| Purpose:Through the research on the action mechanism of Bufei Tongluo Decoction on TLR4/My D88/NF-κB signaling pathway in AECOPD model rats,we explored the action mechanism of AECOPD treatment based on the lung deficiency and collaterals stasis theory,which provided a new idea for the prevention and treatment of AECOPD based on the combination of disease and syndrome in TCM.The research methods of network pharmacology were used to explore the mechanism of Bufei Tongluo Decoction in the treatment of AECOPD at the molecular level,as well as other possible chemical components for the treatment of AECOPD,to further explore the action targets of Bufei Tongluo Decoction,and to enrich the theoretical and scientific connotation of the prevention and treatment of AECOPD based on the combination of disease and syndrome in TCM.Material and method:1.The network pharmacology mechanism of Bufei Tongluo Decoction in the treatment of AECOPD1.1 Search TCMSP,HERB database and BATMAN-TCM analysis tool for the active ingredient compounds of Bufei Tongluo Decoction,and obtain relevant targets according to the active ingredients.1.2 Search for disease targets associated with AECOPD in the OMIM database,Gene Cards database.We used Venn diagram to obtain the cross-targets of Bufei Tongluo Decoction and AECOPD disease to obtain the core targets.1.3 The obtained core targets of Bufei Tongluo Decoction were imported into a STRING database to construct a PPI network of core targets.1.4 Cross-targets generated by Bufei Tongluo Decoction and AECOPD were analyzed for GO functional enrichment and KEGG pathway enrichment using Metascape platform.2.Based on the theory of lung deficiency and collaterals stasis,explore the effect of Bufei Tongluo Decoction on TLR4/My D88/NF-κB signaling pathway in AECOPD model rats.2.1 Male Wistar rats were randomly divided into six groups: blank control group,model control group,western medicine control group(roxithromycin),Bufei Tongluo Decoction low dose group,Bufei Tongluo Decoction medium dose group and Bufei Tongluo Decoction high dose group,12 rats in each group;2.2 The AECOPD animal models were constructed by intratracheal instillation of lipopolysaccharide combined with passive smoking.The corresponding drugs were given for intervention on the next day of modeling.Samples were taken and indicators were tested after14 d of drug intervention.2.3 Observe the general state of rats;2.4 Pathological changes of lung tissue in rats are observed by HE staining.2.5 Influence of expression levels of indexes HMGB1,IL-1β,MCP-1 and TNF-α detected by ELISA;2.6 The protein contents of TLR4,My D88 and NF-κB p65 in the lung tissue of rats in each group were detected and analyzed by Western Blot.Results:1.The network pharmacology mechanism of Bufei Tongluo Decoction in the treatment of AECOPD1.1 The active ingredient compounds of Bufei Tongluo Decoction were searched through TCMSP,HERB database and BATMAN-TCM analysis tool,and a total of 177 active ingredient compounds and 605 related targets were obtained.1.2 A total of 1,759 disease targets related to AECOPD were obtained after sorting and merging through the search results in the OMIM database and the Gene Cards database.Using Venn diagram to obtain 207 cross-over targets of Bufei Tongluo Decoction and AECOPD diseases as the core targets;1.3 The collected 207 core targets were imported into the STRING database,and the PPI network of core targets for Bufei Tongluo Decoction and AECOPD was constructed,yielding a total of 199 core targets.Among them,TNF,IL-6,EGFR,IL1-β,VEGFA,and ESR1 were selected as the key nodes of Bufei Tongluo Decoction related to AECOPD according to the degree value.1.4 Core targets of Bufei Tongluo Decoction in the treatment of AECOPD were subjected to GO and KEGG enrichment analysis using Metascape data platform.In GO enrichment analysis,biological processes mainly include reaction to inorganic substances,reaction to hormones,reaction to xenobiotic stimulation,reaction to lipopolysaccharide,reaction to oxygen level,etc.;The cell components mainly included membrane