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. 2014 May 20;111(20):7421-6.
doi: 10.1073/pnas.1323599111. Epub 2014 May 5.

Regulation of host weight gain and lipid metabolism by bacterial bile acid modification in the gut

Susan A Joyce  1 , John MacSharry  2 , Patrick G Casey  3 , Michael Kinsella  4 , Eileen F Murphy  5 , Fergus Shanahan  6 , Colin Hill  3 , Cormac G M Gahan  7
Affiliations

Regulation of host weight gain and lipid metabolism by bacterial bile acid modification in the gut

Susan A Joyce et al. Proc Natl Acad Sci U S A. .

Abstract

Alterations in the gastrointestinal microbiota have been implicated in obesity in mice and humans, but the key microbial functions influencing host energy metabolism and adiposity remain to be determined. Despite an increased understanding of the genetic content of the gastrointestinal microbiome, functional analyses of common microbial gene sets are required. We established a controlled expression system for the parallel functional analysis of microbial alleles in the murine gut. Using this approach we show that bacterial bile salt hydrolase (BSH) mediates a microbe-host dialogue that functionally regulates host lipid metabolism and plays a profound role in cholesterol metabolism and weight gain in the host. Expression of cloned BSH enzymes in the gastrointestinal tract of gnotobiotic or conventionally raised mice significantly altered plasma bile acid signatures and regulated transcription of key genes involved in lipid metabolism (Pparγ, Angptl4), cholesterol metabolism (Abcg5/8), gastrointestinal homeostasis (RegIIIγ), and circadian rhythm (Dbp, Per1/2) in the liver or small intestine. High-level expression of BSH in conventionally raised mice resulted in a significant reduction in host weight gain, plasma cholesterol, and liver triglycerides, demonstrating the overall impact of elevated BSH activity on host physiology. In addition, BSH activity in vivo varied according to BSH allele group, indicating that subtle differences in activity can have significant effects on the host. In summary, we demonstrate that bacterial BSH activity significantly impacts the systemic metabolic processes and adiposity in the host and represents a key mechanistic target for the control of obesity and hypercholesterolemia.

Keywords: FXR; Lactobacillus salivarius; adiponectin; barrier function; host response.

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Conflict of interest statement

Conflict of interest statement: S.A.J., C.H., and C.G.M.G. have filed a patent relating to a portion of the work outlined in this paper.

