Metformin Ameliorate Autism Spectrum Disorder:Contribution Of Gut Microbiota,Gastrointestinal Barrier Integrity,and Serotonin

Objective Autism spectrum disorder(ASD)is one of the most severe neurodevelopmental disorders in the world,characterized by social impairment,repetitive stereotyped behaviors,interests and activity patterns;the prevalence of ASD has increased dramatically in the last decades.There is growing evidence that intestinal flora and its metabolites play an important role in the pathogenesis of ASD.Studies have shown that a high-fat diet(HFD)can exacerbate symptoms associated with autism,and recent studies suggest that metformin(Met)appears to have a potential role in ameliorating ASD.Our previous studies suggest that the effect of Met derives in part from improving the intestinal barrier and regulating the intestinal flora to act.Based on the above background,we wanted to understand whether a high-fat diet exacerbates autism symptoms and whether metformin could improve ASD symptoms by improving barrier function and modulating intestinal flora and 5-hydroxytryptamine(5-HT)in high-fat mice,as well as the relationship between intestinal flora and metabolites(short-chain fatty acids)and autism symptoms in children with autism.Methods 1,C57BL/6 and BTBR autism mouse models were fed with HFD and normal diet(ND)for 8 weeks,respectively,and some BTBR mice were gavaged with the second part of Met(400 mg/kg/d)for 2 weeks for behavioral testing and evaluation of their intestinal barrier index,5-HT and intestinal flora.2,The following behaviors were evaluated using TSE video behavior tracking analysis system,including stereotyped and repetitive behaviors(marble burying and self-grooming);anxiety-like behaviors(open field test and elevated maze test)socialization test(three chamber test).3,Using immunohistochemistry and protein blotting methods to assess intestinal barrier function including:(1)tight junction proteins:occludin(ocludin)and occluded small band protein(ZO-1);(2)mucus barrier:mucin-2(Muc 2);(3)high performance mass spectrometry to detect 5-HT in the colon and cortex;(3)16S-rRNA method to detect intestinal flora.4.To compare the gastrointestinal symptoms,social,sleep,and dietary characteristics of autism(ASD)and typical development children(TD),to analyze the relationship between intestinal flora and fecal short-chain fatty acid levels and GI,social,sleep,and dietary habits in ASD and TD,and to explore the possible role of intestinal flora and its important metabolite short-chain fatty acids in ASD.Results. Compared to controls,the high-fat diet had significantly abnormal weight,blood glucose,and glucose tolerance;behaviorally,while HFD exacerbated some autistic symptoms(stereotyped,repetitive behaviors),it improved for the core symptom of ASD,social impairment.Metformin improved the behavior of normal diet BTBR autistic mice,HFD increased the expression of Muc 2,Met increased the expression of tight junction protein,HFD increased the level of tryptophan,Met increased the level of 5-HT,and HFD colony results differed significantly from the normal diet group.In addition,78%in children with autism had gastrointestinal symptoms and their presence had no significant effect on the core symptoms of autism;the structure of the ASD microbiota significantly differed from the normal children group and the differences were greater in the presence of GI symptoms and SCFA levels were significantly higher than in children with autism.Conclusion. High-fat diet exacerbated autistic behavior but simultaneously improved social behavior.Metformin improved autistic behavior in BTBR mice,and the mechanism may be related through improving the intestinal barrier and regulating 5-HT.Gastrointestinal symptoms are common in autism,and autism may not directly lead to increased gastrointestinal symptoms;differences in flora and changes in their metabolites,such as short-chain fatty acids,may be a potential cause of gastrointestinal symptoms.