| This paper is divided into two parts:The first part is a literature review,introducing the epidemiological investigation,pathological mechanism research of PM2.5 leading to respiratory diseases and the research progress of Chinese medicine on lung diseases caused by PM2.5 in recent years;reviewing Yupingfengsan and allergy decoction Clinical trials and experimental studies.The second part is experimental research.Objective:1.To compare the mechanism of Yupingfeng Powder and Jiawei Allergy Decoction in inhibiting inflammatory reaction and regulating immune balance to interfere with lung injury induced by PM2.5;2.To investigate lung injury induced by PM2.5 by transcriptomics and proteomics analysis The possible mechanism,as well as the target of Yupingfengsan and flavored allergy.METHODS:Forty male SPF male Wistar rats were randomly divided into normal control group,PM2.5 model group,Jiawei allergy frying group and Yupingfeng powder group.The normal group was given daily saline lml/100g·bw,and the other 3 groups were instilled with PM2.5 suspension 65?g/100?l every 3 days.The PM2.5 model group was given lml/100g·bw of distilled water per day.Stomach,addicted allergy frying group daily add flavor allergy frying lml/100g·bw gavage,Yupingfeng San group daily Yupingfeng loose lml/100g·bw gavage,a total of 28 days.On the 29th day of the experiment,peripheral blood,alveolar lavage fluid,and lung tissue were collected.The pathological changes of bronchi and alveolar in each group were observed by HE staining.The goblet cell metaplasia of airway epithelium in each group was observed by AB-PAS staining.MPO,CD68,CD4,CD8,ICAM-1,MCPwere observed by immunohistochemical staining.1.Expression of MUC5AC,NE,RORyt and FoxP3,ELISA was used to detect the levels of IL-1?,TNF-?,IL-6,IL-10,TGF-?,IL-17A in peripheral blood of each group,alveolar lavage The contents of IL-1?,TNF-?,IL-6,IL-10,TGF-?,IL-17A,IL-4,IL-5,IL-13 and IgE in liquid were detected by real-time fluorescent quantitative PCR.Expression of IL-4,IL-5,IL-13,IgE,Fc?RI,NE,MUC5AC,IL-17,IL-10,RORyt and FoxP3 mRNA in rat lung tissue,TLR2 and MyD88 in lung tissue were detected by Western Blot.Expression of NF?B,p65,p-JNK2,p-ERK1/2,MCP-1,MUC5AC,NE,EGFR,p-EGFR,PI3K,p-PI3K,AKT,p-AKT,ROR?t,FoxP3 protein.The lung tissues of 3 rats in each group were selected,and Agilent expression profiling chip technology was used to detect the whole genome expression and TMT marker quantitative proteomics analysis.The data were analyzed by cluster analysis,GO function analysis and KEGG signal pathway analysis.The analytical method analyzes the data results.Result:The results of HE staining showed that compared with the normal control group,the airway and alveolar structure of the lung tissue of the PM2.5 model group were destroyed,and a large number of inflammatory cell infiltration and glandular hyperplasia were observed,accompanied by cilia adhesion,shedding,lodging;Compared with the Jiaxingfengsan group,the airway epithelial cilia adhesion,lodging and shedding of the rats in the model group were significantly improved,the alveolar collapse fusion was reduced,the inflammatory cell infiltration and the lung tissue congestion were reduced.The airway epithelial structure of Yupingfengsan group was more complete,and the airway smooth muscle thickness of the additive allergy frying group was less than that of PM2.5 model group and Yupingfeng powder group(P<0.05).The results of AB-PAS staining showed that the number of goblet cells in the airway epithelium of the PM2.5 model group was significantly higher than that of the normal group.Compared with the PM2.5 model group,the goblet cells in the flavored allergy decoction group and the Yupingfeng powder group were significantly different.The decrease was(P<0.01,P<0.01),and the inhibition effect of Yupingfengsan on goblet cell metaplasia was significantly better than that of Jiawei allergy decoction group(P<0.01).Immunohistochemical staining showed that the positive staining area and average optical density of MPO,CD68,CD4,CD8,ICAM-1,MCP-1,MUC5AC,NE,RORyt in the PM2.5 model group were significantly higher than those in the normal control group,FoxP3.Positive staining cells were significantly reduced compared with normal controls;MPO,CD68,CD4,CD8,ICAM-1,MCP-1,MUC5AC,NE,RORyt positive staining areas in Jiawei Jianyan group and Yupingfengsan group were more significant than PM2.5 model group.Significantly reduced,the number of positive staining cells of FoxP3 was significantly increased,and the average optical density of the positive staining area of MUC5AC and NE in Yupingfengsan group was