward from cold to treat the disease,the results are remarkable.Through three years of learning with the tutor,under the careful guidance of the tutor,summed up the experience of the tutor from cold to treat CVA and chose Jinfei Zhisou powder applied to experimental research,from the perspective of TRPA1,explore the mechanism of the correlation between CVA and cold,the curative effect and molecular mechanism of Jinfei Zhisou powder in the treatment of CVA guinea pigs.Part I Literature ReviewThis part put cough,chronic cough,recorded in Chinese medical book,the understanding and treatment of CVA in TCM on modern literature,dividing the cough into wind cough,cold cough,hot cough,dry cough,hot and humid cough,phlegm stasis cough,virtual cough,Yu cough to review.Part ? Professor Miao Qing's Experience in Treating Cough Variant Asthma from ColdObjective:To summarize the clinical experience of the tutor in treating CVA from cold,and to deeply understand the tutor's thoughts on the pathogenesis,treatment and medication caused by the cold.Methods:By studying with tutor in out-patient department,ward,sorting out tutor's validity case,asking the tutor questions,combining Chinese medical book and modern literature research,summing up the tutor's thought of treating CVA from cold,the rule of treatment and the choice of drug use,attaching the validity case to proof.Results:The idea of the tutor's treatment of CVA from cold was derived from the intensive reading and deep understanding of the classical Chinese medical book such as Huangdi Neijing,General Treatise on the Cause and Symptoms of Diseases,and Medical Insights.Attaching importance to the difference of cold way in patients with CVA,it is concluded that the cold of six exopathogens in nature,the cold of living condition,the cold of diet and drugs,and the cold of yang deficiency are common ways to feel cold.In the treatment,tutor scrutinizes the difference between the pathogenesis,the disease location,and the viscera,puts forward the classification of tri-jiao viscera.cold in the upper-jiao,Jinfeicao powder or Sanao decoction are common prescription,Zhisou powder is suitable for patients with micro cold,Ephedra decoction is suitable for patients with heavy cold,patients with the cold and rheum could use Xiaoqinglong decoction,Linggui Zhugan decoction could alleviate the uprush of water and qi.Cold in the middle-jiao,Lizhong decoction is suitable for patients with the deficiency-cold in spleen and stomach,patients with the simultaneous occurrence of cold and heat could uses Banxia Xiexin decoction,patients with the liver depression and spleen cold could use Chaihu Guizhi Rhizoma zingiberis decoction.Cold in the lower-jiao,warming the kidney is important,tutor often pays attention to the application of the Er-xian decoction.At the same time,tutor pays attention to the transformation and change of cold in the tri-jiao,emphasizes the prevention before diease onset,preventing disease from exacerbating.Conclusion:Professor Miao Qing believes that we should pay attention to CVA patient's cold path,although eliminating cold to stop cough is important,distinguish the difference between tri-jiao viscera is also cannot be ignored.Part III Mechanism Research of Jinfei Zhisou Powder Regulates TRPA1Objective:To explore the mechanism of the correlation of CVA and cold,the curative effect and mechanism of Jinfei Zhisou powder in the treatment of CVA guinea pigs by the influence of TRPA1 on the activity of human bronchial epithelial cells stimulated by LTD4 combined IL-4 in vitro experiment,and Jinfei Zhisou powder regulates TRPA1 in vivo experiment,in order to provide molecular biological evidence support for clinical application of Jinfei Zhisou powder in the treatment of CVA.Method:1 The influence of TRPA1 on the activity of human bronchial epithelial cells stimulated by LTD4 combined IL-4 in vitro experiment:human bronchial epithelial cells(BEAS-2B)were randomly divided into 6 groups,blank control group,model group,HC-030031 group(TRPA1 antagonist)(0.01?mol/L),HC-030031 group(O.1?mol/L),HC-030031 group(1?mol/L),HC-030031 group(10?mol/L).In addition to the blank control group,this experiment uses LTD4 and IL-4 to stimulate the rest of the human bronchial epithelial cells refer to literature,completes the mold.The cells in blank control group and the model group were cultured with serum-free culture base,and the corresponding concentration of HC-030031 was added to the medication administration group,the cell activity of each group was detected by MTT method.2 Jinfei Zhisou powder regulates TRPA1 in vivo experiment:84 healthy male guinea pigs were randomly divided into 7 groups according to the equilibrium stochastic method:blank control group,model group,glucocorticoid group(prednisone acetate tablets),TRPA1 antagonist group(HC-030031),low,medium and high dose group of Chinese herbal medicine(Jinfei Zhisou powder of different dose gradients).In addition to blank control group,the guinea pigs of rest of the group,according to literature and previous experimental research