Experimental Study Of Yangyin Qingfei Decoction On Pulmonary Epithelial-mesenchymal Transition In SMPP Mice

Purpose:Using network pharmacology methods to study the mechanism of Yangyin Qingfei Decoction Modified in the treatment of severe Mycoplasma pneumonia(SMPP),use Yangyin Qingfei Decoction Modified to intervene in the treatment of severe Mycoplasma pneumoniae pneumonia in mice,and to explore the effect of Yangyin Qingfei Decoction Modified on high load The effect of pulmonary epithelial-mesenchymal transition and related inflammation pathway NF-κB/IL-6 gene expression in the BALB/c mouse model of severe Mycoplasma pneumoniae pneumonia induced by MP nasal drip,to further clarify the intervention of Yangyin Qingfei Decoction Modified in the treatment of severe pneumonia The mechanism and targets of mycoplasma pneumonia.Material and method:Collect the compounds and effective targets of 10 Chinese medicines of Yangyin Qingfei Decoction Modified with TCMSP database,TCMID database and Swiss Target Prediction,screen the prediction targets of severe Mycoplasma pneumoniae pneumonia with the help of Gene Cards database,and perform Venny analysis with the active ingredient targets of Chinese medicine Determine the targeting relationship of the intersection targets;construct a prescription-compound component-key target network diagram through Cytoscape 3.7.2 software;use the String database to construct a key target protein interaction(PPI)network diagram;finally use the Metascape database and the DAVID database to perform GO analysis of gene function and enrichment analysis of KEGG pathway.In the next experiment,120 BALB/c mice were selected and randomly divided into normal group,model group,azithromycin group,traditional Chinese medicine group,integrated traditional Chinese and western medicine group,24 mice in each group,except for the normal group,all were instilled 0.1m L high-load 1010 CCU./m L MP bacterial liquid,establish SMPP model.Each group of mice were randomly killed on the 3rd,7th,10 th,and 14 th days after MP infection for eyeball blood and specimen collection.Observe the general state of the mice and measure the changes in their body temperature.The lung pathology of mice was observed by HE staining method,the expression of interleukin-6(IL-6)in serum was detected by ELISA method;the expression of NF-κB protein in mouse lung tissue was detected by Western blotting;the lung tissue was detected by q PCR method.E-cadherin(E-cad)and nerve cadherin(N-cad)gene content were determined.Results:1.Screened 179 active compounds,786 drug targets,277 disease targets,and 75 intersection targets.The PPI network involves 75 protein nodes.The key targets include IL6,GAPDH,VEGFA,TP53,TNF,JUN,CXCL8,PTGS2,IL10,STAT3,EGFR,TLR4,MYC,MMP9,CASP3,MAPK1,IL1 B,FN1,CRPHMOX1;GO enrichment function analysis is mainly related to cell regulation,cell movement regulation,angiogenesis and development,cytokines,growth factor receptor binding,organelles,cell membranes,etc.;30 KEGG pathways,Yangyin Qingfei Decoction Modified,anti-SMPP Closely related pathways include NF-κB signaling pathway,Toll-like receptor signaling pathway,TNF signaling pathway,MAPK signaling pathway,PI3K-Akt signaling pathway,NOD-like receptor signaling pathway,HIF-1 signaling pathway,etc.2.The pathological results showed that the wall of bronchioles in the model group was thickened,the alveolar structure was severely damaged,the tracheal epithelium fell off,the alveolar interval was significantly widened,a large number of inflammatory cells exuded,the lung tissue capillary congestion,the alveolar,alveolar duct and other structures disappeared;Azithromycin Compared with the model group,the number of inflammatory cells in the mouse group was significantly improved,and a small amount of inflammatory cells infiltrated;there were only a few inflammatory cells in the alveoli of the mice in the traditional Chinese medicine group,and the lung tissue was moderately congested and edema.There are a small amount of inflammatory cells,the alveolar compartment is thickened,and the congestion is lighter than that of the model control group.After infection,the pathological score of the mice was higher than that of the normal group,and reached the peak value on the 7th day after infection;the body temperature of the mice was changed on the 7th day compared with the normal control group.Body temperature reached its peak.Compared with the