| Objective:Based on the “Traditional Chinese Medicine Inheritance Supporting Platform(V2.5)”,this study summarized Prof.Yang lihua’s prescription experience in the treatment of post-stroke cognitive impairment(PSCI),and explained the potential therapeutic mechanism of Prof.Yang lihua’s core prescriptions for the treatment of PSCI through network pharmacology and animal experiments.Methods:1 This study sorted out 317 medical records of patients with PSCI diagnosed and treated by Prof.Yang lihua from August 2018 to March 2021.Using the sub-modules of the "Data Analysis" in the "Traditional Chinese Medicine Inheritance Supporting Platform(V2.5)" software to analyze 317 prescriptions.Based on this,the laws of Prof.Yang lihua’s prescriptions and medications were sorted out.2 This study relied on databases such as The Encyclopedia of Traditional Chinese Medicine(ETCM),Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP),and Gene Card,and used research platforms such as Search Tool for Recurring Instances of Neighboring Genes(STRING)and The Database for Annotation,Visualization and Integrated Discovery(DAVID),and analysis software such as Cytoscape.Subsequently,the study constructed drug-ingredient-target networks and PPI networks,and further established Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analysis,then systematically organized the targets of the core drugs combination.3 In this study,SD rats were randomly divided into blank group,model group,control group,and low-,medium-,and high-dose core drugs combination groups.Rat models with bilateral common carotid artery occlusion(BCCAO)were established except the blank group.After 28 days of drug administration,the cognitive function of rats was examined by Morris water maze(MWM)test,radial arm maze(RAM)test and the novel object recognition(NOR)test;the pathological changes of rat brain tissue were observed by hematoxylin-eosin(HE)staining and transmission electron microscope(TEM);the m RNA expressions of synapse-associated proteins(SAPs)were detected by PCR;the protein expression levels of p-Akt,Akt,p-CREB,CREB,and BDNF were detected by immunohistochemistry(IHC)and Western Blot(WB).Results:1 In the section of clinical medical records,a total of 317 outpatient medical records of Prof.Yang lihua in the treatment of PSCI patients were entered(including spleen-kidney deficiency syndrome,liver and kidney deficiency syndrome,kidney deficiency and blood stagnation syndrome,phlegm turbidity and blood stasis syndrome and kidney essence depletion syndrome).Based on this,60 high-frequency drugs were sorted out,then162 medication patterns,263 prescription rules,and 24 potential new combinations of traditional Chinese medicines were extracted.2 The network pharmacology research in this paper relied on multiple data platforms,then systematically organized the targets of core drugs,as well as potential therapeutic targets and molecular pathways for the treatment of PSCI.The results showed that there were 243 active ingredients and 804 targets in the 13 traditional Chinese medicines we studied,and there were 66 important targets for the treatment of PSCI.PPI network analysis suggested that INS,AKT,BDNF,TNF,IL6,IL1,etc.were the key targets.GO enrichment analysis indicated that these targets involved 361 biological process(BP)items,60 cellular component(CC)items,and 67 molecular function(MF)items,covering the regulation of multiple physiological processes,such as the survival,growth,and plasticity of nerve cells.KEGG enrichment analysis revealed that there were 124 potential pathways involved in the treatment of PSCI by the core drugs combination.All of the above indicated the potential for further investigation of the core drugs combination.3 The MWM test,RAM test and NOR test all indicated that the cognitive function of the rats in the model group was significantly lower than that of the sham group,and after drug administration,the cognitive function of the modeled rats was improved.Among them,the improvement of cognitive function in high-dose core drugs combination group was the most obvious(P<0.05).The results of toluidine blue(TB)solution perfusion indicated that the combination of core drugs could improve the blood perfusion of brain tissue;HE staining results showed that the number of neurons in the hippocampus of rats in the model group decreased,and pathological changes such as hyperchromatism and pyknosis of nuclei appeared;the results of TEM suggested that the rats in the model group showed pathological changes such as chromatin condensation,nuclear membrane rupture,mitochondria and endoplasmic reticulum atrophy,cytoplasmic vacuolization and organelle fragmentation.After drug treatment,the pathological damages of the brain tissue of the rats in each administration group were improved,and the improvement in the high-dose core drugs combination group was the most significant(P<0.05).The results of PCR indicated that the m RNA expression levels of SAPs(SYP,PSD-95,NR2 B and Ca MKⅡ)in the brain tissue of the model group rats were significantly decreased(P<0.05),and the expression levels of the above 4 m RNAs in each administration group all increased(P<0.05),and the increase in high-dose core drugs combination group was the most significant.The results of IHC indicated that the expression levels of p-Akt,p-CREB,and BDNF in the control group were higher than those in the model group,and the expression levels of p-Akt,p-CREB,and BDNF could be further increased after treatment with the combination of core drugs,and the increase was most significant in the high-dose core drugs combination group.The results of WB also indicated that the expression levels of p-Akt,p-CREB,and BDNF in the control group were higher than those in the model group,and each treatment group with core drugs combination was better than those in the control group(P<0.05),and the increase was most significant in the high-dose core drugs combination group.Finally,ELISA was used to detect the expression levels of IL-6,IL-1β,and TNF-α in the brain tissue of rats in each group,and the high-dose core drugs combination group had the most significant reduction.Conclusion:Prof.Yang lihua has rich clinical experience in the treatment of PSCI.In terms of prescription and medication,Prof.Yang lihua pays attention to the importance of tonifying the kidney and replenishing essence,stresses the relationship between kidney essence and brain marrow;emphasizes the homology of the essence and blood and the relationship between congenital essence and acquired essence,highlights that liver and spleen should be taken into account when tonifying the kidney;pays attention to the changes of phlegm turbidity and blood stasis in the course of disease development,and emphasizes the balance between strengthening the body resistance and exorcising evil;The network pharmacology proves that the core drug compatibility of phlegm turbidity and blood stasis syndrome has rich therapeutic targets for the treatment of PSCI,and can regulate the survival and growth of neurons from multiple aspects;Trough animal experiments,We found the core drugs combination for treating phlegm turbidity and blood stasis syndrome can improve the cognitive function of BCCAO rats,reduce the pathological damage of rat brain tissue,regulate synaptic plasticity,reduce the central inflammatory response,and therapy PSCI through the Akt/CREB/BDNF pathway. |
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