Discussion On The Mechanism Of Xinfeng Capsule In The Treatment Of Primary Sjogren’s Syndrome Based On Network Pharmacology

Objective Based on network pharmacology and molecular docking,explore the molecular mechanism of Xinfeng capsule(XFC)in the treatment of primary Sjogren’s syndrome(p SS)and carry out experimental verification in vivo,so as to provide theoretical basis for XFC treatment of p SS and scientific experimental basis for clinical treatment.Methods1.Firstly,the effective ingredients of "Astragalus membranaceus","Coix lacryma-jobi" and "Tripterygium wilfordii" in XFC were obtained from the system pharmacology database and analysis platform of traditional Chinese medicine(TCMSP);The active ingredient of “centipede”,which cannot be found in the TCMSP database,is searched on the HERB Herbal Pharmacopoeia and CNKI platform,and its active ingredient is obtained through Pub-chem database.According to the optimal toxicokinetics ADME,drug class properties and oral bioavailability,the effective components and corresponding targets of XFC were screened.Then,use Gene Card and OMIM database to retrieve the prediction targets related to p SS.2.Import the screened XFC active ingredients and their corresponding target data into the Cytascape software,build the XFC component-target network diagram,and screen the core components.The Venny online tool was used to screen the common targets of XFC active ingredients and p SS,and the obtained common targets were input into the String database to obtain the target protein interaction network(PPI).The core therapeutic targets were selected by the topological attribute analysis of the PPI network with Cytoscape.3.Carry out GO function annotation and KEGG enrichment analysis on the relevant pathway of the core treatment target through Matscape software,and make visual presentation.Finally,the key targets and core components are docked with Autodock Tools software to test their binding ability.4.Animal experiment verification.First,the primary Sjogren’s syndrome mouse model was constructed and C57BL/6 mice were randomly divided into normal group,model group,XFC low-dose group(1.2 g/kg)and XFC high-dose group(4.8 g/kg).The model mice of Sjogren’s syndrome were induced by SG protein antigen and treated with low and high doses of Xinfeng capsule by gavage.The changes of body weight,saliva volume and pathological changes of submandibular gland tissue were observed.The levels of IL-6 and NKCC1 protein in submandibular gland tissue were measured by Western blot,and the level of IL-6 m RNA was detected by real-time quantitative PCR.Result1.After screening according to conditions,XFC included 60 active ingredients,598 related targets and 150 intersections with p SS targets.Through component-target network topology attribute analysis,the top three effective components with degree values are Quercetin(MOL000098),kaempferol(MOL000422),isorhamnetin(MOL000354).The three targets with higher network degree in PPI network are IL-6,TNF and IL-1 β.2.The results of GO enrichment analysis showed that IL-6 was mainly enriched in BP,including the reaction to bacterial molecules,leukocyte differentiation,positive regulation of protein phosphorylation,positive regulation of movement and positive regulation of cell apoptosis.Based on the results of KEGG enrichment,IL-6 is involved in cancer pathway,apoptosis and NF-kappa B signal pathway.3.Effect of XFC on body weight,saliva secretion,pathological changes of submandibular gland and expression of IL-6 and NKCC1 protein in submandibular gland of SS model mice.3.1 With the increase of age,the weight of mice in each group increased,and the weight of mice in the model group was gradually lower than that in the normal group.Compared with the model group,XFC could significantly increase the body weight of SS mice(P<0.01).3.2 The saliva volume of normal group mice gradually increased,and the model group mice were significantly lower than the normal group.Compared with the model group,XFC could significantly improve the saliva volume of SS mice(P<0.01).3.3 Compared with the normal control group,the infiltration of lymphocytes in the submandibular gland tissue of mice in the model group increased,the cells were necrotic,and the lesions were obvious.XFC can significantly recover the pathological changes.3.4 Compared with the blank control group,the expression level of IL-6 and NKCC1 protein in the model group decreased.Compared with the model group,XFC can increase the protein level of IL-6 and NKCC1.3.5 Compared with the blank control group,the level of IL-6 m RNA in the model group decreased,and compared with the model group,the level of IL-6 m RNA in the XFC highdose group increased significantly(P<0.05).ConclusionAs a compound preparation of traditional Chinese medicine,XFC has the characteristics of multi-target,multi-component and multi-system.This study explored the active components,action targets and signal pathways of XFC in the treatment of p SS through the method of network pharmacology.A total of 60 active components of XFC and 1497 p SS-related targets were screened.It was identified that IL-6 was the key target gene for XFC treatment of p SS,which was mainly enriched in cell apoptosis and NF-kappa B signal pathway.At the same time,after the construction of p SS model mice and XFC treatment,the salivary volume and submandibular gland tissue pathology changed.Western blot and RT-PCR results showed that the level of IL-6 protein and m RNA in the model group decreased,and increased to varying degrees after XFC treatment.