The Protective Effect And Mechanism Of Akkermansia Muciniphila On The Intestinal Mucosal Barrier

Part 1Objective:To explore the protective effect of Akkermansia muciniphila on the intestinal mucosal barrier function in mice with acute enteritis induced by Dextran Sodium Sulfate(DSS).Methods:Male C57BL/6 mice were divided into two groups:pseudo germ-free mice and specific-pathogen-free(SPF)mice.Each has 4 groups:the control group,the DSS group,the Akk group and the Akk+DSS group.The mice were treated with antibiotic and antifungal water for one week to become pseudo germ-free,and then the Akk group and Akk+DSS group were gavaged with Akkermansia muciniphila.After 3 weeks of colonization,the DSS group and Akk+DSS group were treated with DSS to establish the acute enteritis model.While the SPF mice were treated with DSS just after one week of adaptive feeding,and at the same time,they were given normal saline or Akkermansia muciniphila by gavage.The disease degree of mice was assessed by disease activity index(DAI).The inflammation of mice was assessed by the length of the colon,the expression level of inflammatory factors in colon tissue and the pathological inflammation score of the colon.The intestinal mucosal barrier function of mice was evaluated by intestinal mucosal permeability test.The expression level of proteins such as ZO-1,E-cadherin,Occludin and Claudin-1 was assessed by Western blot.Results:In the pseudo germ-free mice model,compared with the DSS group,the DAI score of the Akk+DSS group was significantly reduced(P=0.0195),and the expression level of TNF-? in colon tissue was significantly decreased(P=0.0286).What's more,the Akk+DSS group has longer colon,lower pathological inflammation score,lower expression level of IL-6 and IL-1? higher expression level of barrier protein(ZO-1,E-cadherin,Occludin and Claudin-1)and lower mucosal permeability,but there is no statistical significance.In the SPF mice model,The DAI score,colon length,and histopathological score of the Akk+DSS group were not significantly different from those of the DSS group.Conclusion:Akkermansia muciniphila can reduce the permeability of colon mucosa by up-regulating the expression levels of Occludin,ZO-1,claudin-1 and E-cadherin proteins in colon tissue;at the same time,it can reduce the destruction of intestinal mucosal epithelium and crypts,thereby alleviating acute enteritis induced by DSS in pseudo germ-free mice.Part 2Objective:To explore the protective effect of Akkermansia muciniphila on the mucosal barrier function in mouse colon organoids with inflammation induced by TNF-?.Methods:The mouse colon organoid model was constructed and divided into 4 groups:the control group,the Akk group,the TNF-? group,the Akk+TNF-? group.The Akk group and the Akk+TNF-? group were co-cultured with 1 × 105 CFUs/ml Akkermansia muciniphila for 24 hours.At the same time the TNF-? group and the Akk+TNF-? group were stimulated with 120ng/ml TNF-? for 24 hours to construct an inflammatory model.The diffusion of FITC-dextran into the intestinal cavity was used to compare the intestinal mucosal permeability between the groups.Results:The fluorescence intensity of the colon organoids in the TNF-? group was higher than that of the other three groups,but they were not statistically significant(P=0.1000).Conclusion:The co-culture of Akkermansia muciniphila can improve the permeability of mouse colon organoids induced by TNF-? stimulation to a certain extent.Part 3Objective:To investigate the relationship between the disease activity of UC patients and the level of Akkermansia in the intestinal mucosa.Methods:UC patients admitted to Peking Union Medical College Hospital from January 1,2018 to June 30,2019 were enrolled.According to the Mayo score(1-4 points)under endoscopy,the patients were divided into remission group(0 point)and active group(1-3 points).And then the intestinal mucosal tissues of the patients were collected for flora analysis(16S rDNA sequencing),and the level of Akkermansia in the remission group and the active group was compared.Results:The amount of Akkermansia in the colonic mucosa of UC patients in the remission group was significantly higher than that in the active group(P=0.048).Conclusion:Compared with patients with active UC,patients with endoscopic remission have higher levels of Akkermansia in the colonic mucosa.