Modified Bile Acids Research Articles

A Prebiotic Diet Containing Galactooligosaccharides and Polydextrose Produces Dynamic and Reproducible Changes in the Gut Microbial Ecosystem in Male Rats.

Despite substantial evidence supporting the efficacy of prebiotics for promoting host health and stress resilience, few experiments present evidence documenting the dynamic changes in microbial ecology and fecal microbially modified metabolites over time. Furthermore, the literature reports a lack of reproducible effects of prebiotics on specific bacteria and bacterial-modified metabolites. The current experiments examined whether consumption of diets enriched in prebiotics (galactooligosaccharides (GOS) and polydextrose (PDX)), compared to a control diet, would consistently impact the gut microbiome and microbially modified bile acids over time and between two research sites. Male Sprague Dawley rats were fed control or prebiotic diets for several weeks, and their gut microbiomes and metabolomes were examined using 16S rRNA gene sequencing and untargeted LC-MS/MS analysis. Dietary prebiotics altered the beta diversity, relative abundance of bacterial genera, and microbially modified bile acids over time. PICRUSt2 analyses identified four inferred functional metabolic pathways modified by the prebiotic diet. Correlational network analyses between inferred metabolic pathways and microbially modified bile acids revealed deoxycholic acid as a potential network hub. All these reported effects were consistent between the two research sites, supporting the conclusion that dietary prebiotics robustly changed the gut microbial ecosystem. Consistent with our previous work demonstrating that GOS/PDX reduces the negative impacts of stressor exposure, we propose that ingesting a diet enriched in prebiotics facilitates the development of a health-promoting gut microbial ecosystem.

Microbial metabolites as a way to provide crosstalk between gut and liver.

Bile acid metabolism is partially regulated through the activity of the gut microbiota. Primary bile acids can be deconjugated and modified by bacteria expressing bile salt hydrolase and other enzymes, changing bile acid recycling by changing the interactions between enterocytes and hepatocytes. The modified bile acids can also activate signalling in cells regulating metabolism including colonic L-cells, skeletal muscle cells and brown adipocytes. In pregnancy, both bile acid metabolism and gut microbiota composition are altered. In women with intrahepatic cholestasis of pregnancy, the changes in bile acid metabolism are exacerbated and there is some evidence that the gut microbiota composition is also altered. Here we review the crosstalk between the liver and the gut especially in women with intrahepatic cholestasis of pregnancy, with a focus on the role of the gut microbiota in this crosstalk.

24-Norursodeoxycholic acid ameliorates experimental alcohol-related liver disease and activates hepatic PPARγ

Alcohol-related liver disease (ALD) is a global healthcare challenge with limited treatment options. 24-Norursodeoxycholic acid (NorUDCA) is a synthetic bile acid with anti-inflammatory properties in experimental and human cholestatic liver diseases. In the present study, we explored the efficacy of norUDCA in experimental ALD. NorUDCA was tested in a preventive and therapeutic setting in an experimental ALD model (Lieber-DeCarli diet enriched with ethanol). Liver disease was phenotypically evaluated using histology and biochemical methods, and anti-inflammatory properties and peroxisome proliferator-activated receptor gamma activation by norUDCA were evaluated in cellular model systems. NorUDCA administration ameliorated ethanol-induced liver injury, reduced hepatocyte death, and reduced the expression of hepatic pro-inflammatory cytokines including tumour necrosis factor (Tnf), Il-1β, Il-6, and Il-10. NorUDCA shifted hepatic macrophages towards an anti-inflammatory M2 phenotype. Further, norUDCA administration altered the composition of the intestinal microbiota, specifically increasing the abundance of Roseburia, Enterobacteriaceae, and Clostridum spp. In atherapeutic model, norUDCA also ameliorated ethanol-induced liver injury. Moreover, norUDCA suppressed lipopolysaccharide-induced IL-6 expression in human peripheral blood mononuclear cells and evoked peroxisome proliferator-activated receptor gamma activation. NorUDCA ameliorated experimental ALD, protected against hepatic inflammation, and affected gut microbial commensalism. NorUDCA could serve as a novel therapeutic agent in the future management of patients with ALD. Alcohol-related liver disease is a global healthcare concern with limited treatment options. 24-Norursodeoxycholic acid (NorUDCA) is a modified bile acid, which was proven to be effective in human cholestatic liver diseases. In the present study, we found a protective effect of norUDCA in experimental alcoholic liver disease. For patients with ALD, norUDCA could be a potential new treatment option.

