Influence And Mechanism Of Gut Microbes On Cognitive Function In Alzheimer's Disease Model Mice

Background Alzheimer's disease(AD)is one of neurodegenerative diseases,which is associated with gut microbiota.However,studies related to gut microbiota have been stricted to observation of the structure of gut microbiota,while investigations on neurobehavioral changes and the pathogenesis of AD are limited.In our study,we analyzed the relationship between intestinal microflora and pathogenesis of AD by using specific pathogen free(SPF)APPswe/PSI?E9 double transgenic AD model mice(PAP mice),SPF wild type(WT)mice,germ-free(GF)PAP mice and GF WT mice,flited probiotic of AD and produced new ideas of AD therapy.Method 1)Groupl included SPF PAP mice and WT mice at 2,5,10 and 12 months old,group2 included SPF PAP,SPF WT,GF PAP and GF WT mice at 5 months old,group3 included SPF PAP,SPF WT and SPF PAP mice gavaged with supernatant containing Alistipes finegoldii JCM16770(SPF AF)and group4 included GF PAP,GF WT and GF PAP mice gavaged with supernatant containing Alistipes finegoldii JCM16770(GF AF).2)The composition of intestinal bacterial communities in the groupl mice was determined by 16S ribosomal DNA sequencing(16S rDNA Seq)3)Morris water maze,Y maze,T maze,Object recognition test(ORT)and Open field test(OFT)were used to measure cognitive function of group2,3 and 4 mice.4)Pathological examinations,immunohistochemical,immunofluorescence,western blot(WB)were used to observe the changes of amyloid deposition and neuroinflammation.5)Pathological examinations,PAS staining,Elisa and Western blot were used to observe the changes of intestinal barrier function.6)Transcriptome sequencing(RNA Seq)and Gene Expression Omnibus(GEO)database(GSE 36980)were used to filter differentially expressed genes(DEGs)between SPF and germ free(GF)mice.We used Quantitative reverse-transcriptase PCR(qRT-PCR)and western blot(WB)to testify the results of RNA Seq.7)We tested the function of Dusp6 by overexpressing and knocking down of Dusp6 in SH-SY5Y cells and the methods of qRT-PCR,western blot were used to detect the expression level of proteins.Results 1)We found that ? and ? diversity were significantly different between 5-month-old SPF PAP and SPF WT mice.2)Spontaneous alternation of SPF PAP mice,frequency and duration of SPF PAP mice in the novel arm decreased in the Y maze(p<0.05).3)Amyloid ? deposition and neuroinflammation in the brain of SPF PAP mice were much more serious than GF PAP mice.The expression of mucin and tight junction protein of SPF PAP mice's colon decreased.And the levels of IL-6,IL-10 and LPS in the serum of SPF PAP mice increased.4)There were 49 DEGs of SPF PAP mice compared with GF PAP mice,and 19 co-expressed DEGs of SPF PAP mice compared with AD in the GSE36980,which were enriched in the MAPK signaling pathway.5)In vitro,overexpression of Dusp6 inhibited phosphorylation of ERK 1/2 and reduced the expression of APP in the SH-SY5Y cell.Conversely,inhibiting the expression of Dusp6 promoted the phosphorylation of ERK 1/2 and significantly promoted the expression of APP,which were reversed by ERK 1/2 inhibitor.6)? and ? diversity were similar between SPF PAP and SPF AF mice.7)Latency,frequency and duration of SPF AF on the platform and platform zone increased compared with SPF PAP mice in the Morris water maze.Discrimination index of SPF AF mice increased in the ORT test.Spontaneous alternation of SPF AF and GF AF mice,frequency and duration of SPFAF mice in the novel arm increased in the Y maze.Spontaneous alternation of SPF AF mice increased in the T maze.8)Senile plaques in the brain of SPF AF mice decreased,and the expression of TREM2 in the hippocampus of SPF AF and GF AF increased.The form of microglia in the SPF AF and GF AF is amebiform.Conclusion 1)Gut microbiota dysbiosis in SPF PAP mice at 5 months old are most obvious.2)Gut microbiota dybiosis are related with SPF PAP mice cognition impaired.3)Gut microbiota dybiosis are related with low-expressed Dusp6,MAPK signal activation and senile plaques deposition.4)Gut microbiota dybiosis are related with colon barrier impaired and inflammation of SPF PAP mice.5)Dusp6 expression can inhibit the MAPK signal activated and the expression of APP in SH-SY5Y cell.6)Alistipes finegoldii JCM16770 transplantation can improve the structure of gut microbiota of SPF PAP mice,improve the cognition impairment.7)Alistipes finegoldii JCM16770 transplantation can promote the expression of TREM2,promote the phagocytosis of microglia and decreased senile plaques in the hippocampus of SPF PAP and GF PAP mice.