Study On Active Substances And Mechanism Of Naoan Capsule In The Treatment Of Ischemic Stroke

Stroke is one of the three major death diseases in the world nowadays.Ischemic stroke(IS)as most types of stroke is a multi-factor complex cardiovascular and cerebrovascular diseases.Traditional Chinese medic ine(TCM)pays attention to regulate physiological functions through the whole body especially complex diseases like stroke,which have certain therapeutic advantages.AS a chinese patent medic ine,NAC is widely used in clinical treatment of stroke.Studying the active substances and mechanism of NAC in the treatment of IS could improve it's formulations,processes and dosage forms to further improve clinical efficacy.At the same time,there is a systematic and clear analys is of the pathogenes is and possible pathology of the IS.Therefore,in this research,the possible pharmacodynamic substances and mechanisms of NAC in the treatment of IS were studied.In this study,constituents migrating to blood of NAC were collected according to oral bioavailability(OB)and drug-likeness(DL),while IS-related targets were collected from the CTD,TTD,Drugbak database.Using the three current mainstream research methods of network pharmacology,namely basing on intersection genes,intersection pathways,protein-protein interactions(PPIs)network pattern recognition,the main mechanism of NAC treatment for IS was preliminarily analyzed.The differential expression genes(DEGs)were screened by searching the IS gene expression profiles.c Map(Connectivity map)analys is and molecular docking were used to obtain the possible active substances with gene enrichment pathways obtained by Metascape.At the same time,the linked amino acid residues were docked through Sybyl 2.0 software before machine learning classification models were constructed to classify inhibitors and non-inhibitors that may have key targets in NAC compounds.Finally,a middle cerebral artery occlusion(MCAO)model experiment was carried out in rats to determine the overall pesticide effect of NAC and the changes of key proteins content.As a result,54 IS-related targets,51 NAC compounds corresponding to 325 potential targets were collected from the database.Among them,there were 16 intersection targets,13 closely interacting targets,and 13 corresponding association compounds.Besides,there were 5 same pathways,positive regulation of cell death,PID AP1 pathway,reactive oxygen species metabolic process,response to inorganic substance.All PPIs network could be divided into 6 sub-functional modules after it was split.After analysis,56 negative molecules were obtained.Finally,we used PaDEL-Discriptor to calculated 2D molecular descriptors of 51 NAC compounds and 56 negative molecules to obtain s imilar compounds.A total of 29 DEGs,2 up-regulated genes and 27 down-regulated genes.There were 25 similar molecules with Tanimoto coefficient more than 0.7.Besides,there were obvious clusters distribution of the compound skeleton structures,suggesting these 25 compounds may be the basis medicinal substances of NAC.In order to search IS drugs that have been listed on the Drugbank,we found related targets through reverse docking with PharmMapper,transforming it into real gene IDs by Uniprot.The intersection of three gene sets were MMP3,F2,NOS and PLAT.Using molecular docking technology,docking sites and linking res idues of 69 positive drugs and NAC compounds with corresponding targets were analyzed.Meanwhile,the target inhibitors and non-inhibitors were found from CHEMBL as the training sets and validation sets to constructed and screened suitable models RF,ELM,and DT for classifing NAC compounds.Models were run randomly for 50 times with the accuracy of screening and classification models all greater than 92.4%.Beta-carotene of the NAC had effects on the four targets respectively.Diop and Celabezine could act on the three targets.Some literatures had verified that these three substances had repairing effects on IS-related pathological mechanisms.The MCAO model was made by classical Longa method.Effect of each group was evaluated by behavioral score and infarct area ratio.ELISA kits were used to determine Prothrombin(F2),Metalloproteinase matrix 3(MMP3),Tissue plasminogen activator(PLAT),Endothelial nitric oxide enzyme(NOS3).The results showed that NAC could improved behavioral score of rats(P<0.05)and reduced the infarct area ratio(P<0.05).Bsides,NAC could reduce the content of MMP3 and PLAT(P<0.05)and increase the expression of NOS3(P <0.05),while there was no significant difference on F2(P> 0.05).In the process of NAC prevention and treatment in IS,possible medic inal ingredients may be ?-carotene,luteolin,Frutinone A,kaempferol,quercetin,baicalin,myricetin,chrysanthemaxanthin,folic acid and ginsenosides.Possible regulatory mechanisms included NAC could increase the expression of NOS3,promoting the production of NO,maintaining the balance of phosphate metabolism in patients,and thus strengthening neuroprotection.Besides,NAC moderately reduced MMP3 and induced the expression of t-PA,while slows down the apoptosis and death of brain nerve cells.Reducing the probability of stroke type from ischemic stroke to hemorrhagic stroke and tissue edema caused by hemorrhagic brain.