Analysis of gut microbiota in rats with bile duct obstruction after biliary drainage

The abundance of specific gut microorganisms is strongly associated with the concentrations of microbially modified bile acids. This study aimed to investigate the composition of intestinal microbiota in rats subjected to bile duct ligation or biliary drainage. Extrahepatic bile duct ligation was conducted to induce bile duct obstruction in rats. The bile was drained via a percutaneous biliary drainage catheter to cause bile deficiency. The total DNA extracted from fecal samples was sequenced with 16S DNA sequencing. Taxonomic classifications were conducted using the Mothur algorithm and SILVA138 database and were presented along with the abundance presented using a heatmap. The inter- and intra-group differences in the intestinal microbiome composition were analyzed by ANOSIM test. The biomarker microorganisms were screened using the Linear discriminant analysis Effect size method. The possible functional pathways were predicted using the Tax4Fun package. A total of 3277 operational taxonomic units (OTUs) were examined, with 2410 in the Kongbai group, 2236 in the Gengzu group, and 1763 in the Yinliu group. The composition of microorganisms at the levels of phylum, class, order, family, and genus was altered in rats with bile duct obstruction. This composition was then restored by biliary drainage. The top 10 predominant microorganisms were identified that led to the inter-group differences. Functional annotation revealed that the potential functions of the microorganisms with significant differences were enriched in metabolism, cellular processes, and genetic and environmental information processing. The intestinal microbial community was significantly changed in rats with bile duct obstruction. The changes in the abundance of intestinal microbiota Prevotellaceae and Enterobacteriaceae were statistically significant after biliary drainage treatment.

Obeticholic Acid—A Pharmacological and Clinical Review

Obeticholic acid (OCA) or 6-alpha-ethyl-chenodeoxycholic acid is a semisynthetic modified bile acid derivative that acts on the farnesoid X receptor (FXR) as an agonist with a higher potency than bile acid. The FXR is a nuclear receptor highly expressed in the liver and small intestine and regulates bile acid, cholesterol, glucose metabolism, inflammation, and apoptosis. The FXR group of bile acid receptors is currently under investigation for their potential role in the treatment of primary biliary cirrhosis (PBC), non-alcoholic steatohepatitis (NASH), and primary sclerosing cholangitis (PSC). Recent clinical studies suggest OCA may work synergistically with lipid modifying medications to further improve long-term outcomes with primary sclerosing cholangitis. Specifically, OCA can improve clinical outcomes in NASH patients with their different histological, metabolic, and biochemical issues as well as improve morbidity and mortality in patients suffering from PBC, PSC, or liver disease. This improvement is noted in both improved histological examination and reduced need for transplantation. In this review, we examine the pharmacology of OCA towards the treatment of PBC refractory and steatohepatitis (NASH). In addition, we examine future directions and applications of OCA for PBC, PSC, NASH, and NAFLD.

Synthesis of modified bile acids via palladium-catalyzed C(sp3)-H (hetero)arylation.

A Pd(II)-catalyzed strategy for the diastereo- and regioselective (hetero)arylation of unactivated C(sp3)-H bonds in bile acids is accomplished with aryl and heteroaryl iodides under solvent-free conditions using the 8-aminoquinoline auxiliary as a directing group. This methodology demonstrated excellent functional group tolerance with respect to aryl/heteroaryl iodides on O-protected N-(quinolin-8-yl)cholyl/deoxycholyl amides to afford β-C(sp3)-H (hetero)arylated products in good-to-excellent yields. Moreover, the 8-aminoquinoline (AQ) auxiliary can easily be removed to obtain modified bile acids.

Secretin alleviates biliary and liver injury during late-stage primary biliary cholangitis via restoration of secretory processes.

