| Objective:As a neurodegenerative disease,there is no drug that can reverse the course of alzheimer’s disease.Ginsenoside Rgl is currently found to have an improvement effect on alzheimer’s disease,and is also an important active ingredient in pseudo-ginseng.The Wnt signaling pathway plays an important role in neurodegenerative diseases,especially AD.To investigate whether the improvement of cognitive function of ginsenoside Rgl in the model of AD tree shrew induced by d-galactose combined with agent A 25-35 was related to Wnt signaling pathway and intestinal microbial balance.Methods:The experiment is divided into two parts.First part D-galactose combined with agent A β25-35 induced the establishment of AD tree shrew model.Forty 6 month old males were randomly divided into the control group(CT),d-galactose group(D),A β group(Aβ),and d-galactose group combined with Aβ25-35(D+A),with 10 in each group.The hippocampus of tree shrews in group Aβ and group D+A were injected with 4 ul of Aβ25-35(5mg/ml)at one time.CT and D group tree shrews received a single injection of normal saline.Group D and group D+A were subcutaneously injected with d-galactose(125mg/kg/day),while the CT group and group A β were subcutaneously injected with the same dose of normal saline for 8 weeks.The Morris water maze test was performed at the ninth week.The hippocampus and cortex of the brain were taken for pathological analysis.Immunohistochemistry detected the expression of APP in the cortex and hippocampus,HE staining detected the number of active cells in the cortex,and detected the expression of phosphorylated Tau protein in the cortex and hippocampus.Second part Relationship between ginsenoside Rgl and Wnt signaling pathway in improving cognitive impairment in tree shrews with alzheimer’s disease.Sixty 6 month old males were randomly divided into control group(CT),model group(D+A),model + donepezil hydrochloride group(30mg/kg/D,DN group),Rgl low concentration group(7.5mg/kg,L group),Rg1 medium concentration group(15mg/kg,M group),and Rgl high concentration group(30mg/kg,H group),with 10 males in each group.The dosages of d-galactose and agent A β 25-35 is the same as in the first part.The water maze test was conducted in week 9.After the test,the hippocampus and cortex of the brain were taken for pathological analysis to explore the effects of ginsenoside Rgl on the expression of APP,DNA,iba-1,b-catenin,p-GSK-3 β/GSK-3 β,p-tau/Tau,MAP2,NeuN and LRP1.The contents of each group of large intestine were taken,the v3-v4 region was amplified with bacterial 16SrDNA universal primer,and lllumina PE250 platform was used for sequencing.Results:First part ①D-galactose in combination with A β 25-35 significantly reduced the learning and memory abilities of tree shrews.Morris water maze results showed that tree shrews in group D,group A,and group D+A showed impaired cognitive and memory functions in the evasive latency and crossing times.Compared with the CT group,the evasive incubation period of group D,group A β,and group D+A were significantly increased(P<0.01).The evasive incubation period of group D+A was significantly longer than that of group D and group A β(P<0.05).In the experiment of crossing the platform,compared with the CT group,the crossing times of tree shrews in group D,group Aβ and group D+A were significantly reduced(P<0.05).The crossing times of tree shrews in group D+A were significantly lower than those in group D and group A β(P<0.05).②D-galactose in combination with A βsignificantly enhanced the pathological features of AD.In western blot results,compared with the CT group,there was a significant increase in the level of A β 1-42 in the hippocampus and cortex of tree shrews in group D,group Aβ and group D+A(P<0.05),while the level of A β 1-42 in group D+A was significantly higher than that in group D and group Aβ(P<0.05).The expression of phosphorylated Tau-404 protein and total Tau in cortex and hippocampus showed that the ratio in CT group was significantly lower than that in group D,group A β and group D+A(P<0.01),and that in group D+A was significantly higher than that in group D and group A β.The statistical results of HE staining showed that the number of active cells in group D+A was significantly lower than that in group D and group Aβ.Second part ①Rg1 enhanced AD tree shrews learning ability and memory ability.Compared with D+A group and D+A group,the incubation period of CT,DN,M and H groups was significantly shorter during the 4-6 days of Morris water maze experiment,and the difference was extremely significant(P<0.01).Compared with group L,the incubation period of group M and group H was significantly shortened(P<0.05).Compared with group D+A,the number of crossing platforms in group L increased significantly(P<0.05),compared with group D+A,the number of crossing platforms in group CT,DN,M and H increased significantly(P<0.01),compared with group L,the number of crossing platforms in group M and H increased significantly(P<0.01).②Ginsenoside Rg1 inhibits Tau phosphorylation and improves the pathological features of AD tree shrews.Western blot analysis of the expression of phosphorylated Tau 404 protein/Tau in the cortex and hippocampus showed that the ratio of the two groups in group D+A was significantly higher than that in group DN,group L,group M and group H(P<0.01),and that in group H was significantly lower than that in group L and group M.There was no significant difference in the expression of APP in cortex and hippocampus of six groups of tree shrews(P>0.05).③Ginsenoside Rg1 inhibits inflammatory gliosis in AD tree shrews.Immunohistochemical analysis showed that the expression of microglia marker iba-1 in the hippocampus and cortex of tree shrew in D+A group was significantly higher than that in DN group,L group,M group and H group(P<0.01),and that in H group was significantly lower than that in L group and M group(P<0.01).④Ginsenoside Rg1 increased the expression of neuronal markers in hippocampus and cortex of AD tree shrews.Western blot statistical results of MAP2 and NeuN showed that group D+A was lower than group L,group M and group H(P<0.01),and group H was significantly higher than group L and group M(P<0.01).⑤Activation of ginsenoside Rg1 on Wnt signaling pathway.The expression of β-catenin in cortex and hippocampus showed that the content of-catenin in D+A group was significantly lower than that in DN group,L group,M group and H group(P<0.01),and that in H group was significantly higher than that in L group and M group(P<0.01).The ratio of expression D+A in cortex and hippocampus in group D+A was significantly lower than that in group DN,group L,group M and group H(P<0.01),and significantly higher in group H than that in group L and group M(P<0.01).⑥Ginsenoside Rg1 improves intestinal flora disorder in AD tree shrews.In the coliform group of tree shrews,the bacteroides abundance of group D+A was 0.2867%,which was significantly higher than that of group CT(0.0255%,P<0.01).However,Rg1 treatment significantly reduced the abundance of bacteroides in the intestinal tract of tree shrews of AD(0.0486%),with no significant difference from the CT group(P>0.05).Conclusion:The model tree shrews induced by d-galactose combined with Aβ 25-35 were consistent with the characteristics of AD in behavior and pathology,the learning and memory ability of tree shrews decreased,and the alzheimer’s disease model was successfully established.Ginsenoside Rg1 can improve the pathological features of AD,and its mechanism involves the activation of Wnt signaling pathway.Ginsenoside Rg1 can improve neuronal inflammation in cortex and hippocampus of AD tree shrews by activating Wnt signaling pathway,and protect neurons in hippocampus and cortex of AD tree shrews and regulate microbial balance in the gut. |
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