Cyp3a11 is not essential for the formation of murine bile acids

Humans and mice differ substantially in their bile acid profiles as mice in addition to cholic acid (CA) predominantly synthesize 6β-hydroxylated muricholic acids (MCAs) whereas humans produces chenodeoxycholic acid (CDCA) and CA as primary bile acids. Identifying the gene performing 6β-hydroxylation would be useful for ‘humanizing’ the bile acid profile in mice for studies of the interaction between bile acids, gut microbiota, and host metabolism. We investigated the formation of MCAs in primary murine hepatocytes and found that αMCA is synthesized from CDCA and βMCA from UDCA. It is commonly assumed that the P450-enzyme CYP3A11 catalyzes 6β-hydroxylation of bile acids, thus we hypothesized that mice without the Cyp3a11 gene would lack MCAs. To test this hypothesis, we analyzed bile acid profiles in Cyp3a deficient mice, which lack 7 genes in the Cyp3a gene cluster including Cyp3a11, and compared them with wild-type littermate controls. Bile acid composition in liver, gallbladder, caecum and serum from Cyp3a knock out mice and wild-type littermate controls was analyzed with UPLC-MS/MS and revealed no major differences in bile acid composition. We conclude that Cyp3a11 is not necessary for 6β-hydroxylation and the formation of MCAs

Regulation of Serum Amyloid A3 (SAA3) in Mouse Colonic Epithelium and Adipose Tissue by the Intestinal Microbiota

The gut microbiota has been proposed as an environmental factor that affects the development of metabolic and inflammatory diseases in mammals. Recent reports indicate that gut bacteria-derived lipopolysaccharide (LPS) can initiate obesity and insulin resistance in mice; however, the molecular interactions responsible for microbial regulation of host metabolism and mediators of inflammation have not been studied in detail. Hepatic serum amyloid A (SAA) proteins are markers and proposed mediators of inflammation that exhibit increased levels in serum of insulin-resistant mice. Adipose tissue-derived SAA3 displays monocyte chemotactic activity and may play a role in metabolic inflammation associated with obesity and insulin resistance. To investigate a potential mechanistic link between the intestinal microbiota and induction of proinflammatory host factors, we performed molecular analyses of germ-free, conventionally raised and genetically modified Myd88−/− mouse models. SAA3 expression was determined to be significantly augmented in adipose (9.9±1.9-fold; P<0.001) and colonic tissue (7.0±2.3-fold; P<0.05) by the presence of intestinal microbes. In the colon, we provided evidence that SAA3 is partially regulated through the Toll-like receptor (TLR)/MyD88/NF-kappaB signaling axis. We identified epithelial cells and macrophages as cellular sources of SAA3 in the colon and found that colonic epithelial expression of SAA3 may be part of an NF-kappaB-dependent response to LPS from gut bacteria. In vitro experiments showed that LPS treatments of both epithelial cells and macrophages induced SAA3 expression (27.1±2.5-fold vs. 1.6±0.1-fold, respectively). Our data suggest that LPS, and potentially other products of the indigenous gut microbiota, might elevate cytokine expression in tissues and thus exacerbate chronic low-grade inflammation observed in obesity

Host-microbiota interaction induces bi-phasic inflammation and glucose intolerance in mice

Objective: Gut microbiota modulates adiposity and glucose metabolism in humans and mice. Here we investigated how colonization of germ-free (GF) mice affects kinetics of adiposity and glucose metabolism. Methods: Adiposity and glucose metabolism were evaluated at different time points in ex-GF and antibiotic treated mice after colonization with gut microbiota from a conventionally raised (CONV-R) mouse. Mouse physiology, microbiome configuration, serum cytokine levels, and gene expression for inflammatory markers were performed in different tissues. Results: Colonization resulted in a bi-phasic glucose impairment: the first phase occurring within 3 days of colonization (early phase) and the second 14–28 days after colonization (delayed phase). The early phase co-occurred with an inflammatory response and was independent of adiposity, while the delayed phase was mostly ascribed to adipose tissue expansion and inflammation. Importantly, re-colonization of antibiotic treated mice displays only the delayed phase of glucose impairment and adiposity, suggesting that the early phase may be unique to colonization of the immature GF mice gut. Conclusions: Our results provide new insights on host–microbiota interaction during colonization of GF mice and the resulting effects on adiposity and glucose metabolism in a time resolved fashion

