基于网络药理学和动物实验探讨好忘方治疗血管性痴呆作用机制OA

Objective To explore the mechanism of Haowang Prescription in the treatment of vascular dementia(VD)using network pharmacology and animal experiments.Methods Active components and corresponding targets of Haowang Prescription were retrieved from the TCMSP databases;the targets of VD were obtained through the OMIM and GeneCards databases.The protein-protein interaction network was established through STRING database.GO and KEGG enrichment analyses of key targets were carried out by using R language.The VD rat model was established by modified bilateral carotid artery ligation.The modeled rats were divided into model group,donepezil group,Haowang Prescription low-dosage group,Haowang Prescription high-dosage group,Haowang Prescription high-dosage+AMPK inhibitor(Compound C)group,and the sham-operation group was set up.The corresponding intervention was given.The learning and memory abilities of rats were tested by Morris water maze test.The pathomorphological changes in the hippocampus were observed by HE staining.Transmission electron microscopy was employed to observe mitochondrial morphology in hippocampus neurons.The content of inflammatory factors in hippocampus was detected by ELISA.Flow cytometry was used to detect changes in mitochondrial membrane potential and the level of reactive oxygen species(ROS).The expressions of proteins and mRNA of core targets and pathways in the hippocampus were detected by Western blot and RT-qPCR.Results A total of 39 active components of Haowang Prescription and 91 main targets for treating VD were screened,with the core targets including PINK1,NLRP3,CASP1,GSDMD,IL1B,PARK2,SQSTM1,TOM7,TNF and ROS1.KEGG pathway enrichment analysis suggested that AMPK signaling pathway,mitophagy,NOD-like receptor signaling pathway,autophagy and mTOR signaling pathway may play a core regulatory role.Animal experiments showed that Haowang Prescription could significantly improve the learning and spatial memory abilities of model rats(P<0.05,P<0.01),alleviate pathological morphology damage in hippocampal tissue,and alleviate mitochondrial morphological and structural damage in hippocampal tissue.The contents of IL-1β,IL-18,TNF-α and ROS in hippocampus decreased significantly;the mitochondrial membrane potential significantly increased,the expressions of AMPK,PINK1,Parkin,LC3B protein and mRNA were up-regulated,and the expressions of p62,TOMM20,NLRP3,ASC,Caspase-1,GSDMD-N proteins and mRNA were down-regulated(P<0.05,P<0.01).AMPK inhibitor partially reversed the therapeutic effects of Haowang Prescription on mitophagy and pyroptosis.Conclusion Haowang Prescription can reduce pyroptosis in hippocampal tissues of VD rats,and the mechanism may be related to its activation of AMPK/PINK1/Parkin-mediated mitophagy.