Qiweiduqi Pills Recipe Is Based On MKK3/6-p38MAPK Signaling Pathway To Interfere With The Efficacy And Mechanism Of COPD In Stable Phase

Purpose:1.To evaluate the efficacy and safely of traditional Chinese medicine compound in the treatment of chronic obstructive pulmonary disease(CQPD)in stable phase based on systematic evaluation and meta analysis.2.Based on the data mining to explore the understanding and medication rule of conte mporary doctors for COPD.3.Based on the network pharmacology to analyze the possible mechanism and potenti al targets of Qiwei Duqi Pills in the treatment of stable COPD.4.To explore the therapeutic effect and mechanism of modified Qiwei Duqi Pills on st able COPD by animal experiments based on MKK3/6-p38 MAPK signal pathway.Material and method:1 Systematic evaluation and meta analysis of the efficacy and safety of traditional Chinese medicine compounds in the treatment of stable COPD at stable stage.Searching for randomized controlled trials about traditional Chinese medicine combined with western medicine in the conventional treatment of chronic obstructive pulmonary disease in 6 electronic databases.For the retrieved literature,two independent researchers will screen the literature,extract the data,evaluate the quality of the literature and evaluate the risk of bias.If there are differences,they will be settled through negotiation;otherwise,the third researcher will make a decision.Using Revman 5.3 software to carry out meta-analysis on total effective rate,lung function(FEV1%、FVC、FEV1/FVC),blood gas,TCM symptom scores,etc.2 Based on data mining technology to explore the prescription regularity of traditional Chinese medicine compound in the treatment of stable COPD.By utilizing frequency analysis,factor analysis,cluster analysis,association analysi s and other data mining methods to explore the basic principles,common methods,dr ug selection and compatibility of stable COPD.3 Based on network pharmacology to explore the mechanism about how modified Qiw eiduqi Pills works on stable COPDUsing TCMSP and BATMAN-TCM databases to sdect the active ingredients and targets in modified Qiwdduqi Pills.The rdevant genes of COPD were obtained thro ugh OMIM and Genecards databases.Selecting the key genes between drugs and disea se,and then PPI network was constructed.Making an analysis about the result throng h GO functional enrichment analysis and KEGG pathway enrichment analysis.4 Explore the effect and mechanism of modified Qiweiduqi Pills on stable COPD has ed on mkk3/6-p38 MAPK signaling pathway4.1 Grouping and Dosing60 rats were randomly divided into blank group and modd group by using a random number table method,with 12 rats in blank group and 48 rats in modd group.In the modd group After the modd was successfully established,the modd rats were randomly divided into 3 groups,namely modd group,aminophylline group and traditional Chinese medicine group.The COPD rat modd was established by tracheal instillation of lipopolysaccharide combined with continuous smoking for 28 days.The next day after the end of modeling,gastric dosing was started.The blank group and the model group were given normal saline twice a day.The aminophylline group and the traditional Chinese medicine group were gavaged with corresponding medicines respectivdy,of winch the aminophylline group was gavaged twice a day,with 0.3125 mg/mL aminophylline suspension in the early stage and normal saline of the same volume in the late stage.The traditional Chinese medicine group was given 0.452 g/ml of traditional Chinese medicine twice a day.The volume of liquid medicine in each group was 1ml/100g,and the body weight of rats was weighed once a week.The dosage of liquid medicine was adjusted according to the change of body weight for 28 days.4.2 Get samplesSamples were taken on the 57th day after the modd was established.AH the rats were fasted but could drink for 12 hours before taking the drug.Put the rats to deal h by overdose of anesthesia{ 10%chloral hydrate),then get samples.Getting lung tiss ue and prepare alveolar lavage fluid.4.3 Research contentsHE staining was used to observe the pathological changes of lung tissue of rats in each group.The expression of IL-1β,TNF-α and IFN-γ in alveolar lavage fluid of rate was detected by Elisa method The expression of Bax/Bd-2,Fas/FasL,MKK3/p-MKK3,MKK6/p-MKK6 and p38 MAPK/p-p38 MAPK in rat lung was detected by Western-Blot;The mRNA transcription levels of Bax/Bd-2,Fas/FasL,MKK3,MKK6 and p38 MAPKmRNA were detected by RT-PCR.4.4 Statistical methodThe results were statistically analyzed