Mechanistic Research Of Yiqi Huoxue Tuodu Recipe In Trating CP/CPPS By Mediating NF-κB Pathway

CP/CPPS is a common disease in the male genitourinary system,characterized by high incidence rate and high recurrence rate.Due to the complex pathogenesis of CP/CPPS,the current study is inconclusive and there is a lack of effective treatment options for this disease in clinical practice.In this study,CP/CPPS was used as the research goal to analyse the clinical data of patients with CP/CPPS treated with Yiqi Huoxue Tuodu Recipe,to virtually construct the possible pathogenesis network of Yiqi Huoxue Tuodu Recipe in the treatment of CP/CPPS by combing with modern scientific cutting-edge technology,and to verify the predicted molecular pathway model by animal experiments finally.The aim of this study is to preliminarily explore the possible mechanism of Yiqi Huoxue Tuodu Recipe in the treatment of CP/CPPS,and to provide theoretical and experimental basis for the clinical application and promotion of Yiqi Huoxue Tuodu Recipe in the treatment of CP/CPPS.This study mainly includes four parts:literature review,clinical study analysis,network pharmacology research and experimental basic research.1.Literature reviewThis study concludes the research progress of diagnosis,pathogenesis and treatment of CP/CPPS in Chinese and Western medicine,and knows about the current research progress and shortcomings of CP/CPPS systematically.Based on the current research on the pathogenesis of CP/CPPS,the fact that inflammation and immunity play a key role in the pathogenesis of CP/CPPS is well known.The relevant inflammation,immune factors and pathway reports involved in the literature are also systematically collated in order to know about the possible pathogenesis of inflammation and immune factors on CP/CPPS.2.Clinical study:Clinical observation data analysis of Yiqi Huoxue Todu Recipe in improving patients with CP/CPPS and prescription optimizationObjective:This study analysis is to explore the association between inflammatory factors and CP/CPPS by analyzing the clinical observation results of Yiqi Huoxue Tuodu Recipe in improving NIH-CPSI score in patients with CP/CPPS.Methods:The results of TNF-α and IL-10 in the expressed prostatic secretion(EPS)of 120 patients with CP/CPPS,correlation with patient symptoms,and changes before and after treatment were analyzed.Results:EPS cytokines TNF-α and IL-10 were also measured in the two groups before and after treatment.It was found that cytokine TNFa in the two groups after treatment was significantly lower than that before treatment,the decrease range in the treatment group was significantly higher than that in the control group,IL-10 in the two groups after treatment was significantly lower than that before treatment,and the decrease range in the treatment group was significantly higher than that in the control group.The results of general linear analysis showed that TNF-αcontent in EPS was positively correlated with CPSI-pain score(r=0.37,P<0.05).TNFa and TCM symptom score were positively correlated(r=0.30,P<0.05),but not with NIH-CPSI total score,CPSI-life quality score and CPSI-urination score(P>0.05);the results of general linear analysis showed that IL-10 content in EPS was linearly correlated with pain score(r=0.38,P<0.01),but not significantly linearly correlated with NIH-CPSI total score,urination score,life quality score and TCM symptom score(P>0.05).Conclusion:Yiqi Huoxue Tuodu Recipe may play a role in the treatment of CP/CPPS by decreasing the expression of cytokines and inhibiting inflammation and immune response.3.Network pharmacology research:Based on network pharmacology research methods to explore the mechanism of Yiqi Huoxue Tuodu Recipe in the treatment of CP/CPPS.Objective:This study is to study the mechanism of action of Yiqi Huoxue Tuodu Recipe in the treatment of CP/CPPS by using network pharmacology and systems biology research methods,and to explore the key molecules and pathway models of its therapeutic effect.Methods:The effective chemical constituents of Yiqi Huoxue Tuodu Recipe were selected from the pharmacology database of traditional Chinese medicine system(TCMSP)and published literatures,and