| Objectives:In this study,we investigated the effect of glucagon like peptide-1(GLP-1)analogues on cognitive impairment of Alzheimer's disease(AD),and its regulatory effect on cerebral glucose metabolism,so as to explore the possible mechanism of GLP-1 analogues in improving cognitive impairment in AD.Methods:4-month-old 5×FAD transgenic mice were randomly divided into the AD group and the liraglutide group(AD+lira group).4-month-old wild-type littermates were used as controls,referred to as the wild-type(WT)group.In AD+lira group,liraglutide(25?g/kg)was injected subcutaneously once a day for 8 weeks;in AD group and WT control group,the same volume of saline was given instead.Morris water maze,Nissl staining and transmission electron microscope were used to evaluate the spatial memory function and brain nerve damage of mice in each group;Western blot was used to evaluate the expressions of PSD95 and SYN.Itraq labeled proteomics was used to analyze the proteomic changes in the cortex of mice in each group and evaluate the changes of metabolic pathways.Western blot was used to detect key enzymes of glycolysis and oxidative phosphorylation,as well as the expression of PI3K/Akt;the content of cerebral ATP,lactate and NAD+and the levels of reactive oxygen species(ROS)and glutathione(GSH)were also tested.Primary astrocytes of Sprague Dawley(SD)rats were treated with A?1-42oligomers to AD model of astrocytes.GLP-1 was used to intervene A?treated astrocytes.MTT assay was used to evaluate cell viability.LY294002 was applied to inhibit the PI3K/Akt pathway and 2-deoxy-D-glucose(2-DG)to inhibit cell glycolysis,respectively;RT-q PCR was used to detect the gene transcription level of glycolysis related enzymes;Western blot and immunofluorescence were used to detect the key enzymes of glycolysis and oxidative phosphorylation,as well as the protein expression of PI3K/Akt pathway.Real-time oxygen consumption rate(OCR)was monitored and the degree of mitochondrial aerobic respiration was detected.DCFH-DA probe and mitotracker probe were used to detect the content of ROS in cells and mitochondria,and the GSH content in cells was detected to evaluate oxidative stress levels.Primary neurons were co-cultured with astrocytes.Survival of neurons in the co-culture system was evaluated by Hoechst/PI staining.The length and branches of axons and dendrites,and the expression of PSD95and SYN were compared by immunofluorescence and Western blot.Results:(1)Liraglutide(a GLP-1 analogue)improved the spatial cognition of 5×FAD mice,with improved survival and synaptic structures of neurons in hippocampus and cerebral cortex;Liraglutide enhanced ATP content in AD mice,regulated metabolic pathways,elevated expressions of PDK2 and glycolytic enzymes,and increased lactate and NAD+contents;meanwhile,it reduced the phosphorylation level of PDH,reduced ROS content,enhanced GSH level,thus alleviated oxidative stress.In addition,Liraglutide activated the PI3K/Akt pathway in the brain of 5×FAD mice.(2)The intervention of 100 n M GLP-1 significantly enhanced the viability of A?-treated astrocytes,increased the expression of glycolytic enzymes and the production of lactate and NAD+;it also elevated the expression of PDK2 and the phosphorylation level of PDH,but decreased OCR and the production of ROS.However,when PI3K/Akt pathway was inhibited,the ability of GLP-1 to enhance glycolysis was aborted,and the level of oxidative phosphorylation was increased.The level of oxidative phosphorylation also increased when glycolysis was inhibited by 2-DG.GLP-1 promoted the survival of neurons in co-culture system,enhanced the growth of axons and dendrites;however,when cellular glycolysis was inhibited by 2-DG,cell survival and synaptic growth were impaired.Conclusion:GLP-1 analogue improved cognitive impairment in AD by increasing brain glycolysis,and down regulating oxidative phosphorylation,so as to reduce oxidative stress and improve astrocyte's neuronal supportive functions. |
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