Primary biliary cholangitis (PBC) is characterised by ductopenia, ductular reaction, impairment of anion exchanger 2 (AE2) and the 'bicarbonate umbrella'. Ductulo-canalicular junction (DCJ) derangement is hypothesised to promote PBC progression. The secretin (Sct)/secretin receptor (SR) axis regulates cystic fibrosis transmembrane receptor (CFTR) and AE2, thus promoting choleresis. We evaluated the role of Sct/SR signalling on biliary secretory processes and subsequent injury in a late-stage PBC mouse model and human samples. At 32 weeks of age, female and male wild-type and dominant-negative transforming growth factor beta receptor II (late-stage PBC model) mice were treated with Sct for 1 or 8 weeks. Bulk RNA-sequencing was performed in isolated cholangiocytes from mouse models. Biliary Sct/SR/CFTR/AE2 expression and bile bicarbonate levels were reduced in late-stage PBC mouse models and human samples. Sct treatment decreased bile duct loss, ductular reaction, inflammation, and fibrosis in late-stage PBC models. Sct reduced hepatic bile acid levels, modified bile acid composition, and restored the DCJ and 'bicarbonate umbrella'. RNA-sequencing identified that Sct promoted mature epithelial marker expression, specifically anterior grade protein 2 (Agr2). Late-stage PBC models and human samples exhibited reduced biliary mucin 1 levels, which were enhanced by Sct treatment. Loss of Sct/SR signalling in late-stage PBC results in a faulty 'bicarbonate umbrella' and reduced Agr2-mediated mucin production. Sct restores cholangiocyte secretory processes and DCJ formation through enhanced mature cholangiocyte phenotypes and bile duct growth. Sct treatment may be beneficial for individuals with late-stage PBC. Secretin (Sct) regulates biliary proliferation and bicarbonate secretion in cholangiocytes via its receptor, SR, and in mouse models and human samples of late-stage primary biliary cholangitis (PBC), the Sct/SR axis is blunted along with loss of the protective 'bicarbonate umbrella'. We found that both short- and long-term Sct treatment ameliorated ductular reaction, immune cell influx, and liver fibrosis in late-stage PBC mouse models. Importantly, Sct treatment promoted bicarbonate and mucin secretion and hepatic bile acid efflux, thus reducing cholestatic and toxic bile acid-associated injury in late-stage PBC mouse models. Our work perpetuates the hypothesis that PBC pathogenesis hinges on secretory defects, and restoration of secretory processes that promote the 'bicarbonate umbrella' may be important for amelioration of PBC-associated damage.

Synthesis of Novel C/D Ring Modified Bile Acids.

Bile acid receptors have been identified as important targets for the development of new therapeutics to treat various metabolic and inflammatory diseases. The synthesis of new bile acid analogues can help elucidate structure–activity relationships and define compounds that activate these receptors selectively. Towards this, access to large quantities of a chenodeoxycholic acid derivative bearing a C-12 methyl and a C-13 to C-14 double bond provided an interesting scaffold to investigate the chemical manipulation of the C/D ring junction in bile acids. The reactivity of this alkene substrate with various zinc carbenoid species showed that those generated using the Furukawa methodology achieved selective α-cyclopropanation, whereas those generated using the Shi methodology reacted in an unexpected manner giving rise to a rearranged skeleton whereby the C ring has undergone contraction to form a novel spiro–furan ring system. Further derivatization of the cyclopropanated steroid included O-7 oxidation and epimerization to afford new bile acid derivatives for biological evaluation.

Analysis of fecal bile acids and metabolites by high resolution mass spectrometry in farm animals and correlation with microbiota

There is a growing interest in the named “acidic sterolbiome” and in the genetic potential of the gut microbiome (GM) to modify bile acid (BA) structure. Indeed, the qualitative composition of BAs in feces correlates with the bowel microorganisms and their collective genetic material. GM is responsible for the production of BA metabolites, such as secondary and oxo-BAs. The specific BA profiles, as microbiome-host co-metabolic products, could be useful to investigate the GM-host interaction in animals under physiological conditions, as well as in specific diseases. In this context, we developed and validated an ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry method for the simultaneous analysis of up to 21 oxo-BAs and their 9 metabolic precursors. Chromatographic separation was achieved in 7 min with adequate analytical performance in terms of selectivity, sensitivity (LOQ from 0.05 to 0.1 µg/mL), accuracy (bias% < 5%), precision (CV% < 5%) and matrix effect (ME% < 10%). A fast solvent extraction protocol has been fine-tuned, achieving recoveries > 90%. In parallel, the gut microbiota assessment in farming animals was evaluated by 16S rRNA next-generation sequencing, and the correlation with the BA composition was performed by multivariate analysis, allowing to reconstruct species-specific associations between the BA profile and specific GM components.