by SPSS 19.0 software,and me measured data were expressed by the standard deviation(x±S)of mean addition and subtraction.The comparison between the two groups was tested by t test,and the comparison between multiple groups was analyzed by variance.P<0.05 was considered to be statistically significant.Results:1 Systematic evaluation and meta analysis of the efficacy and safety of traditional Chinese medicine compound in the treatment of stable COPD at stable stage.Compared with conventional western medicine treatment,the total effective rate of traditional Chinese medicine compound combined with conventional western medicine treatment in intervention of COPD is significantly increased.It can better improve some indexes of lung function:FEV1%,FVC,FEV1/FVC;Improving blood gas analysis indexes:increasing PaO2 value and decreasing PaCO2 value;Reduce the score of TCM symptoms:reduce the score of cough and expectoration;Improving quality of life:increasing 6MWT value;Reduce CAT score;Decrease the mMRC value;Lower the BODE index value-Besides,TCM compound were with good security.2 Based on data mining technology to explore the prescription regularity of traditional Chinese medicine compound in the treatment of stable COPD.Data mining of TCM compound in the treatment of stable COPD included 232 prescriptions of COPD in stable period,involving 206 kinds of traditional Chinese medicines.According to the results of frequency analysis,Astragalus propinquus Schischkin,Glycynhiza uralensis Fisch and Poria cocos are the most frequently used single medicine.The types of medicines are mostly medicines for tonifying deficiency,eliminating phlegm,relieving cough and asthma,promoting diuresis and dampness,etc.The medicinal properties are mostly warm,flat and mild.Among the five flavors,sweet,bitter and pungent are more common.Medicines for meridians to meridians of lung and spleen are often selected.A total of 17 common factors were extracted by factor analysis,which mainly reflected the basic therapeutic principles of strengthening body resistance and eliminating pathogenic factors.The treatment methods included in strengthening body resistance mainly include tonifying lung and kidney,invigorating qi and nourishing yin,astringing lung and consolidating exterior,etc.Eliminating pathogenic factors mainly includes eliminating phlegm,eliminating dampness,dredging collaterals,promoting blood circulation,etc.Based on the therapeutic principles,it reflects the basic pathogenesis of COPD in remission period.The analysis of the medicine combination of public factors shows that it is mainly the medicine core composition of classic prescriptions such as Liuwei Dihuang Pills.Erchen Decoction,Sanzi Yangqin Decoction and Qiwei Duqi Pills.The results of cluster analysis showed that Alisma plantago-aquatica Linn-Cortex Moutan,Radix Paeoniae Alba-Cinnamomi,Radix Bupleuri-Ligusticum Chuanxiong,Pericarpium Citri Tangerinae-Pinellia Ternata,Radix Codonopsis-Atractylodis Macrocephalae-Poria-Glycyrrhiza,Radix Rehmanniae Preparata-Comus officinalis-Yam and other medicine pairs/groups could provide reference for clinical3 Based on network pharmacology to explore the mechanism about how modified Qiw eiduqi Pills works on stable COPDUsing network pharmacology technology,99 active components,352 targets and 5762 genes related to COPD were screened in modified Qiwei Duqi Pills.Screening intersection genes found that 253 possible targets were found in modified Qiwei Duqi Pills for the treatment of COPD diseases.Targets located in the center of protein interaction network were MAPK1,MAPK3,TNF,IL6,FOS,JUN,etc.The results of GO function analysis suggest that the functional pathways involved in the related target genes mainly include lipopolysaccharide reaction,reactive oxygen species reaction,nuclear receptor activity,cofactor binding,membrane raft,receptor complex,etc.The results of KEGG pathway enrichment suggest that MAPK,NF-kB,apoptosis,AGE-RAGE,TNF,IL-17,HIF-1,cAMP and other signal pathways may be the targets of modified Qiwei Duqi Pills in COPD.4 Explore the effect and mechanism of modified Qiweiduqi Pills on stable COPD bas ed on mkk3/6-p38 MAPK signaling pathway4.1 General state and pathological results of lung tissue of rats:Aminophylline group and traditional Chinese medicine group can obviously relieve symptoms like cough,shortness of breath and of rats,and the effect of traditional Chinese medicine group is better than that of aminophylline group.Pathological results of lung tissue showed that the alveoli in the blank control group were uniform in size,moderate