The Gut Microbiota-bile Acids-anorexigenic Neuropeptide Axis Modulates The Sensitivity To High Fat Diet-induced Obesity In Mice

Background: High calorie diet is an important factor leading to obesity and overweight,but the phenomenon of high-fat diet-induce obesity resistance is widespread in people and animals,and the mechanism is not clear.Gut microbiota and its metabolites secondary bile acids affect the occurrence and development of obesity by regulating energy metabolism.Serum bile acids concentration is closely related to the appetite of obese people.In this study,C57 BL / 6J mice were randomly fed with either a high-fat diet(HFD)or a low-fat diet(LDF)for 12 weeks.According to the different body weight gains,we selected the diet-induced obesity resistance(DIR)and diet-induced obesity(DIO),and the metabolic characteristics were observed.We investigated the changes of bile acid profiling in plasma,intestinal content and feces among LFD,DIO and DIR mice.Correlations between changes in microbiota composition and host metabolic parameters were investigated by performing Spearman’s correlation analysis.To investigate the effects and mechanisms of bile acids on orexin regulation,GT1-7 cells were treated with bile acids in different concentration.Methods: In this study,Six-week-old C57BL/6J male mice were randomly assigned to receive either a high-fat diet(HFD)diet or a low-fat diet(LFD)for 12 weeks.After 10-week HFD feeding,we defined those with significantly increased body weight gains on the HFD as ‘‘diet-induced obese’’(DIO)mice,and those with body weights ranging between the average ± 3 standard deviations of the LFD group as ‘‘diet-induced obese resistant’’(DIR)mice.Metabolic cage experiments were performed before the mice were anesthetized for blood harvesting.After the mice were sacrificed,serum,small intestine contents and feces were collected,and bile acid concentrations were measured by ultraperformance liquid chromatography mass spectrometry.Spearman correlation analysis was performed to study the correlation between changes in microbiota composition and host metabolic parameters.GT1-7 cells were treated with 10 nM,100nM,1μM,10μM tLCA or CDCA for 12,24 and 48 h,respectively.Real-time PCR was used to detect the mRNA expression level of appetite-suppressing signal peptide(POMC)in GT1-7 cells after treatment with different concentrations of bile acids for different times;ELISA method was used to detect the level of orexin α-melanin Cytokine(α-MSH)which was the processing products of POMC.Western blot was used to detect the expression of G protein-coupled bile acid receptor(TGR5)and farnesol X receptor(FXR),phosphorylated signal transduction transcription activation factor 3(p-STAT3),and phosphorylated protein kinase(p-AKT),the expression of cytokine signaling inhibitor 3(SOCS3)protein.Results: We found increase energy expenditure in DIR mice.In addition,the plasma bile acid of DIR mice was significantly decreased(CA,-MCA,-MCA and UDCA),while intestinal bile acids were significantly increased(T-CA,T--MCA,T--MCA and T-UDCA).The composition of the gut microbiota changed drastically(higher levels of Ruminiclostridium and Mucispirillum,and reduction of Bacteroides).Our results indicated that the DIR mice exhibited higher energy expenditure,which may be related to the altered gut microbiota and increased levels of intestinal T--MCA.The level of POMC mRNA expression and the release of α-MSH were increased in GT1-7 cells treated with bile acids.Bile acid treatment up-regulated the expression of p-STAT3 and p-AKT proteins in GT1-7 cells,and increased the expression of SOCS3 protein that regulates the STAT3 signaling pathway.Conclusions: Resistant to high-fat induced obesity is possibly related to increased energy expenditure.Gut microbiota alterations and increased levels of tauro--muricholic acid(T--MCA)is one of the important mechanisms that determine the susceptibility to high-fat diet-induced obesity.The inhibition of appetite signal in the hypothalamus by bile acids may prevent obesity development.