Aberrant Gut-To-Brain Signaling in Irritable Bowel Syndrome - The Role of Bile Acids

Functional bowel disorders such as irritable bowel syndrome (IBS) are common, multifactorial and have a major impact on the quality of life of individuals diagnosed with the condition. Heterogeneity in symptom manifestation, which includes changes in bowel habit and visceral pain sensitivity, are an indication of the complexity of the underlying pathophysiology. It is accepted that dysfunctional gut-brain communication, which incorporates efferent and afferent branches of the peripheral nervous system, circulating endocrine hormones and local paracrine and neurocrine factors, such as host and microbially-derived signaling molecules, underpins symptom manifestation. This review will focus on the potential role of hepatic bile acids in modulating gut-to-brain signaling in IBS patients. Bile acids are amphipathic molecules synthesized in the liver, which facilitate digestion and absorption of dietary lipids. They are also important bioactive signaling molecules however, binding to bile acid receptors which are expressed on many different cell types. Bile acids have potent anti-microbial actions and thereby shape intestinal bacterial profiles. In turn, bacteria with bile salt hydrolase activity initiate the critical first step in transforming primary bile acids into secondary bile acids. Individuals with IBS are reported to have altered microbial profiles and modified bile acid pools. We have assessed the evidence to support a role for bile acids in the pathophysiology underlying the manifestation of IBS symptoms.

Aryl Hydrocarbon Receptor (AhR) Activation by 2,3,7,8-Tetrachlorodibenzo-p-Dioxin (TCDD) Dose-Dependently Shifts the Gut Microbiome Consistent with the Progression of Non-Alcoholic Fatty Liver Disease.

Gut dysbiosis with disrupted enterohepatic bile acid metabolism is commonly associated with non-alcoholic fatty liver disease (NAFLD) and recapitulated in a NAFLD-phenotype elicited by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in mice. TCDD induces hepatic fat accumulation and increases levels of secondary bile acids, including taurolithocholic acid and deoxycholic acid (microbial modified bile acids involved in host bile acid regulation signaling pathways). To investigate the effects of TCDD on the gut microbiota, the cecum contents of male C57BL/6 mice orally gavaged with sesame oil vehicle or 0.3, 3, or 30 µg/kg TCDD were examined using shotgun metagenomic sequencing. Taxonomic analysis identified dose-dependent increases in Lactobacillus species (i.e., Lactobacillus reuteri). Increased species were also associated with dose-dependent increases in bile salt hydrolase sequences, responsible for deconjugation reactions in secondary bile acid metabolism. Increased L. reuteri levels were further associated with mevalonate-dependent isopentenyl diphosphate (IPP) biosynthesis and o-succinylbenzoate synthase, a menaquinone biosynthesis associated gene. Analysis of the gut microbiomes from cirrhosis patients identified an increased abundance of genes from the mevalonate-dependent IPP biosynthesis as well as several other menaquinone biosynthesis genes, including o-succinylbenzoate synthase. These results extend the association of lactobacilli with the AhR/intestinal axis in NAFLD progression and highlight the similarities between TCDD-elicited phenotypes in mice to human NAFLD.

The relevance of DHA with modulating of host-gut microbiome signatures alterations and repairing of lipids metabolism shifts

