Mechanistic Study Of Shenqi Yizhi Granules Intreating Alzheimer’s Disease

BackgroundAlzheimer’s disease(AD)is a neurodegenerative disease closely related to aging.It is clinically manifested as a progressive cognitive impairment such as memory decline.Currently,available Western medicine can only improve certain clinical symptoms of AD patients but may have accompanying side effects.Traditional Chinese medicine has a long history of clinical application and offers significant advantages in the comprehensive prevention and treatment of AD.These advantages include precise clinical efficacy,wide range of sources,multiple targets,synergistic effects,and fewer adverse reactions,which warrant further in-depth research.Shenqi Yizhi Granules,based on years of clinical experience,is a formula used for the treatment of mild to moderate AD.The prescription consists of five traditional Chinese medicines:Panax ginseng C.A.Mey,Astragalus membranaceus(Fisch.)Bunge,Scutellaria baicalensis Georgi,Salvia miltiorrhiza Bunge and Alisma plantago-aquatica L.Through optimization of the manufacturing process and confirmation of efficacy,it has been developed into a new Chinese medicine preparation.The formula exhibits effects of invigorating Qi and strengthening the spleen,warming Yang and clearing turbidity,promoting blood circulation and removing blood stasis,as well as detoxifying and dredging collaterals.It is utilized for the treatment of mild senile dementia characterized by Qi deficiency,blood stasis,collateral obstruction,and symptoms such as memory loss,sluggish thinking,fatigue,shortness of breath,and dizziness.Clinical practice of traditional Chinese medicine has demonstrated that Shenqi Yizhi Granules can effectively intervene in multiple pathological processes of AD,yielding positive clinical outcomes without adverse reactions.However,the mechanism underlying its effect on improving cognitive function disorders remains unclear and warrants further research.Modern omics technologies can elucidate the intricate mechanism of action of Chinese medicine from various levels and perspectives.This approach aligns with the characteristics of multi-component,multi-pathway,and multi-target effects exhibited by Chinese medicine formulas and offers an effective means to investigate their mechanisms of action.ObjectiveThe study aims to explore the interaction between the main active ingredients of Shenqi Yizhi granules and protein targets associated with AD.It intends to analyze the binding capacity of the drug with its target proteins,observe the impact of Shenqi Yizhi granules on cognitive impairment,protein expression profiles,metabolite contents,and intestinal flora in the cerebral cortex and hippocampus tissues of APP/PS1 transgenic mice,and elucidate the mechanism by which the drug ameliorates cognitive dysfunction in APP/PS1 mice.MethodsNetwork pharmacology and molecular docking techniques were applied to analyze the interaction network and drug-target molecular binding ability of the main active ingredients of Shenqi Yizhi granules and AD protein targets.Using bioinformatics methods,we investigated the biological functions of the relevant differential proteins,and selected the potential targets for the treatment of AD by network analysis,correlation analysis and protein function analysis.Using molecular docking technology,we analyzed the binding ability of the measured active compounds and potential medicinal compounds of Shenqi Yizhi granules with AD therapeutic targets,and screened the core targets and action pathways of Shenqi Yizhi granules for the treatment of AD.The behavioral effects of Shenqi Yizhi granules on AD model mice were assessed using a nesting test,new object recognition test,and Morris water maze test.4D-DIA proteomics was applied to identify protein expression in mouse cortical and hippocampal tissues.We used plausible protein analysis,fold change analysis and ANOVA to screen plausible proteins and differential proteins,and bioinformatics to perform functional analysis of differential proteins and identify key protein targets,biological processes and pathways of action of Shenqi Yizhi granules.A dual-platform metabolomics approach combining gas chromatography-mass spectrometry(GC-MS)and liquid chromatography-mass spectrometry(LC-MS)was employed to investigate the metabolic effects of Shenqi Yizhi granules on the cortex and hippocampus of APP/PS1 mice.Multivariate statistical analysis,combined multidimensional and unidimensional analysis,and correlation analysis were utilized to screen differential metabolites and quantify the degree of association among significant differential metabolites.Bioinformatics methods were applied to identify the biological functions and metabolic pathways associated with differential metabolites.The impact of Shenqi Yizhi granules on the gut microbiota of APP/PS1 mice was assessed through 16S rRNA high-throughput sequencing technology.Alpha diversity analysis was employed to assess species richness and evenness,whereas beta diversity analysis was performed to evaluate significant variations in community distribution.Additionally,bioinformatics methods were employed to predict gut microbiota functions.ResultsNetwork pharmacology and molecular docking studies revealed that ALB,IL6,TNF,TP53,IL1B,VEGFA,APOE,CTNNB1,and PPARG are potential key targets in the treatment of AD with Shenqi Yizhi granules.Shenqi Yizhi granules have the potential to ameliorate AD through the regulation of lipid metabolism,metal ion metabolism,inflammation,neurotransmitters,and other pathways.Their mechanism of action is closely linked to the active ingredients Ginsenoside Rgl and Aβ1-42,tanshinone ⅡA and BACE 1,as well as baicalin and AchE,which exhibit enhanced binding affinity.The results of behavioral evaluation demonstrated that Ginseng Astragalus Puzzle Granules enhanced the discrimination index of AD model mice in the novel object recognition test and increased the swimming time of AD model mice in the target quadrant during the Morris water maze spatial exploration experiment.Proteomic analysis revealed that Ginseng Astragalus Puzzle Granules up-regulated 130 proteins and down-regulated 166 proteins in the cortical and hippocampal tissues of AD mice.Key nodes in the up-regulated differential protein network included Mapk8ip2,Hltf,Isynal,Kcna4,and Timm17b,which were associated with functions such as membrane transport of amino acids,regulation of metal ions,complement activation,and mitochondrial energy metabolism,thereby modulating pathways related to glucose metabolism,the complement system,and lipid metabolism.Similarly,key nodes in the down-regulated differential protein network included Rnasel,Nfkb1,Mapk14,Sh3pxd2b,Nedd8,Isg15,and Acvrl,which were involved in functions associated with synaptic translation and MAPK signaling pathways,contributing to the regulation of lipid metabolism,immune regulation,and inflammatory pathways.Metabolomics analysis revealed that Shenqi Yizhi granules exerted significant regulatory effects on 144 metabolites in the cortex and hippocampus of AD model mice,with 71 metabolites upregulated and 73 metabolites downregulated.These metabolites encompassed 6 benzene compounds,ConclusionThe mechanism of action is intricately associated with the strong binding affinity of ginsenoside Rg1 to Aβ1-42,tanshinone IIA to BACE1,and baicalin to AchE as their bioactive constituents.By modulating the expression of complement system components,lipid metabolism,amino acid transmembrane transport,and proteins associated with mitochondrial energy metabolism in the cerebral cortex and hippocampus of APP/PS1 transgenic mice,Ginsenoside Rg1 influences the biosynthesis of lipids,lipid-like molecules,organic acids and their derivatives,organic oxygen compounds,nucleosides,nucleotides,and their analogs,thereby inducing alterations in the composition and population structure of the mice’s intestinal microbiota.Consequently,this leads to improvements in the intestinal microenvironment,enhancement of learning and memory capacity in APP/PS1 mice,and provision of cerebral protection.Through a variety of modern pharmacological experimental methods,this study explains the mechanism of Shenqi Yizhi granule in improving cognitive dysfunction in AD from multiple levels and perspectives,and provides data support and scientific basis for the prevention and treatment of cognitive dysfunction with traditional Chinese medicine.