Duodenal-jejunal bypass ameliorates MASLD in rats by regulating gut microbiota and bile acid metabolism through FXR pathways

Abstract

Background: Although bariatric and metabolic surgical methods, including duodenal-jejunal bypass (DJB), were shown to improve metabolic dysfunction-associated steatotic liver disease (MASLD) in clinical trials and experimental rodent models, their underlying mechanisms remain unclear. The present study therefore evaluated the therapeutic effects and mechanisms of action of DJB in rats with MASLD.

Methods: Rats with MASLD were randomly assigned to undergo DJB or sham surgery. Rats were orally administered a broad-spectrum antibiotic cocktail (Abx) or underwent fecal microbiota transplantation to assess the role of gut microbiota in DJB-induced improvement of MASLD. Gut microbiota were profiled by 16S rRNA gene sequencing and metagenomic sequencing, and bile acids (BAs) were analyzed by BA-targeted metabolomics.

Results: DJB alleviated hepatic steatosis and insulin resistance in rats with diet-induced MASLD. Abx depletion of bacteria abrogated the ameliorating effects of DJB on MASLD. Fecal microbiota transplantation from rats that underwent DJB improved MASLD in high-fat diet-fed recipients by reshaping the gut microbiota, especially by significantly reducing the abundance of Clostridium. This, in turn, suppressed secondary BA biosynthesis and activated the hepatic BA receptor, farnesoid X receptor. Inhibition of farnesoid X receptor attenuated the ameliorative effects of post-DJB microbiota on MASLD.

Conclusions: DJB ameliorates MASLD by regulating gut microbiota and BA metabolism through hepatic farnesoid X receptor pathways.

Graphical abstract

FIGURE 1

DJB ameliorates HFD-induced MAFL in rats. (A) Relative changes in body weight after DJB or sham surgery. (B) Liver weight and liver/body weight ratio at 6 weeks postoperatively. (C) Serum ALT and AST concentrations. (D) Serum TG and TC concentrations. (E) Liver TG contents. (F) Representative images of rat liver tissues stained with H&E and Oil Red O. (G–I) Serum glucose concentrations measured by (G) ipGTT, (H) oGTT, and (I) ITT. n = 6 individuals/group. Each point represented an individual rat. Data were represented as mean ± SD. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. Data were analyzed using an unpaired Student t test. Scale bar, 100 μm. Abbreviations: DJB, duodenal-jejunal bypass; H&E, hematoxylin and eosin; HFD, high-fat diet; ipGTT, intraperitoneal glucose tolerance test; ITT, insulin tolerance test; MAFL, metabolic dysfunction–associated fatty liver; oGTT, oral glucose tolerance test; TC, total cholesterol; TG, triglyceride.

FIGURE 2

Bacterial depletion abrogates the ameliorating effects of DJB on MASLD. (A) Relative changes in body weight in DJB-operated rats after 5 weeks of Abx administration. (B) Liver weight and liver/body weight ratio. (C) Serum ALT and AST concentrations. (D) Serum TG and TC concentrations. (E) Liver TG contents. (F) Representative images of rat liver tissues stained with H&E and Oil Red O. (G–I) Serum glucose concentrations measured by (G) ipGTT, (H) oGTT, and (I) ITT. n = 6 individuals/group. Each point represented an individual rat. Data were represented as mean ± SD. *p < 0.05, **p < 0.01, ***p < 0.001. Data were analyzed using an unpaired Student t test. Scale bar, 100 μm. Abbreviations: Abx, oral broad-spectrum antibiotic cocktail; DJB, duodenal-jejunal bypass; H&E, hematoxylin and eosin; ipGTT, intraperitoneal glucose tolerance test; ITT, insulin tolerance test; MASLD, metabolic dysfunction–associated steatotic liver disease; oGTT, oral glucose tolerance test; TC, total cholesterol; TG, triglyceride.

FIGURE 3

FMT from DJB-operated rats improves MASLD in HFD-fed recipients. (A) Diagram of the FMT1 experiment. (B) Relative changes in body weight after 2 weeks of daily FMT. (C) Liver weight and liver/body weight ratio of rats receiving microbiota from DJB-operated or sham-operated rats and fed an HFD for 8 weeks. (D) Serum ALT and AST concentrations. (E) Serum TG and TC concentrations. (F) Liver TG contents. (G) Representative images of rat liver tissues stained with H&E and Oil Red O. (H–J) Serum glucose concentrations measured by (H) ipGTT, (I) oGTT, and (J) ITT. n = 6 individuals/group. Each point represented an individual rat. Data were represented as mean ± SD. *p < 0.05, **p < 0.01, ***p < 0.001. Data were analyzed using an unpaired Student t test. Scale bar, 100 μm. Abbreviations: DJB, duodenal-jejunal bypass; FMT, fecal microbiota transplantation; H&E, hematoxylin and eosin; HFD, high-fat diet; ipGTT, intraperitoneal glucose tolerance test; ITT, insulin tolerance test; MASLD, metabolic dysfunction–associated steatotic liver disease; oGTT, oral glucose tolerance test; TC, total cholesterol; TG, triglyceride.