model group,each administration group started to decrease on the 7th day(P<0.05).3.After high-load MP infection,the expression of IL-6 in serum was detected by ELISA.Compared with the normal group,the levels of IL-6 in the serum of mice were increased at each time point(P<0.05)and reached the peak on the 7th day;compared with the model group,the levels of IL-6 in the serum of the mice in the three administration groups were equal Some decrease(P<0.05);but the level of reduction in the traditional Chinese medicine group is not obvious,and the level of reduction in the azithromycin group and the integrated traditional Chinese and western medicine group is more obvious,indicating that the three administration groups can inhibit the expression of IL-6 in the serum of mice;Today,the curative effect of the integrated traditional Chinese and western medicine group is better than that of the azithromycin group and the traditional Chinese medicine group(P<0.05).On the10 th and 14 th day of administration,there is no significant difference between the integrated traditional Chinese and western medicine group and the azithromycin group(P>0.05),but compared with the traditional Chinese medicine group both were better than the traditional Chinese medicine group(P<0.05),indicating that the effect of Yangyin Qingfei Decoction Modified combined with azithromycin is better.4.Western blotting to detect the expression of NF-κB protein in mouse lung tissue: After MP infection,compared with the normal group,NF-κB in the serum of mice increased at each time point(P<0.05),and reached the peak on the 3rd day,And then gradually decreased;after drug intervention,compared with the model group,it started to take effect on the 7th day.The level of NF-κB in the serum of mice in the three-dose group decreased(P<0.05),at the10 th and 14 th days The integrated traditional Chinese and western medicine group is better than the azithromycin group and the traditional Chinese medicine group(P<0.05).5.The q PCR method to detect the expression of E-cad and N-cad m RNA in the lung tissues of mice showed that: After MP infection,the expression of E-cad m RNA in mice decreased at each time point,and the difference was statistically significant compared with the normal control group(P<0.05).After drug intervention,compared with the model group,on the 3rd and 7th day,each administration group had no obvious effect(P>0.05);on the 10 th day,each administration group started to take effect,but there was no difference between the three administration groups(P>0.05),but on the 14 th day,the therapeutic effect of the integrated traditional Chinese and western medicine group was better than that of the traditional Chinese medicine group and the azithromycin group(P<0.05);after MP infection,the expression of N-cad m RNA in mice increased at each time point,and was compared with normal The difference in the control group was statistically significant(P<0.05).After drug intervention,compared with the model group,the effect of each administration group was not obvious on the 3rd day(P>0.05),and the integrated traditional Chinese and western medicine group started to take effect on the 7th day,but the azithromycin group and the Chinese medicine group did not take effect;on the 10 th day All treatment groups were effective,but there was no significant difference between the three treatment groups(P>0.05);on the 14 th day,the treatment effect of the integrated traditional Chinese and western medicine group was better than that of the traditional Chinese medicine group and the azithromycin group(P<0.05).Conclusion:1.The quercetin,wogonin,aloe-emodin and other ingredients in Yangyin Qingfei Decoction Modified are likely to act on the NF-κB signaling pathway and Toll-like through targets such as IL6,TNF,IL10,TLR4,MAPK1,IL1 B,and CRP.Receptor signaling pathway,TNF signaling pathway,MAPK signaling pathway and many other signal transduction pathways play an anti-inflammatory effect.2.Yangyin Qingfei Decoction Modified can reduce the pulmonary fibrosis process of severe Mycoplasma pneumoniae pneumonia by intervening in pulmonary epithelial-mesenchymal transition,and may be a target for the treatment of SMPP.3.Yangyin Qingfei Decoction Modified can effectively reduce lung inflammation in SMPP mice,which may be related to the inhibition of NF-κB/IL-6 inflammation signaling pathway,and the combined use of azithromycin is more effective.