Huge of previous reports recommended that gut microbiome have a crucial role in the human health and its change was profound impact for the metabolic improvements associated with lipids metabolism. In order to explore the relevance of a direct dysbiosis effect of gut microbiome on lipids metabolism shifts and repaired position of DHA, we built the animal model for the study with gut microbiome dysbiosis administrated by i.g. with CRO and intervened by DHA in the present work. Gut microbiome was analyzed by high throughput sequencing and bioinformatics analyses of bacteria. The composition of fatty acids and short chain fatty acids (SCFAs) were determined by gas chromatography. Blood lipids and bile acids were assayed by kit and UPLC-MS/MS, respectively. The expressions of enzymes of long chain fatty acid metabolism were analyzed by qRT-PCR. The results showed that gut microbiome dysbiosis caused lipid metabolism abnormal, and DHA was able to repair the lipids metabolism shifts resulted from gut microbiome dysbiosis. DHA could modulate host-gut microbiome signatures, improve concentrations of SCFAs, regulate fatty acids metabolism but modify bile acid profiles. In conclusion, we considered that DHA repaired lipid metabolism by modulating gut microbiome and regulating fatty acids metabolism pathway.

Modified bile acids and androstanes—Novel promising inhibitors of human cytochrome P450 17A1

Cytochromes P450 are key enzymes for steroid hormone biosynthesis in human body. They are considered as targets for the screening of novel high efficient drugs. The results of screening of bile acids and androstane derivatives toward human recombinant steroid 17α-hydroxylase/17,20-lyase (CYP17A1) are presented in this paper. A group of steroids, binding with micromolar or submicromolar affinity (in a range from 9 μM – less than 0.1 μM), was identified. Results presented here showed that these steroidal compounds are able to decrease rate of hydroxylation of essential CYP17A1 substrate – progesterone, while some compounds completely inhibited enzyme activity. Structure-activity relationship (SAR) analysis based on in vitro and in silico studies showed that high affinity of the enzyme to bile acids derivatives is correlated with side chain hydrophobicity and presence of hydroxyl or keto group at C3 position. From the other side, bile acid-derived compounds with more polar side chain or substituents at C7 and C12 positions possess higher Kd values. Among androstane-derived steroids couple of Δ5-steroids with hydroxyl group at C3 position, as well as 16,17-secosteroids, were found to be high affinity ligands of this enzyme. The data obtained could be useful for the design of novel highly efficient inhibitors of CYP17A1, since the bile acids-derived compounds are for first time recognized as effective CYP17A1 inhibitors.

Infection with Bacteroides Phage BV01 Alters the Host Transcriptome and Bile Acid Metabolism in a Common Human Gut Microbe

SUMMARYGut-associated phages are hypothesized to alter the abundance and activity of their bacterial hosts, contributing to human health and disease. Although temperate phages constitute a significant fraction of the gut virome, the effects of lysogenic infection are underexplored. We report that the temperate phage, Bacteroides phage BV01, broadly alters its host’s transcriptome, the prominent human gut symbiont Bacteroides vulgatus. This alteration occurs through phage-induced repression of a tryptophan-rich sensory protein (TspO) and represses bile acid deconjugation. Because microbially modified bile acids are important signals for the mammalian host, this is a mechanism by which a phage may influence mammalian phenotypes. Furthermore, BV01 and its relatives in the proposed phage family Salyersviridae are ubiquitous in human gut metagenomes, infecting a broad range of Bacteroides hosts. These results demonstrate the complexity of phage-bacteria-mammal relationships and emphasize a need to better understand the role of temperate phages in the gut microbiome.

Metabolite-Sensing G Protein Coupled Receptor TGR5 Protects Host From Viral Infection Through Amplifying Type I Interferon Responses.

The metabolite-sensing G protein–coupled receptors (GPCRs) bind to various metabolites and transmit signals that are important for proper immune and metabolic functions. However, the roles of metabolite-sensing GPCRs in viral infection are not well characterized. Here, we identified metabolite-sensing GPCR TGR5 as an interferon (IFN)-stimulated gene (ISG) which had increased expression following viral infection or IFN-β stimulation in a STAT1-dependent manner. Most importantly, overexpression of TGR5 or treatment with the modified bile acid INT-777 broadly protected host cells from vesicular stomatitis virus (VSV), newcastle disease virus (NDV) and herpes simplex virus type 1 (HSV-1) infection. Furthermore, VSV and HSV-1 replication was increased significantly in Tgr5-deficient macrophages and the VSV distribution in liver, spleen and lungs was increased in Tgr5-deficient mice during VSV infection. Accordingly, Tgr5-deficient mice were much more susceptible to VSV infection than wild-type mice. Mechanistically, TGR5 facilitates type I interferon (IFN-I) production through the AKT/IRF3-signaling pathway, which is crucial in promoting antiviral innate immunity. Taken together, our data reveal a positive feedback loop regulating IRF3 signaling and suggest a potential therapeutic role for metabolite-sensing GPCRs in controlling viral diseases.

