Exploring The Pathogenesis Of Alzheimer’s Disease Based On Gut Microbiota-Gut-Brain Pathway Through Whole-intestinal Microbiota Transplantation

Objective: To explore the pathogenesis of Alzheimer’s disease based on the gut microbiota-gut-brain pathway and to provide the experimental basis and new ideas for the research and development of anti-Alzheimer’s disease drugs by transplanting the whole gut microbiota of Alzheimer’s disease(AD)model mice into normal mice.Methods: The 6-month-old SPF male APP/PS1 double-transgenic AD model mice were used as donors,and the whole intestinal contents were taken to prepare bacterial suspension.The normal C57BL/6J mice of the same age and background treated with antibiotics were continuously gavaged with the whole intestinal flora suspension for microbiota transplantation for 14 days,once a day.The model control group and the normal control group were given an equal volume of normal saline every day during the same period.After the microbiota transplantation and colonization for 2 weeks,Congo red staining was used to observe the deposition of amyloid plaques in the hippocampus of mice,and hematoxylin-eosin(HE)staining was used to observe the pathological changes of the mouse hippocampus,ileum and colon tissue.Enzyme-linked immunosorbent assay(ELISA)was used to detect the content of tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β)and interleukin-10(IL-10)in hippocampus,ileum and colon of mice.The 16 S microbial three-generation high-throughput sequencing was used to analyze the microbiota composition and diversity of ileal and colon contents,including Alpha diversity,Beta diversity,microbiota composition at different taxonomic levels and differential species analysis.Results: 1.The results of Congo red staining showed that there were no amyloid plaques in the hippocampus of the mice in the normal control group,obvious red amyloid plaques were seen in the hippocampus of the mice in the model control group,and red amyloid plaques were occasionally seen in the hippocampus of the mice in the whole-intestinal microbiota transplantation group.The results of HE staining showed that there was no obvious pathological damage in the hippocampus,ileum and colon tissue of the normal control group.The neuronal cells in the hippocampus of the model control group were shrunken and hyperchromatic,the intercellular space was widened,and the mucosal tissue in the ileum was shrunken and hyperchromatic.The structure is incomplete,the cells at the top of the intestinal villi are shed,the mucosa in the colon is eroded or ulcerated,the mucosal epithelium is shed in a large area,the lamina propria is damaged,and the fibrous tissue is hyperplasia.The degree of pathological damage in the hippocampus,ileum and colon of the mice in the wholeintestinal microbiota transplantation group was between that of the normal control group and the model control group.2.The results of inflammatory indicators showed that compared with the normal control group and the model control group,the ileal TNF-α content of the mice in the whole-intestinal microbiota transplantation group was significantly increased(P<0.05,P<0.01);Compared with the normal control group,the contents of IL-1β in the hippocampus and colon tissue of the mice in the model control group and the whole-intestinal microbiota transplantation group were significantly increased(P<0.05,P<0.01),and the IL-1β content in the ileum tissue of the mice in the whole-intestinal microbiota transplantation group was also significantly increased compared with the normal control group(P<0.05).However,there was no significant difference in IL-10 content in the hippocampus,ileum and colon tissue of each group(P>0.05).3.The results of ileal flora analysis showed that compared with the normal control group,the Chao1,Simpson,Shannon,and PD indices of the model control group and the wholeintestinal microbiota transplantation group were significantly decreased(P<0.01),while there was no significant difference in each index in the model control group and the whole-intestinal microbiota transplantation group.In addition,the Beta diversity and the community composition at the six taxonomic levels of phylum,class,order,family,genus,and species were also significantly different between the normal control group and the model control group,while the ileal microbiota of the mice in the whole-intestinal microbiota transplantation group approached the ileal microbial system of the model control group as a whole.Among them,Lactobacillus,Muribaculum and Pseudomonas played the most important roles.4.The results of colonic flora analysis showed that there was no significant difference in the Alpha diversity index of mice in each group,and the composition of dominant species at different taxonomic levels was also not significantly different.However,the colony structure and community composition at different taxonomic levels in the whole-intestinal microbiota transplantation group were still close to those in the model control group.Conclusion: Whole-intestinal microbiota transplantation in normal mice can form a gut microbial system similar to that of the donor model mice,indicating that the gut microbiota of the Alzheimer’s disease model mice has successfully colonized normal mice.Whole gut microbiota transplantation can not only induce pathological damage and inflammatory response in ileal and colonic mucosa of normal mice,but also lead to injury and inflammatory response of central nervous cells,and a certain degree of Aβ deposition in hippocampus.In addition,Lactobacillus,Muribaculum and Pseudomonas may be potential target bacteria for the treatment of Alzheimer’s disease based on Intestinal Microflora.