rafts,vesicle cavities,and plasma membrane protein complexes.Molecular functions mainly include oxidoreductase activity,transcription factor binding,protein domain-specific binding,and signal receptor activator activity.Based on KEGG analysis,the core targets of Bufei Tongluo Decoction in the treatment of AECOPD are mainly related to cancer signaling pathway,TNF signaling pathway,IL-17 signaling pathway,PI3K-Akt signaling pathway,NF-κB signaling pathway,etc.2.Based on the theory of lung deficiency and collaterals stasis,explore the effect of Bufei Tongluo Decoction on TLR4/My D88/NF-κB signaling pathway in AECOPD model rats.2.1 Bufei Tongluo Decoction can improve the mental status,food consumption,body weight change,and two-stool status of AECOPD rats.2.2 Bufei Tongluo Decoction can improve the massive inflammatory cell infiltration in the airway as well as the lumen stenosis and cilia shedding in AECOPD rats.2.3 Detection of expression levels of indicators HMGB1,IL-1β,MCP-1 and TNF-α by ELISA: Compared with the rats of the blank control group,the expression levels of HMGB1,IL-1β,MCP-1 and TNF-α in serum of rats in the model group,roxithromycin group,and all dose groups treated with Bufei Tongluo Decoction were significantly increased(P<0.01),and the differences were statistically significant.The expression levels of serum HMGB1,IL-1β,MCP-1,and TNF-α in the western medicine(roxithromycin)group and Chinese medicine(Bufei Tongluo Decoction)low-,medium-and high-dose groups were significantly lower than those in the model group(P< 0.01).The expression levels of serum HMGB1,MCP-1 and TNF-α in the low-dose group of Chinese medicine were higher than those in the western medicine group(P<0.05),while the expression levels of serum HMGB1,IL-1β,MCP-1 and TNF-α in the medium-dose group and high-dose group of Chinese medicine were not significantly different from those in the western medicine group(P>0.05).2.4 The protein contents of TLR4,My D88 and NF-κB p65 in the lung tissue of rats in each group were detected by Western Blot.Compared with the blank control group,the protein expression levels of TLR4,My D88 and NF-κB p65 in the lung tissue of rats in the model group,western medicine roxithromycin group and each dose group of Bufei Tongluo Decoction were increased significantly(P<0.01 or P<0.05),with statistically significant differences.The expression levels of TLR4 and My D88 proteins in lung tissue of western medicine(roxithromycin)group and Chinese medicine(Bufei Tongluo Decoction)medium-and high-dose groups were significantly lower than those in the model group(P<0.01),and the expression levels of these proteins in the low-dose group were lower than that in the model group(P<0.05).The protein expression levels of TLR4 and My D88 in lung tissue of Chinese medicine low dose group were higher than that of western medicine group(P<0.05),while the protein expression levels of TLR4,My D88 and NF-κB p65 in lung tissue of Chinese medicine medium dose group and high dose group were not significantly different from those of western medicine group(P>0.05).Conclusion:1.By using the research method of network pharmacology,we obtained the active components,action targets and action pathways of Bufei Tongluo Decoction in the treatment of AECOPD through preliminary analysis,and further analyzed that Bufei Tongluo Decoction might regulate the inflammatory reaction process of TLR4/My D88/NF-κ B signaling pathway to achieve the purpose of treating AECOPD.2.Bufei Tongluo Decoction can improve the general state of AECOPD model rats and pathological morphological changes in lung tissue,reduce the expression levels of serum HMGB1,IL-1 β,MCP-1,and TNF-α in AECOPD model rats,and reduce the protein expression levels of TLR4,My D88 and NF-κB p65 in lung tissue of AECOPD model rats.Comprehensive analysis showed that the biological process of Bufei Tongluo Decoction in the treatment of AECOPD model rats was to improve the inflammatory response process by regulating the TLR4/My D88/NF-κB signaling pathway. |
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