| Objective: Alzheimer’s disease(AD)is a neurodegenerative disease characterized by neurological and psychiatric symptoms such as progressive cognitive and memory impairment,personality changes and language disorders.AD is caused by the imbala Normale between the production and degradation of β-amyloid(Aβ)in the brain,which leads to the deposition of Aβ in the brain.The accumulation of neurotoxic Aβ and oxidative stress lead to neuronal death,which causes a series of pathological changes in the brain.Studies have shown that human symbiotic microorganisms are also very important environmental factors that affect host health.They not only play an important role in digestion,nutrition,growth,immunity,inflammation,etc.,but also are closely related to the occurre Normale of many diseases.Recent studies have shown that changes in intestinal flora are closely related to the occurre Normale of central nervous system diseases.Changes in the tract flora will affect the fu Normaltion and behavior of the brain.Some scholars have proposed the co Normalept of intestinal flora-gut-brain axis,and believe that the imbala Normale of the intestinal flora causes the intestinal epithelium to be affected,thereby damaging the intestinal barrier,and a series of inflammatory factors that are produced cross the intestinal barrier and enter the brain The tissue further exerts a series of pathophysiological effects on the brain,resulting in diseases.Traditional Chinese medicine and traditional Chinese medicine compounds produce extensive and complex interactions between the gastrointestinal tract and the flora.Punicaridin is a relatively specific component of the traditional Chinese medicinal material pomegranate peel and has many pharmacological activities.Among them,anti-inflammatory and antibacterial is one of the important pharmacological effects of punica granatum.At present,there are few researches on the relationship between punicalagin and AD.The research in this class aims to explore safety by using punicalarin as an experimental drug based on the adjustment of the "gut-brain" axis and the effect of punicalagin on the cognitive function of AD mice and its possible mechanism.Methods: 48 APP/PSN AD mice were divided into 4 groups,and the following methods were used for intragastric administration,PBS solution(APP/PSN group),low-dose punicalagin(12.5 mg per kilogram of body weight,Pun-L group),and medium-dose safety Punicarin(25mg per kilogram of body weight,Pun-M group),high-dose punicalagin(50mg per kilogram of body weight,Pun-H group).Twelve C57 b L/6 mice were intragastrically administered with PBS as a normal control group(Normal Control group,NORMAL group).Gavage was continued for 45 days.The water maze space exploration and positioning cruise test were performed on each group of mice to judge the cognitive fu Normaltion of the mice.Take mouse intestinal contents,intestinal tissue and brain tissue.The contents of the intestine were handed over to BGI for 16 s DNA seque Normaling and bioinformatics analysis.Q-PCR experiment detects the expression of inflammatory mediators(IL-6,TNF-α,NLr P-3,Nod-2)m RNA in the colon tissue of mice.Immunohistochemical method was used to detect the expression of ZO-1 and Occludin protein in mouse intestinal epithelium and the expression of Aβ42 and tau protein in the CA1 region of mouse hippocampus.Western Blot experiment detects the expression of β-amyloid(Aβ)in mouse brain tissue.Results:(1)The effect of punicalagin on the cognitive function of AD mice.As the learning progressed,the escape latency of the five groups of mice all became a decreasing trend.Compared with the Normal group,the mice in the Model group had significantly longer time(P<0.01).Compared with the Model group mice,the time of the AD mice given punicalagin by gavage was reduced.Among them,the Pun-H group mice were significantly lower than the Pun-L group(P<0.01),and with the increase of the gavage dose,the daily escape latency of mice has a downward trend.After the platform was removed,as the number of days increased,the staying time of the five groups of mice in the target quadrant all tended to increase.Among them,the Model group and the Normal group are significantly different(P<0.01).Compared with the Model group mice,the AD mice given punicalagin had a longer stay in the target quadrant.And with the increase of the gavage dose,the stay time of AD mice in the target quadrant tended to increase.The Pun-H group mice were significantly higher than the Pun-L group mice(P<0.01).From this we can conclude that punicalagin can improve the spatial cognitive function of AD mice,and the higher the dose,the more significant the improvement effect.(2)The effect of punicalagin on the gut microbiota of AD mice The difference in species composition between the Normal group and the Model group is small,while there are certain differences in the species composition between the punicalagin group and the Model group.Among them,the phylum Bacteroides,Firmicutes,and Verrucomicrobia belong to Ekmania big different.It can be seen from the overall scope that the composition of the flora of AD mice is significantly different from that of normal mice,but the distribution of the flora is changed after gavage punicalagin.Among them,the difference of the overall composition between the Pun-M group and the Normal group is the smallest and closest to normal mice.(3)Effects of punicalagin on the gene expression of related inflammatory mediators in the intestinal tissue of AD mice Compared with the mice in the Normal group,the m RNA levels of pro-inflammatory factors IL-6,TNF-α,NLr P-3 and Nod-2 genes in the Model group were significantly increased(P<0.01).After intragastric administration of punicalagin,the m RNA expression of intestinal pro-inflammatory factors in AD mice was significantly reduced(P<0.01).As the gastric dose increased,the expression level showed a significant decrease(P<0.01).Among them,the expression levels of related genes in the Pun-H group mice were even lower than those in the Normal mice(P<0.01).(4)The effect of punicalagin on the expression of intestinal barrier protein in the intestinal wall of AD mice.Normal mouse ZO-1 and occludin proteins are distributed neatly along the top of the mouse intestinal mucosal epithelial cell membrane,and the brown signal is strong.The positive staining in the model group was significantly weakened,and the expression of ZO-1 and occludin protein was disordered,showing a discontinuous distribution.After intragastric administration of punicalagin,the expression of both was significantly increased,and the higher the intragastric dose,the stronger the expression and the more continuous the distribution.It is suggested that punicalagin can up-regulate the expression of tight junction proteins ZO-1 and occludin in the intestinal wall of mice,and the expression of high-dose group is basically the same as that of normal group.(5)The effect of punicalagin on the expression of Aβ42 and tau in the hippocampal CA1 area of AD mice A large number of normal nerve cells can be seen in the hippocampal CA1 area of normal mice,and there is almost no deposition of Aβ42 and tau protein in the cytoplasm.A large amount of Aβ42 and tau protein can be seen in the nerve cells of the hippocampal CA1 area of AD model mice.After intragastric administration of punicalagin,the two were significantly reduced,and the hippocampal CA1 area of AD mice in the middle and high dose groups of punicalagin was close to that of normal mice.(6)The effect of punicalagin on the accumulation of Aβ amyloid in the brain tissue of AD mice Normal mice had the least expression of Aβ protein,and the Model group had the most expression of Aβ protein.After intragastric administration of punicalagin,the expression of Aβ protein in AD mice decreased significantly,and the expression of Aβ protein in the low,medium,and high dose groups decreased sequentially.Conclusions: The punicalagin can significantly improve the cognitive fu Normaltion of AD mice,and can effectively regulate the structure of the intestinal flora of AD mice to make it close to normal mice.The punicalagin can alleviate the inflammatory response in the intestines of AD mice,can improve intestinal barrier function.Punicalagin can reduce the expression of Aβ42 and tau protein in the hippocampus of AD mice and reduce the expression of Aβ protein in the brain of AD mice,and the inhibitory effect of the low,middle,and high dose groups of punicalagin was successively enha Normaled. |
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