experience to establish guinea pig CVA model:day 1,guinea pigs were intraperitoneally injected cyclophosphamide 30mg/kg;2 days later,guinea pigs were intraperitoneally injected 2mg of ovalbumin and 100mg of aluminum hydroxide(4%aluminum hydroxide gel 2.5ml),initial sensitization;after 3 weeks,boosting was carried out by intraperitoneal injection of 0.01mg ovalbumin and 100mg aluminum hydroxide(4%aluminum hydroxide gel 2.5ml);3 weeks after the boosting,with lOmg/ml ovalbumin solution aerosolized 90s to stimulate cough,before inhaled ovalbumin about 30min,guinea pigs were intraperitoneal injection of diphenhydramine 20mg/kg.The challenging of ovalbumin once every other day,7 times in total,establish guinea pig CVA model.At the same time,guinea pigs of blank control group were given physiological saline in the same volume.Guinea pigs of TRPA1 antagonist group inhaled 0.3mmol/L HC-030031 solution by a nebulizer for 10min before the detection of the indicators;the rest of guinea pigs of the medication administration group were orally given corresponding Chinese herbal medicine and glucocorticoid the day before starting ovalbumin challenging for 14 consecutive days.After reaching the drug delivery cycle,the bronchoalveolar lavage fluid and lung tissue were obtained.Wright's staining,observing and calculating EOS%in bronchoalveolar lavage fluid of guinea pigs under oil immersion lens.Detecting the expression of TRPA1 in guinea pigs' lung tissue by Western blotting.Hematoxylin eosin staining,observing the pathological morphological changes of lung tissue in guinea pigs under microscope.Results:1 The influence of TRPA1 on the activity of human bronchial epithelial cells stimulated by LTD4 combined IL-4 in vitro experiment:Compared with blank control group,the activity of human bronchial epithelial cells decreased significantly after the joint intervention of LTD4 and IL-4 in the model group(P<0.05).Compared with model group,10?mol/L HC-030031 can significantly improve the inhibition of cell activity by LTD4 and IL-4(P<0.05).The other concentration gradients HC-030031 group,there was a increasing tendency of the activity of human bronchial epithelial cells in different degrees(P>0.05).2 Jinfei Zhisou powder regulates TRPA1 in vivo experiment:(1)Behavioral characteristics of guinea pigs in each group:there were no special behavioral abnormalities in guinea pigs in the blank control group;after intraperitoneal injection of ovalbumin in the model group,guinea pigs scratched the nose,restless,breathed faster,and the abdominal limbs twitched slightly;after the challenging of ovalbumin,the guinea pigs showed shortness of breath,cough and so on;with the increase of the strength of molding,the hair of guinea pigs was gradually sparse and loose,the gloss was poor,and the weight was reduced to different degrees.In the medication administration group,the behavioral performance of guinea pigs improved compared with that of the model group.(2)The results of EOS%in bronchoalveolar lavage fluid of guinea pigs:Compared with blank control group,EOS%in bronchoalveolar lavage fluid of guinea pigs in model group increased significantly(P<0.05);compared with model group,EOS%in each medication administration group decreased in different degrees(P>0.05);the decline trend of glucocorticoid group,low and high dose group of Chinese herbal medicine were the most obvious,followed by medium dose group of Chinese herbal medicine,the decline trend of EOS%in TRPA1 antagonist group was not obvious(P>0.05).(3)The results of TRPA1 expression in lung tissue of guinea pigs:Compared with blank control group,there was an increasing trend of TRPA1 expression in lung tissue of guinea pigs in the model group(P>0.05),and compared with model group,the expression of TRPA1 in lung tissue of each medication administration group decreased in different degrees(P>0.05).Among them,the decrease trend of glucocorticoid group was the most obvious,followed by the low dose group of Chinese herbal medicine,the trend of the high dose group of Chinese herbal medicine and TRPA1 antagonist group were relatively not obvious(P>0.05).(4)Pathological morphological results of lung tissue in various groups of guinea pigs:The pathological morphology of lung tissue in the blank control group showed that the bronchial and alveolar structures were clear and complete,the mucosal epithelium of the lumen was intact,there were no shedding cells,there was no obvious inflammatory cell infiltration,the alveolar septum of the model group was widened,the inflammatory cells infiltrated,and compared with model group,the infiltration of different levels of inflammatory cells in the bronchial tube of each dosing group decreased.Conclusion:(1)TRPAl is involved in the process of eosinophilic chronic airway inflammatory of CVA.(2)Jinfei Zhisou powder has the trend to reduce the EOS%in bronchoalveolar lavage fluid of guinea pigs and the TRPA1 expression in lung tissue.(3)Jinfei Zhisou powder has the effect of alleviating the infiltration of inflammatory cells in lung tissue of CVA guinea pigs.