Figures

Fig. 1.
Fig. 1.
Expression of cloned BSH in E. coli MG1655 and activity in murine gallbladder bile in vitro. (A) Cloning strategy for expression of BSH enzymes in E. coli MG1655. (B) Ninhydrin assay showing the release of taurine from conjugated bile acids as an index of BSH activity. *P < 0.05; **P < 0.005, (n = 5 per group). (C) Heat maps summarizing UPLC-MS analysis of individual bile acids in murine bile in vitro following 90-min exposure to EC, to plasmid-borne ECpBSH1 or ECpBSH2, or to the empty vector control, ECpNZ44. Results represent analysis of three biological replicates. MBA, murine bile acids.
Fig. 2.
Fig. 2.
Alterations of host bile acid signatures through gastrointestinal expression of cloned BSH in gnotobiotic mice. (A) Total plasma bile acids (assessed by UPLC-MS) in germ-free (GF) mice, mice monocolonized with EC, ECBSH1, or ECBSH2, and conventionalized mice (CONV-D). (B) Total tauroconjugated bile acids (assessed by UPLC-MS) in plasma of GF mice, mice monocolonized with EC, ECBSH1, or ECBSH2, and CONV-D mice. (C) Total tauroconjugated muricholic acid moieties in plasma of GF mice, mice monocolonized with EC, ECBSH1, or ECBSH2, and CONV-D mice. (D) Total unconjugated muricholic acid moieties in plasma of GF mice, mice monocolonized with ESC, ECBSH1, or ECBSH2, and CONV-D mice. In all cases statistical comparisons were made relative to E. coli controls using the Student t test and were corrected for false discovery using the Benjamini–Hochberg procedure. *P < 0.05; **P < 0.005 (n = 5 per group). (E) Influence of gastrointestinal BSH expression upon Cyp7a expression in the livers of monocolonized mice. Cyp7a transcript was measured by qRT-PCR (n = 5 per group). Data are presented as means ± SEM; *P < 0.05 vs. GF; **P < 0.01 vs. GF; #P < 0.05 vs. ECBSH1; ###P < 0.001 vs. ECBSH1. (F) Total levels of secondary and tertiary bile acids in plasma of GF mice, mice monocolonized with EC, ECBSH1, or ECBSH2, and CONV-D mice. (n = 5 per group).
Fig. 3.
Fig. 3.
BSH expression in the gastrointestinal tract of gnotobiotic mice significantly alters gene-expression patterns in ileal and hepatic tissue. Microarray analysis of ileal and liver tissue from GF mice, CONV-D mice, and mice monocolonized with EC, ECBSH1, or ECBSH2. Shown are heat maps representing gene-expression profiles of selected genes that were significantly (P < 0.05) altered through BSH1 expression in our system. Pathways related to lipid digestion and absorption, circadian rhythm, adipo-signaling, and immune homeostasis were most significantly affected, as determined by pathway analysis, and are shown here (n = 5 mice per group). The schematic indicates key transcriptional changes affected by BSH1 expression. Genes increased in ECBSH1-colonized mice relative to EC-colonized mice are indicated in red, genes decreased in ECBSH1-colonized mice relative to EC-colonized mice are indicated in blue.
Fig. 4.
Fig. 4.
Gastrointestinal expression of cloned BSH in conventionally raised mice alters plasma bile acid profiles. Mice were provided with streptomycin (5 mg/mL) ad libitum in drinking water to promote stable high-level colonization of the host E. coli MG1655 StrepR strain as described previously (16). (A) Total plasma bile acids (assessed by UPLC-MS) in conventional mice (not treated, NT), conventional mice treated with antibiotic only (Ab), and mice colonized with EC, ECBSH1, or ECBSH2). (B) Total tauroconjugated plasma bile acids in uncolonized untreated or antibiotic-treated mice and mice colonized by EC, ECBSH1, or ECBSH2. (C) Relative proportions of primary bile acids (pBAs), secondary and tertiary bile acids (stBAs), and tauroconjugated bile acids (TcBAs) in the plasma of uncolonized untreated and antibiotic-treated mice and mice colonized by EC, ECBSH1, or ECBSH2. (D) Secondary and tertiary bile acid moieties in uncolonized, untreated and antibiotic-treated mice and mice colonized by EC, ECBSH1, or ECBSH2. (E) Total tauroconjugated muricholic acid moieties in plasma of conventionally raised untreated and antibiotic-treated mice and mice colonized by EC, ECBSH1, or ECBSH2. (F) Total unconjugated muricholic acid moieties in plasma of untreated mice, antibiotic-treated mice, and mice colonized by EC, ECBSH1, or ECBSH2. In all cases comparisons were made relative to E. coli controls using the Student t test and were corrected for false discovery using the Benjamini–Hochberg procedure. *P < 0.05; **P < 0.005; ***P < 0.0005 (n = 5 per group).
Fig. 5.
Fig. 5.
Gastrointestinal expression of cloned BSH in conventionally raised mice reduces weight gain, serum cholesterol, and liver triglycerides. (A) Average weight gain over time measured in grams following colonization of mice with EC or ECBSH1. Data represent antibiotic-treated mice (solid circles), EC-colonized mice (solid squares), and ECBSH1-colonized mice (solid triangles) with weight gain monitored over 10 wk. **P < 0.005 and ***P < 0.0005 for ECBSH1 mice relative to EC-colonized controls. (B) Weight of total excised fat from mice undergoing antibiotic treatment alone and mice colonized by EC, ECBSH1, or ECBSH2. *P < 0.05 relative to the EC-colonized control (n = 5 per group). (C) Levels of LDL cholesterol in plasma in mice colonized by EC, ECBSH1, or ECBSH2 and control uncolonized mice. *P < 0.05 relative to controls (n = 5 per group). (D) Levels of liver triglycerides in mice colonized by EC, ECBSH1, or ECBSH2 and in antibiotic-treated controls (Ab). *P < 0.05. (n = 5 per group). All analyses used the Student t test.
Fig. 6.
Fig. 6.
Gastrointestinal expression of BSH influences gene-expression patterns in ileal tissue in conventionally raised mice. Mice given streptomycin were colonized by E. coli MG1655 StrepR as outlined in our model system. Gene-expression patterns of selected genes were examined using qRT-PCR in ileal tissue of mice colonized by EC, ECBSH1, or ECBSH2 and of uncolonized animals (n = 5 per group). Statistical significance was determined using ANOVA. Data are presented as means ± SEM; *P < 0.05 vs. GF; $P < 0.05 vs. EC.
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