significantly lower than that in the flavored allergy group.The results of ELISA showed that IL-6 was significantly increased in the PM2.5 model group compared with the normal control group,and IL-1?,IgE,and IL-17A were significantly increased in BALF.Compared with the PM2.5 model group,the additive was allergic.The contents of IgE and IL-4 in the BALF group were significantly lower than those in the PM2.5 model group.Compared with the PM2.5 model group,IL-1?,IgE and IL-4 were significantly decreased in the BALF group.Real-time quantitative PCR showed that compared with the normal control group,the expression of Fc?RI,IL-13,MUC5AC and RORyt mRNA in lung tissue of PM2.5 model group was significantly increased,IL-10 mRNA expression was significantly decreased;and PM2.5 model Compared with the group,the expression of Fc?RI mRNA in lung tissue of the Jiaweifengjian group was significantly decreased,and the expression of IL-10 mRNA was significantly increased.Compared with the PM2.5 model group,the expression of Fc?RI and MUC5AC mRNA in the lung tissue of Yupingfengsan group decreased significantly,IL-10 mRNA expression was significantly elevated.Western Blot results showed that PM2.5 significantly up-regulated the levels of MCP-1,pERK1/2,MUC5AC,NE,p-PI3K/PI3K,p-AKT/AKT,and ROR?t in lung tissue,inhibiting pJNK2 And the expression of FoxP3;the expression of MUC5AC,NE,MCP-1 was significantly decreased compared with PM2.5 model group,the level of AKT phosphorylation was decreased,the expression of FoxP3 was significantly increased;Yupingfengsan group and PM2.5 Compared with MUC5AC,NE,and MCP-1,the expression of the model group was significantly decreased,the phosphorylation levels of PI3K and AKT were decreased,and the expression of FoxP3 was significantly increased.Transcriptomic studies showed that the PM2.5 model group compared with the normal control group(PM2.5 vs Control),a total of 431 differential genes,of which 322 up-regulated genes,109 down-regulated genes;flavored allergy frying group and PM2.5 Compared with the model group(GMJ vs PM2.5),there were 184 differential genes,including 49 up-regulated genes and 135 down-regulated genes;Yupingfengsan group compared with PM2.5 model group(YPF vs PM2.5),the difference There are 767 genes,of which 421 are up-regulated and 346 are down-regulated.The genes with differential expression in PM2.5 vs Control and GMJ vs PM2.5 were extracted.It was found that PM2.5 significantly increased the expression of Aloxl2e,Brs3 and T1r12 mRNA in rat lung tissue.The expression level of mRNA in this part of the gene was significantly decreased.The expression of Cldn19,Cry1,Fos and Slc26a3 mRNA in the PM2.5 model group was significantly lower than that in the normal control group.The mRNA expression level of this part of the gene was significantly increased after the intervention of allergy.Genes differentially expressed in PM2.5 vs Control and YPF vs PM2.5 were extracted.It was found that PM2.5 significantly increased Rph3a,Gabrg2,Olr1413,Adad2,B3gnt6,Olr132,Gbpl,Vwa5b2 in rat lung tissue.,Rhox2,Kptn,Diol,Epgn,Cobl,Vomlr57,Rspo2,Gal3st4,Stonl,Slc37a3,Hoxc8,RT1-M4,Kiaa1671,Cldnd2,Fam163b mRNA expression,the expression level of this part of the mRNA after intervention by Yupingfengsan Significantly decreased;the expression of Abcal,Asb5,Rasd1,Upk3a,Slc26a3,Pdgfd,Gas2,Cryl mRNA was significantly lower in the PM2.5 model group than in the normal control group,and the mRNA expression level of this part of the gene was significantly increased after Yupingfengsan intervention.The GO enrichment analysis found that it was not only related to the differentially expressed genes in the PM2.5 model group vs.normal control group,but also related to the differentially expressed genes in the flavored allergy decoction group vs PM2.5 model group including:SMAD protein signal transduction And growth factor activities.It is not only related to the differentially expressed genes in the PM2.5 model group vs.normal control group,but also related to the differentially expressed genes in the Yupingfengsan group vs PM2.5 model group including:phospholipase C activation,G protein coupled receptor Signal pathway,regulation of circadian rhythm,positive regulation of myoblast differentiation,extracellular domain,synaptic vesicle membrane,selenium binding,growth factor activity,sputum binding,receptor binding function.The KEGG signaling pathway analysis revealed that it was a pathway related to the differentially expressed genes in the PM2.5 model group vs.the normal control group,and was also associated with the differentially expressed genes in the