the target of chemical constituents was predicted by TCMSP and SwissTargetPrediction database.OMIM,GeneCards,and DisGeNET databases were applied to screen CP/CPPS disease-related targets,followed by taking the intersection targets of Yiqi Huoxue Tuodu Recipe and CP/CPPS-related genes to construct a protein-protein interaction(PPI)network by using STRING.Protein interaction networks were constructed by using Cytoscape and GO analysis and KEGG signaling pathway enrichment analysis is performed by utilizing STRING and DAVID databases.Results:145 active components of 7 herbs in Yiqi Huoxue Tuodu Recipe were selected.961 potential targets were predicted.2871 CP/CPPS diseaserelated genes,420 drug-disease intersection gene targets and 380 related signaling pathways were obtained.Conclusion:Yiqi Huoxue Tuodu Recipe has the characteristics of multiple targets and multiple pathways in the treatment of CP/CPPS.After screening and analysis,it is predicted that it may mainly play a therapeutic role through NF-κB pathway,JAK-STAT signaling pathway,p53 signaling pathway,Inflammatory mediator regulation of TRP channels etc.to inhibit gene transcription,prevent apoptosis,decrease inflammation and reduce immune response.4.Experimental study:The mechanism of Yiqi Huoxue Tuodu Recipe in regulating inflammation and immunotherapy of CP/CPPS by mediating NF-κB pathwayObjective:To observe the expression of inflammatory factors,immune molecules and NF-κB pathway-related proteins in different groups and to explore the possible mechanism of Yiqi Huoxue Tuodu Recipe in treating CP/CPPS by constructing EAP model in mice and intervening with Yiqi Huoxue Tuodu Recipe.Methods:Sixty C57BL/6 male mice over 8 weeks old were randomly divided into control group,high dose Herbal group,low dose Herbal group,western medicine group and model group,each group with 10 mice.Except for the control group,the remaining 50 mice were given multipoint subcutaneous injection of prostate protein purified solution+complete Freund’s complete adjuvant at day 0,day 15 and day30 respectively and EAP model was constructed at day 45.After that,high dose Herbal group and low dose Herbal group were intragastrically administered with different doses of Yiqi Huoxue Tuodu Decoction,the western medicine group was intragastrically administered with tansoprazole aqueous solvent,and the model group and blank group were intragastrically administered with purified water.28 days later,the serum of mice was collected,and the prostate tissue of mice was isolated.HE staining was used to observe the pathomorphological changes and inflammatory scores of prostate tissues under a light microscope.TNF-α,IL-1β,IgA,IgG,and IgM in serum were detected by ELISA method,and the protein expression of TNF-α,IL-1β,pIKK-β,and pNF-κBP65 in prostate tissues was detected by Western blot.Results:Yiqi Huoxue Tuodu Recipe could improve improve the pathological structure of prostate tissue according to that the inflammation score of high dose Herbal group was superior to that of low dose Herbal group(P<0.05);Yiqi Huoxue Tuodu Recipe could reduce the expression of TNF-αand IL-1β in serum since high dose(Chinese traditional medicine)group was superior to that of low dose(Chinese traditional medicine)group(P<0.05)on reducing the expression of TNF-α and could decrease the levels of IgA,IgG and IgM in serum since high dose(Chinese traditional medicine)group was superior to that of low dose(Chinese traditional medicine)group(P<0.05)on decreasing the expression of IgA and IgM.Yiqi Huoxue Tuodu Recipe could inhibit the expression of TNF-α,IL-1β,pIKK-β,and pNF-κBP65 in NF-κB pathway-related proteins,phosphorylation and pathway activation.Conclusion:The mouse model was established by multipoint subcutaneous injection of prostate protein purified solution+complete Freund’s adjuvant,showing the characteristics of chronic prostatitis.Yiqi Huoxue Tuodu Recipe can improve the histopathological morphology of prostate,reduce the content of serum inflammatory factors and regulate the immune level.Its mechanism is related to inhibiting the expression of TNF-α,IL-1β,pIKK-β,pNF-κBP65 protein,inhibiting phosphorylation and inhibiting pathway activation.