in thickness and no inflammatory cell infiltration was found.In the model group,the alveolar wall was thickened,the alveoli were uneven,and a large number of inflammatory cells infiltrated into the alveolar wall.Compared with the model group,the pathological changes of lung tissue in aminophylline group and traditional Chinese medicine group were reduced to different degrees.4.2 Expression levels of inflammatory factors IL-1β,TNF-α and IFN-γ:Compared with the blank group,the expression levels of IL-1,TNF and IFN-in alveolar lavage fluid of rate in each group of model group were up-regulated to different degrees.Compared with the model group,the expression levels of IL-lβ,TNF-α and IFN-γ in aminophylline group and traditional Chinese medicine group were significantly down-regulated,and the degree of down-regulation in traditional Chinese medicine group was more obvious.4.3 Expression level of apoptosis molecules and key node proteins of pathways:4.3.1 Bax/Bcl-2:Compared with the blank group,the expression level of Bax protein in hing tissue of model group was significantly up-regulated and the expression level of Bcl-2 protein was significantly down-xegulated.Compared with the model group,the expression level of Bax protein in aminophylline group and traditional Chinese medicine group was significantly down-regulated,and the down-regulated level in traditional Chinese medicine group was more significant than that in aminophylline group.The expression level of Bcl-2 protein was significantly up-regulated,and the up-regulation level of traditional Chinese medicine group was more significant than that of aminophylline group.4.3.2 Fas and FasL:Compared with the blank group,the expression level of Fas and FasL protein in lung tissue of model group was significantly up-regulated;Compared with the model group,the expression levels of Fas and FasL proteins in aminophylline group and traditional Chinese medicine group were significantly down-regulated,and the down-regulation level in traditional Chinese medicine group was more significant than that in aminophylline group.4.3.3 MKK3/p-MKK3:Compared with the blank group,the expression level of MKK3 protein in lung tissue of model group was significantly down-regulated and the expression level of p-MKK3 protein was significantly up-regulated.Compared with the model group,the expression level of MKK3 protein in aminophylline group and traditional Chinese medicine group was significantly up-regulated,and the up-regulation level in traditional Chinese medicine group was more significant than that in aminophylline group.The expression level of p-MKK3 protein was sigmficantly down-regulated,and the down-regulated level in the traditional Chinese medicine group was more significant than that in the aminophylline group.4.3.4 MKK6/p-MKK6:Compared with the blank group,the expression level of MKK6 protein in lung tissue of model group was significantly down-regulated and the expression level of p-MKK6 protein was significantly up-regulated.Compared with the model group,the expression level of MKK6 protein in aminophylline group and traditional Chinese medicine group was significantly up-regulated:and the up-regulation level in traditional Chinese medicine group was more significant than that in aminophylline group.The expression level of p-MKK6 protein was significantly down-regulated,and the down-regulated level in the traditional Chinese medicine group was more significant than that in the aminophylline group.4.3.5 p38 MAPK’p-p38 MAPK:Compared with the blank group,the expression level of p38 MAPK protein in lung tissue of model group was significantly down-regulated and the expression level of p-p38 MAPK protein was significantly up-regulated.Compared with the model group,the expression level of p38 MAPK protein in aminophylline group and traditional Chinese medicine group was significantly up-regulated and the up-regulation level in traditional Chinese medicine group was more significant than that in aminophylline group.The expression level of p-p38 MAPK protein was significantly down-regulated,and the down-regulation level of Chinese medicine group was more significant than that of aminophylline group.4.4 mRNA expression4.4.1 Bax/Bcl-2;Compared with the blank group,the expression level of Bax mRNA in lung tissue of model group was sigmficantly up-regulated and the expression level of Bcl-2 mRNA was significantly down-regulated.Compared with the model group,the expression level of Bax mRNA in aminophylline group and traditional Chinese medicine group was significantly down-regulated,and