Infection Regulates Pro-Resolving Mediators that Lower Antibiotic Requirements

Underlying mechanisms for how bacterial infections contribute to active resolution of acute inflammation are unknown. Here, we performed exudate leukocyte trafficking and mediator-metabololipidomics of murine peritoneal Escherichia coli (E. coli) infections with temporal identification of pro-inflammatory (prostaglandins and leukotrienes) and specialized pro-resolving mediators (SPM). In self-resolving E. coli exudates ($10^5$ CFU), the dominant SPM identified were resolvin (Rv) D5 and protectin D1 (PD1), which at 12 h were significantly greater than levels in exudates from higher titer E. coli ($10^7$ CFU) challenged mice. Germ-free mice displayed endogenous RvD1 and PD1 levels higher than in conventional mice. RvD1 and RvD5 (ng/mouse) each reduced bacterial titers in blood and exudates, E. coli-induced hypothermia and increased survival, demonstrating the first actions of RvD5. With human polymorphonuclear neutrophils (PMN) and macrophages, RvD1, RvD5, and PD1 each directly enhanced phagocytosis of E. coli, and RvD5 counter-regulated a panel of pro-inflammatory genes, including NF-κB and TNF-α. RvD5 activated the RvD1 receptor, GPR32, to enhance phagocytosis. With self-limited E. coli infections, RvD1 and the antibiotic ciprofloxacin accelerated resolution, each shortening resolution intervals (Ri). Host-directed RvD1 actions enhanced ciprofloxacin’s therapeutic actions. In $10^7$ CFU E. coli infections, SPM (RvD1, RvD5, PD1) together with ciprofloxacin also heightened host antimicrobial responses. In skin infections, SPM enhanced vancomycin clearance of Staphylococcus aureus. These results demonstrate that specific SPM are temporally and differentially regulated during infections and that they are anti-phlogistic, enhance containment and lower antibiotic requirements for bacterial clearance

A Changing Gastric Environment Leads to Adaptation of Lipopolysaccharide Variants in Helicobacter pylori Populations during Colonization

The human gastric pathogen Helicobacter pylori colonizes the stomachs of half of the human population, and causes development of peptic ulcer disease and gastric adenocarcinoma. H. pylori-associated chronic atrophic gastritis (ChAG) with loss of the acid-producing parietal cells, is correlated with an increased risk for development of gastric adenocarinoma. The majority of H. pylori isolates produce lipopolysaccharides (LPS) decorated with human-related Lewis epitopes, which have been shown to phase-vary in response to different environmental conditions. We have characterized the adaptations of H. pylori LPS and Lewis antigen expression to varying gastric conditions; in H. pylori isolates from mice with low or high gastric pH, respectively; in 482 clinical isolates from healthy individuals and from individuals with ChAG obtained at two time points with a four-year interval between endoscopies; and finally in isolates grown at different pH in vitro. Here we show that the gastric environment can contribute to a switch in Lewis phenotype in the two experimental mouse models. The clinical isolates from different human individuals showed that intra-individual isolates varied in Lewis antigen expression although the LPS diversity was relatively stable within each individual over time. Moreover, the isolates demonstrated considerable diversity in the levels of glycosylation and in the sizes of fucosylated O-antigen chains both within and between individuals. Thus our data suggest that different LPS variants exist in the colonizing H. pylori population, which can adapt to changes in the gastric environment and provide a means to regulate the inflammatory response of the host during disease progression

The endocannabinoid system links gut microbiota to adipogenesis

We investigated several models of gut microbiota modulation: selective (prebiotics, probiotics, high-fat), drastic (antibiotics, germ-free mice) and mice bearing specific mutations of a key gene involved in the toll-like receptors (TLR) bacteria-host interaction (Myd88−/−). Here we report that gut microbiota modulates the intestinal endocannabinoid (eCB) system-tone, which in turn regulates gut permeability and plasma lipopolysaccharide (LPS) levels.The activation of the intestinal endocannabinoid system increases gut permeability which in turn enhances plasma LPS levels and inflammation in physiological and pathological conditions such as obesity and type 2 diabetes.The investigation of adipocyte differentiation and lipogenesis (both markers of adipogenesis) indicate that gut microbiota controls adipose tissue physiology through LPS-eCB system regulatory loops and may play a critical role in the adipose tissue plasticity during obesity.In vivo, ex vivo and in vitro studies indicate that LPS acts as a master switch on adipose tissue metabolism, by blocking the cannabinoid-driven adipogenesis

The gut microbiota modulates host amino acid and glutathione metabolism in mice

The gut microbiota has been proposed as an environmental factor that promotes the progression of metabolic diseases. Here, we investigated how the gut microbiota modulates the global metabolic differences in duodenum, jejunum, ileum, colon, liver, and two white adipose tissue depots obtained from conventionally raised (CONV-R) and germ-free (GF) mice using gene expression data and tissue-specific genome-scale metabolic models (GEMs). We created a generic mouse metabolic reaction (MMR) GEM, reconstructed 28 tissue-specific GEMs based on proteomics data, and manually curated GEMs for small intestine, colon, liver, and adipose tissues. We used these functional models to determine the global metabolic differences between CONV-R and GF mice. Based on gene expression data, we found that the gut microbiota affects the host amino acid (AA) metabolism, which leads to modifications in glutathione metabolism. To validate our predictions, we measured the level of AAs and N-acetylated AAs in the hepatic portal vein of CONV-R and GF mice. Finally, we simulated the metabolic differences between the small intestine of the CONV-R and GF mice accounting for the content of the diet and relative gene expression differences. Our analyses revealed that the gut microbiota influences host amino acid and glutathione metabolism in mice

A POSSIBLE ROLE OF GUT MICROBIOTA IN THE BEHAVORIAL CONTROL OF ALCOHOL-DEPENDENT SUBJECTS

These observations suggest that alterations at the level of the gut microbiota influence the gut permeability and activate specific inflammation pathways that are related to psychological symptoms of alcoholdependence. Altogether these observations are consistent with a role of inflammation as one mediator of a gut-brain communication in AD patients