The Study On The Mechanism Of Xifeng Qinghuo Huoxue Huatan Decoction In Treating Ischemic Stroke Based On Network Pharmacology

Objective:1.Clinical research: To observe the effect of Xifeng Qinghuo Huoxue Huatan decoction on plasma IL-6、IL-8、IL-10 and IFN-γ levels,clinical efficacy,to explore the possibility mechanism of them,and this may pave a new way for therapy.2.Network pharmacology research and molecular docking verification: To explore potential mechanism of xifeng qinghuo huoxue huatan decoction in treating ischemic stroke(IS)based on network pharmacological method to provice new ideas for Compatibility of traditional Chinese medicine and new drug research and development.3.Experimental study on MCAO model animal: Based on the analysis results of network pharmacology,the key targets were screened out and the in vivo experiments were carried out to verify the mechanism of Xifengqinghuo Huoxuetan decoction in MCAO model animals.Method: 1.Clinical research: 60 patients were randomly divided into traditional Chinese medicine(TCM)group and control group.The control group was given standard western medicine treatment,and the TCM group received standard western medicine treatment and Xifeng Qinghuo Huoxue Huatan decoction.After 10 days of treatment,we evaluated the clinical efficacy rate,National Institutes of Health Stroke Scale score(NIHSS),Barthel index(BI)and Modified Rankin scores(m RS)and examine the plasma IL-6,IL-8,IL-10 and IFN-γ levels between the two groups.2.Network pharmacology research and molecular docking verification: Active components of xifeng qinghuo huoxue huatan decoction were screened out through TCMSP,ETCM,TCMID,BATMAN-TCM and literature.Potential targets of the active components were predicted according to the target predictive model of TCMSP,databases of STITCH,Swiss Target and SEA.Ischemic Stroke-related targets were searched filtered out through databases of Genecards,OMIM,Disgenet and Dig See and take the targets-intersection of disease and drugs.STRING database and Cytoscape 3.6.0 software were used to establish the protein-protein interaction network of the core targets associated with this disease.Enrichment analyses of GO and KEGG on the key targets were performed by Metascape.Aout Dock was used to carry out molecular docking between main components and key targets,and Pymol 2.5 was used to visualize the results.3.Experimental study on MCAO model animal: Sixty SD rats were randomly divided into three groups,Blank control group,model group and Chinese medicine group,there were 20 rats in each group.The model group and the Chinese medicine group were made into MCAO models.The Zea Longa5 scoring system was used to evaluate the degree of neurological deficit.Each rat in the blank group and the model group was given 1ml/(100g·d)normal saline,and each rat in the TCM group was given TCM 1ml/(100g·d),morning and evening.On day 7,samples were taken 2 h after gavage.In the model group and the Chinese medicine group,the brain tissue of the lesion side was taken,and The corresponding brain tissues were taken in the blank group.The protein levels of p-AKT1/AKT1,p-P53/P53,p-ERK2/ERK2,p-JNK1/JNK1 and IL-6 were detected by Western blot.Results:1.Clinical research: 10 days after treatment,the total effective rate of TCM group were higher than control group(P<0.05),the neurological functional deficit scale of TCM group were lower than control group(P<0.05),the stroke specific quality of life scale of TCM group were higher than control group(P<0.05),and Scalethe plasma level of IL-8 in TCM group were lower than control group(P<0.05).2.Network pharmacology research and molecular docking verification: There were 251 active components,3971 medicinal targets,1798 targets for Ischemic Stroke,60 key targets and the first signal pathways screened by enrichment analysis.The main components of xifeng qinghuo huoxue huatan decoction in treating Ischemic Stroke were quercetin,kaempferol,beta-sitosterol,luteolin,stigmasterol,etc.The key targets were AKT1,TP53,IL-6,MAPK1,MAPK8,etc.The key biological processes and pathways might include signaling pathways of AGE-RAGE,Fluid shear stress and atherosclerosis,PI3k-Akt,NF-k B,VEGF,etc.The results of molecular docking showed that the binding stability between target protein and ligand was strong.3.Experimental study on MCAO model animal: Western blot results showed that compared with the blank group,the expression of p-Akt1,p-p53,p-JNK1 and p-ERK2 in the model group was increased(P<0.05).Compared with the model group,the expression of p-Akt1,p-p53,p-JNK1,p-ERK2 and IL-6 in the Chinese medicine group was decreased.AKT1 expression was increased(P<0.05).Conclusion:1.Clinical research: Xifeng Qinghuo Huoxue Huatan decoction has an exact effect in treatment of the patients with wind-fire-phlegm-stasis syndrome of acute ischemic stroke,which could improve the alleviate neuronal deficits quality of life by decreasing plasma level of IL-8.2.Network pharmacology research and molecular docking verification: This study reveals that Xifeng Qinghuo Huoxue Huatan decoction may play a role in the treatment of ischemic stroke by acting on the AGE-RAGE,PI3k-Akt,NF-k B and other signaling pathways through AKT1,TP53,IL-6 and other targets to participate in inflammatory response,immune response,nerve cell apoptosis and other processes.3.Experimental study on MCAO model animal: The effect of this prescription on the expression of target proteins AKT1,TP53,IL-6,MAPK1 and MAPK8 was preliminarily confirmed.It provides the basis for the clinical development and utilization of Xifeng Qinghuo Huoxue Huatan decoction in the treatment of ischemic stroke.