Profile Of Bile Acids-FXR-FGF15 Pathway And Gut Microbiota In The Glycolipid Metabolism Disorder Of Diabetic Mice Suffered With Chronic Stress

OBJECTIVEMutual comorbidity of diabetes and depression is a challenge for clinical chronic disease management.Diabetes increases the risk of depression,meanwhile,depressive disorder exacerbates the occurrence and development of diabetes and increases diabetes-related distress.The underlying mechanism of this process is still unclear.This study used the diabetes and depression comorbidity mice model and focused on the profile changes of glycolipid metabolism,bile acid FXR-FGF15 and gut microbiota in type 2 diabetes mice exposed to chronic stress,aimed to explore the mechanism of the role of bile acids-FXR-FGF15 pathway and gut microbiota in the disruption of glucolipid metabolism in diabetic mice exacerbated by chronic stress and to provide a reference for seeking new strategies for prevention and treatment of type 2 diabetes and depression comorbidity.METHODSC57BL/6 male mice were selected and fed with high-fat chow for 4 weeks and then combined with an intraperitoneal injection of streptozotocin to induce a T2DM mouse model,followed by chronic unpredictable mild stress(CUMS)to induce depression-like phenotype to construct a comorbidity model of diabetes and depression.The mice were randomly divided into the Control group,CUMS group,T2DM group,and T2DM+CUMS group by random number method according to the experimental design.Body weight and random blood glucose were monitored regularly every week during CUMS establishment.After 4 weeks of CUMS model construction,behavioral tests(open field test,tail suspension test,forced swimming test)and sucrose preference test were performed to verify the depression model.After the successful establishment of the comorbidity model,fasting blood glucose was measured and blood samples were collected from the mice as well as the liver,pancreas,and ileum.Detection of blood glucose,blood lipids,liver function,total bile acids,and insulin level in mice,tissue weighing and pathological section analysis of liver and pancreas,and detection of mRNA level expression and protein expression of FXR and its related genes in liver and ileum by molecular biology methods.Simultaneously,at the end of the experiment,the feces of four groups of mice were collected,and the abundance of gut microbiota in the colon contents of each group was analyzed by using bacterial 16S rDNA amplicon analysis.RESULTSBlood biochemical analysis showed that chronic stress induced depression-like phenotype given to type 2 diabetic mice had a significant effect on glucolipid metabolism.Comparing with T2DM group,fasting glucose,CHO,TG,LDL and NEFA were significantly increased in mice after giving to chronic stress,with increased liver damage(AST)and increased liver/body ratio,along with increased total bile acid levels.Pathological examination revealed that diabetes and depression comorbidity resulted in moderate to severe hepatic steatosis,islet atrophy,and reduced number of islets βcell clusters in mice.This suggests that comorbidity can further disrupt glucolipid metabolism levels,impair liver and pancreatic function,and worsen pre-existing type 2 diabetes.Molecular biology studies revealed that chronic stress induced depression-like phenotype given to type 2 diabetic mice inhibited bile acid receptor FXR signaling,and that mRNA and protein levels of FXR were reduced in the liver,whereas mRNA and protein levels of SHP,a downstream target gene of FXR in the liver,were higher,suggesting that the disruption of lipid metabolism in diabetic mice following administration of chronic stress may be mediated by the FXR-SHP.In the ileum,both mRNA and protein expression levels of FXR and its downstream target gene FGF15 were reduced.This suggests that the disruption of glucose metabolism in diabetic mice following administration of chronic stress may be mediated by the FXR-FGF15 pathway.Analysis of the gut microbiota results revealed that(1)The sequencing yielded 165 unique OTUs(operational taxonomic units)for the Control group,285 unique OTUs for the CUMS group,168 unique OTUs for the T2DM group,and 188 unique OTUs for the T2DM+CUMS group;(2)In the Alpha diversity analysis,the species richness of the T2DM+CUMS group was not significantly different in Chao1,Ace,Shannon and Simpson indices compared to the T2DM group,indicating that the type 2 diabetic mice were given chronic stress with insignificant differences in flora diversity;(3)In the beta diversity analysis,principal coordinate analysis(PCoA)results showed that CUMS or T2DM significantly altered the intestinal flora of mice;(4)In the species composition analysis,at the phylum level,compared with the T2DM group,the abundance of the mouse intestinal flora of the T2DM+CUMS group was increased in the Firmicutes,Bacteroidetes,Proteobacteria,and decreased in the Verrucomicrobia,Actinobacteria.At the genus level,the abundance of harmful bacteria Aerococcus increased and beneficial bacteria Lactobacillus and Akkermansia decreased in the T2DM+CUMS group compared with the T2DM group.(5)In the indicator species analysis,the dominant bacterial group categories in mice after type 2 diabetic comorbid depression by LEFSe analysis were Deltaproteobacteria,Desulfovibrionales,Desulfovibrionaceae,Proteobacteria,Lachnospiraceae_NK4A136_group and Blautia.By indicator analysis,at the phylum level,the indicator values of mice in the T2DM+CUMS group were significantly increased in Proteobacteria,Deferribacteres and significantly decreased in Verrucomicrobia;in the microbial genus level,indicator values in the T2DM+CUMS group were significantly increased in the harmful bacteria genera Aerococcus,Ruminiclostridium,Rikenella,Ruminococcaceae_UCG-004,GCA-900066225,Peptococcus,Acetivibrio_ethanolgignens_group,Bilophila significantly increased,and in the beneficial genus Akkermansia significantly decreased.CONCLUSIONDepression like behavior induced by chronic stress further interferes with glucose and lipid metabolism in diabetes mice.These changes are associated with bile acid receptor FXR,SHP and FGF15 signaling,and FXR and its downstream target gene FGF15 may be targets for comorbid treatment of T2DM and depression.Diabetes and depression comorbidity is accompanied by a decrease in the relative abundance of beneficial bacteria and an increase in the relative abundance of pathogenic bacteria during the course of disease development,altering the gut microbiota structure,suggesting that diabetes and depression comorbidity may exacerbate the disruption of gut microbiota distribution.