flavored allergy frying group vs PM2.5 model group including:arachidonic acid metabolism,cytochrome P450 metabolism of xenobiotics and drug metabolism-cytochrome P450 pathway.It is a pathway related to the differentially expressed genes in the PM2.5 model group vs.normal control group,and is also related to the differentially expressed genes in the Yupingfengsan group vs PM2.5 model group including:neuroactive ligand-receptor interaction,drug Metabolismcytochrome P450 and circadian rhythm pathway.Proteomics studies showed that the PM2.5 model group compared with the normal control group(PM2.5 vs Control),a total of 73 differential proteins,including 50 up-regulated proteins and 23 down-regulated proteins;flavored allergy frying group and PM2.5 Compared with the model group(GMJ vs PM2.5),there were 77 differential proteins,including 34 upregulated proteins and 43 down-regulated proteins;Yupingfengsan group compared with PM2.5 model group(YPF vs PM2.5),the difference There are 100 proteins,of which 53 are up-regulated and 47 are down-regulated.Extracting proteins with differential expression in PM2.5 vs Control and GMJ vs PM2.5 found that PM2.5 significantly increased Shmt2,Hist3h2bb,Histlh2bh,Ednra,Arl5a,Dynclil,Slc4a2,and Col14al in rat lung tissue.The expression of Hspbpl,Histlh1e,Ctla2a,Lgals5,Lgals9,Fabp4,Ca3 and Serpina3c protein was significantly decreased after the intervention of Jiawei Allergy Decoction.PM2.5 exposure can significantly down-regulate the levels of Nudt2 and Erp29 in lung tissue.Adding allergies can significantly increase the content of Nudt2 and Erp29 in lung tissue.Extracting proteins with differential expression in PM2.5 vs Control and YPF vs PM2.5 found that PM2.5 significantly increased Serpina3c,Arl5a,Aspscrl,P2rx7,Histlhle,Hspbpl,Ctcf,Timp2 in rat lung tissue.The expression of Slc4a2,Ctla2a,Ednra,Histlh2bh,Mzb1,Shmt2,Dyncli1 protein was significantly decreased after the intervention of Yupingfengsan;PM2.5 exposure could significantly down-regulate Krt10,Defb4,Aqp4 in lung tissue.Protein level,Yupingfeng powder can significantly increase the content of this part of the protein in the lung tissue.The GO enrichment analysis found that it was not only related to the differentially expressed protein in the PM2.5 model group vs.the normal control group,but also related to the differentially expressed proteins in the flavored allergy frying group vs PM2.5 model group including:defense response,stress Functions such as response,innate immune response,cell adhesion,regulation of cellular protein metabolism,and response to cytokines.It is not only related to the differentially expressed protein in the PM2.5 model group vs.normal control group,but also related to the differentially expressed proteins in the Yupingfengsan group vs.PM2.5 model group including:immune response,stress response,hydrolase activity Negative regulation,innate immunity,proteolysis and other functions.Conclusion:1.Yupingfeng Powder and Jiawei Allergy Decoction can improve the inflammation and damage of lung tissue caused by PM2.5 by inhibiting the expression of ICAM-1 and MCP-1 in airway epithelial cells;2.Yupingfengsan and flavored allergy frying Down-regulation of Fc?RI gene expression and IgE levels in lung tissue,reducing eosinophilic airway inflammation;3.Yupingfengsan and flavored allergy decoction relieves high secretion of airway mucus caused by PM2.5 by inhibiting EGFR-PI3K-AKT signaling pathway,The intervention effect of Yupingfeng Powder is better than that of Jiawei Allergy Decoction;4.Yupingfeng Powder and Jiawei Allergy Decoction can alleviate the inflammation of lung tissue caused by PM2.5 by regulating the balance of Th17/Treg;5.Transcriptomics and proteomics research It is indicated that Jiawei Allergy Decoction can inhibit the expression of inflammatory genes Aloxl2e and TLR12 in lung tissue of PM2.5 model rats,inhibit the oncoprotein,down-regulate histone and galectin,and up-regulate the oncoprotein;Yupingfengsan can inhibit G protein coupling.Receptor gene expression,upregulation of tumor suppressor genes Rasdl,Gas2,inhibition of oncogenic proteins,downregulation of histones,up-regulation of aquaporins,suggesting that these molecules are potential targets for traditional Chinese medicine compounds,for further in-depth The study provides a reference;6.There is a significant difference between the transcriptomics and proteomics results of the comparison between groups,which may be related to the regulation of mRNA translation and the interaction between proteins by non-coding RNAs. |
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