the down-regulation level in traditional Chinese medicine group was more significant than that in aminophylline group.The expression level of Bcl-2 mRNA was significantly up-regulated,and the up-regulation level of Chinese medicine group was more significant than that of aminophylline group.4.4.2 Fas/FasL:Compared with the blank group,the expression levels of Fas and FasL mRNA in lung tissue of model group were significantly up-regulated Compared with the model group,the expression levels of Fas and FasL mRNA in aminophylline group and traditional Chinese medicine group were significantly down-regulated,and the down-regulation level in traditional Chinese medicine group was more significant than that in aminophylline group.4.4.3 MKK3,MKK6,p38 MAPK:The expression levels of MKK3,MKK6,p38 MAPK mRNA in lung tissue of rats in the model group were similar to those in the blank group,and there was no significant difference.Conclusion:1.The results of systematic evaluation and meta analysis suggest that the total effective rate of traditional Chinese medicine compound combined with conventional western medicine in the treatment of stable COPD is higher than that of conventional western medicine alone,and the relief of lung function index and blood gas analysis index is more significant,which can reduce the score of traditional Chinese medicine symptoms,improve the quality of life,and has good security.2.1he results of data mining research suggest that the disease is located in the lung in stable COPD,and then involves the spleen and kidney,and damages the heart and liver for a long time.Its basic pathogenesis is deficiency of origin and excess,deficiency of origin and excess are mainly qi deficiency,and excess is mainly phlegm and dampness.Strengthening body resistance and eliminating pathogenic factors is the therapeutic principle for COPD in remission period.The therapeutic methods included in strengthening body resistance mainly include tonifying lung and kidney,invigorating qi and nourishing yin,astringing lung and consolidating exterior,etc.Eliminating pathogenic factors mainly includes eliminating phlegm,eliminating dampness,dredging collaterals,promoting blood circulation,etc.Clinical prescriptions are mostly based on Liuwei Dihuang Pills,Qiwei Duqi Pills,Erchen Decoction,etc.The choice of medicines is mostly qi-invigorating medicines,phlegm-resolving medicines,dampness-removing medicines,etc.,and the warm,flat,lung meridian,spleen meridian,sweet,bitter and pungent medicines are more often selected.Cluster analysis and correlation analysis have obtained the common compatibility of some included prescriptions,which can provide reference for future clinical prescriptions.3.According to the results of network pharmacology research,the traditional Chinese medicine compound modified Qiwei Duqi Pills,which has the effects of tonifying lung and kidney,resolving phlegm and dredging collaterals,may treat COPD through multiple targets and ways.The prescription involves 99 active ingredients,352 potential targets and 5762 genes involved in diseases.Results a total of 253 cross genes were screened and obtained.The results of GO enrichment analysis of intersection genes suggest that the genes regulated by modified Qiwei Duqi Pills mainly involve functional pathways such as lipopolysaccharide reaction,reactive oxygen species reaction,nuclear receptor activity,cofactor binding,membrane raft,receptor complex,etc.The results of KEGG enrichment analysis suggest mat MAPK signal pathway,NF-κB signal pathway,apoptosis,AGE-RAGE signal pathway,TNF signal pathway,IL-17 signal pathway,HIF-1 signal pathway,cAMP signal pathway and other signal pathways may play an important role as potential targets in the intervention of modified Qiwei Duqi Pills on COPD.Among them,MAPK signal pathway is closely related to the occurrence and development of COPD,so regulating MAPK signal pathway may be the potential entry point for modified Qiwei Duqi Pills to intervene in COPD.Based on this,in the next study,the mechanism of modified Qiwei Duqi Pills in treating COPD and the internal relationship between MAPK signal pathway and apoptosis are verified through experiments.4.The animal experiment confirmed that modified Qiwei Duqi Pills could play an important role in treating COPD by down-regulating the expression of IL-1β,TNT-α,IFN-γinflammatory factors,down-regulating the phosphorylation levels of MKK3,MKK6,p38 MAPK proteins,inhibiting MKK3/6-p38 MAPK signal pathway,and further regulating endogenous and exogenous apoptosis pathways in the stable phase of COPD.