Molecular Mechanism Of Ginsenoside CK Targeted Regulation Of A? Deposition To Protect Neurons And Attenuate Memory Impairment Based On Nrf2/keap1 Signaling Pathway

Objective: This study was based on the Nrf2/Keap1 signaling pathway to investigate the regulation of ginsenoside CK on A? oligomers and neuroprotection.To elucidate the role of CK in the prevention and treatment of molecular mechanisms of Alzheimer's disease(AD),and provide theoretical support and experimental basis for clinical ginseng prevention and treatment of AD.Methods: This study was to investigate the effects of CK on A? ligand binding and regulation,and on the regulation of Nrf2/Keap1 signaling pathway,from in vitro,cellular and animal experiments.To provide a theoretical basis for clearing CK prevention and treatment of AD.1 In vitro CK regulates A? oligomersThe targeting affinity of CK for A?42 ligand was detected by OpenSPR technology.The regulation of CK on A? oligomers was detected by ThT fluorescence assay.Depolymerization of CK to A? oligomers was verified by SDS-PAGE electrophoresis.The inhibitory effect of CK on A? oligomers was observed by electron microscopy.The binding regions and binding levels of CK and A? were estimated by molecular modeling.2 CK protects against damaged SH-SY5 Y cellsThe effect of CK on the survival rate of damaged SH-SY5 Y cells was detected by CCK-8 assay.The regulation of oxidative free radicals in damaged SH-SY5 Y cells was verified by ROS detection technique.The regulation of CK on A? was detected by Western blotting.Role and regulation of Nrf2/Keap1 signaling pathway.3 CK protects against damaged HT22 cellsReal-time monitoring technology was used to monitor the protective effect of CK on damaged HT22 cells.The effect of CK on the survival rate of damaged HT22 cells was detected by MTT technique.The morphology of cells was observed by light microscopy.The free radicals and fluorescence of CK in damaged HT22 cells were detected by ROS.The regulation of intensity was observed by immunofluorescence,Western blotting and RT-PCR to observe the localization of CK to A?42 and Nrf2 cells,and to detect the regulation of A? and Nrf2 signaling pathway in damaged HT22 cells.4 CK protects mice from AD modelThe effects of CK on memory and learning function of AD model mice were detected by water maze and platform test.The effects of CK on SOD activity,MDA and GSH content in brain of AD model mice were detected by kit.The effects of morphological changes on neuronal apoptosis in AD model mice were observed by TUNEL method.Regulation of A? and Nrf2/Keap1 signaling pathway in brain of AD model mice were detected by immunohistochemistry,Western blotting and RT-PCR.Results:1 CK regulates A? oligomersIn vitro,CK can dock with A?42 monomer and target binding in the ?-sheet domain.The optimal conformational docking Ki value is 4.51 ?M.CK can dissociate A? aggregates and reduce tetramers and high molecular weight multimers.Inhibition of the formation of A? oligomers.2 Protective mechanism of CK on damaged SH-SY5 Y cellsCK could increase the survival rate of damaged SH-SY5 Y cells(P<0.05,P<0.01),reduce intracellular ROS(P<0.05),down-regulate the expression of APP and A?(P<0.05,P<0.01),and down-regulate the expression of BACE1 and PS1(P<0.05,P<0.01).,upregulated the expression of IDE(P<0.05,P<0.01),up-regulated the expression of Nrf2/Keap1 signaling pathway(P<0.05,P<0.01).3 Protective mechanism of CK on damaged HT22 cellsCK increased the survival rate of damaged HT22 cells(P<0.05,P<0.01),increased cell growth curve,decreased ROS production in HT22 cells(P<0.05,P<0.01),up-regulated the expression of SYP and PSD95(P<0.05),and maintained neuronal synapses.Structure and function,down-regulated the expression of APP,BACE1 and PS1(P<0.05,P<0.01),upregulated the expression of IDE(P<0.05,P<0.01),down-regulated the expression of A?(P<0.05,P<0.01),decreased the aggregation of A? between neurons,and up-regulated Bcl-The expression of 2(P<0.05,P<0.01)down-regulated the expression of Bax and Caspase3(P<0.05,P<0.01),promoted the entry of Nrf2 into the nucleus and up-regulated the expression of Nrf2/Keap1 signaling pathway(P<0.05,P<0.01).4 Protective mechanism of CK on AD model miceCK improved the memory function and learning ability of AD model mice,increased the expression levels of SYP and PSD95(P<0.05,P<0.01),increased SOD activity and GSH content in AD mice(P<0.05,P<0.01),and decreased MDA content(P<0.05,P<0.01).The expression levels of APP,BACE1 and PS1 were decreased(P<0.05,P<0.01),the expression level of IDE was increased(P<0.05,P<0.01),the expression level of A? was decreased(P<0.05,P<0.01),and the expression level of Bcl-2 was increased(P<0.05,P<0.01).The expression levels of Bax and Caspase3 were decreased(P<0.05,P<0.01),and the expression of Nrf2/Keap1 signaling pathway was increased(P<0.05,P<0.01).Conclusion: CK can bind to the ?-sheet region of A?42 monomer,promote the depolymerization of A? oligomers,reduce the accumulation of A? in neurons and form A? aggregates,activate Nrf2/Keap1 signaling pathway,and decrease neuronal A?.Inducing neuronal peroxidation damage and toxicity,inhibiting neuronal mitochondrial apoptosis,maintaining synaptic structure and function,improving memory function and learning ability of AD model mice,providing theoretical basis for prevention and treatment of AD.