FIGURE 4

DJB reshapes the gut microbiota and reduces the abundance of Clostridium. (A) PCoA plot of 16S rRNA gene sequencing data sets based on weighted UniFrac. (B) Relative abundances at the phylum level. (C) Relative abundances of Firmicutes and Bacteroidetes. (D) The Firmicutes/Baceteroidetes ratio. (E) Relative abundances at the genus level. (F) Relative abundance of selected microbial genera that differed significantly in the DJB-operated and sham-operated groups. (G) Relative abundances of Clostridium. (H) LEfSe identification of taxa that differed significantly in abundance in the DJB-operated and sham-operated groups (LDA ≥4, p < 0.05). n = 6 individuals/group. Each point represented an individual rat. Data were represented as mean ± SD. **p < 0.01. Data were analyzed using the Mann-Whitney U test. Abbreviations: DJB, duodenal-jejunal bypass; LDA, linear discriminant analysis; LEfSe, linear discriminant analysis effect size; PCoA, principal coordinate analysis.

FIGURE 5

DJB reduces the biosynthesis of secondary BAs and activates hepatic FXR. (A) PCA plot of BA-targeted metabolomics data sets from systemic circulation (left) and cecal contents (right) of DJB-operated and sham-operated rats. (B) Total, primary, secondary, unconjugated, and conjugated BAs in systemic circulation (left) and cecal contents (right) as determined by BA-targeted metabolomics. (C) BA contents in the systemic circulation of DJB-operated and sham-operated rats. (D) Cecal BA contents in DJB-operated and sham-operated rats. (E) Serum C4 levels in DJB-operated and sham-operated rats. (F) Pearson correlation analysis of cecal DCA and LCA contents with the relative abundance of Clostridium. (G) Metagenomic analysis of the relative abundance of Bai operon genes. (H, I) Levels of mRNA expression of Fxr and its target genes in the (H) livers and (I) intestines of DJB-operated and sham-operated rats. (J) Serum FGF15 levels in DJB-operated and sham-operated rats. (K) Serum GLP-1 levels in DJB-operated and sham-operated rats. (L) Levels of mRNA expression of Fxr and its target genes in the livers of HFD-fed rats receiving microbiota from DJB-operated or sham-operated rats. n = 6 individuals/group. Each point represented an individual rat. Data were represented as mean ± SEM in (B–D) and mean ± SD in (E) and (G–K). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. Data were analyzed using an unpaired Student t test. Abbreviations: BA, bile acid; C4, 7α-hydroxy-4-cholesten-3-one; DCA, deoxycholic acid; DJB, duodenal-jejunal bypass; FMT, fecal microbiota transplantation; FXR, farnesoid X receptor; GLP-1, glucagon-like peptide-1; HFD, high-fat diet; LCA, lithocholic acid; PCA, principal component analysis.

FIGURE 6

Inhibition of FXR attenuates the ameliorative effects of post-DJB microbiota on MASLD. (A) Diagram of the experiment. (B) Relative changes in body weight of rats receiving microbiota from DJB-operated or sham-operated rats and administered the FXR inhibitor Z-Gu. (C) Liver weight and liver/body weight ratio. (D) Serum ALT and AST concentrations. (E) Serum TG and TC concentrations. (F) Liver TG contents. (G) Representative images of rat liver tissues stained with H&E and Oil Red O. (H, I) Serum glucose concentrations measured by (H) ipGTT and (I) ITT. n = 6 individuals/group. Each point represented an individual rat. Data were represented as mean ± SD. *p < 0.05, **p < 0.01, ***p < 0.001. Data were analyzed using 1-way ANOVA. Scale bar, 100 μm. Abbreviations: DJB, duodenal-jejunal bypass; FMT, fecal microbiota transplantation; FXR, farnesoid X receptor; HFD, high-fat diet; ITT, insulin tolerance test; ipGTT, intraperitoneal glucose tolerance test; MASLD, metabolic dysfunction–associated steatotic liver disease; TC, total cholesterol; TG, triglyceride.