Gut microbiome–mediated bile acid metabolism regulates liver cancer via NKT cells

Primary liver tumors and liver metastasis currently represent the leading cause of cancer-related death. Commensal bacteria are important regulators of antitumor immunity, and although the liver is exposed to gut bacteria, their role in antitumor surveillance of liver tumors is poorly understood. We found that altering commensal gut bacteria in mice induced a liver-selective antitumor effect, with an increase of hepatic CXCR6+ natural killer T (NKT) cells and heightened interferon-γ production upon antigen stimulation. In vivo functional studies showed that NKT cells mediated liver-selective tumor inhibition. NKT cell accumulation was regulated by CXCL16 expression of liver sinusoidal endothelial cells, which was controlled by gut microbiome-mediated primary-to-secondary bile acid conversion. Our study suggests a link between gut bacteria-controlled bile acid metabolism and liver antitumor immunosurveillance.

Bile acids and liver cancer

Microbiome Liver cancer is a leading cause of cancer-related deaths in the United States. The composition of the gut microbiome influences many human diseases, including liver inflammatory disorders. Ma et al. found that commensal gut bacteria can recruit the immune system to control the growth of liver tumors in mice (see the Perspective by Hartmann and Kronenberg). Clostridium species modified bile acids to signal liver sinusoidal endothelial cells to produce the chemokine CXCL16. This recruited natural killer T (NKT) immune cells to perform antitumor surveillance of the liver. Growth of both primary and metastatic cancer was reduced by NKT cell–driven killing. Science , this issue p. [eaan5931][1]; see also p. [858][2] [1]: /lookup/doi/10.1126/science.aan5931 [2]: /lookup/doi/10.1126/science.aat8289

Crystalline arrays of side chain modified bile acids derivatives. Two novel self-assemblies based on π-π and belly-to-belly interactions

Crystalline derivatives of side chain modified bile acids were efficiently prepared from the naturally occurring steroids by palladium-catalyzed cross coupling reaction as a key step. The solvent-free crystalline bile acids derivatives 2b–e are readily accessed by slow evaporation from selected solvents. A variety of steroidal scaffolds were found and elucidated by SXRD studies. The crystal packing of the title compounds are dominated by hydrogen-bonding interactions established between differently positioned acetyl protecting groups, which in the case of 2b and 2e take advantage of the facial amphiphilicity producing two novel steroidal supramolecular self-assemblies combining π-π and strong facial interactions. Thus, these crystalline arrays of side chain modified bile acids represent promising scaffolds for research and implementation in biomolecular materials or inclusion phenomena.

Oral treatment with Eubacterium hallii improves insulin sensitivity in db/db mice.

An altered intestinal microbiota composition is associated with insulin resistance and type 2 diabetes mellitus. We previously identified increased intestinal levels of Eubacterium hallii, an anaerobic bacterium belonging to the butyrate-producing Lachnospiraceae family, in metabolic syndrome subjects who received a faecal transplant from a lean donor. To further assess the effects of E. hallii on insulin sensitivity, we orally treated obese and diabetic db/db mice with alive E. hallii and glycerol or heat-inactive E. hallii as control. Insulin tolerance tests and hyperinsulinemic-euglycemic clamp experiments revealed that alive E. hallii treatment improved insulin sensitivity compared control treatment. In addition, E. hallii treatment increased energy expenditure in db/db mice. Active E. hallii treatment was found to increase faecal butyrate concentrations and to modify bile acid metabolism compared with heat-inactivated controls. Our data suggest that E. hallii administration potentially alters the function of the intestinal microbiome and that microbial metabolites may contribute to the improved metabolic phenotype.

Application of palladium-catalyzed carboxyl anhydride-boronic acid cross coupling in the synthesis of novel bile acids analogs with modified side chains

Palladium-catalyzed cross coupling of 4-methoxycarbonyl phenyboronic acid with acetylated bile acids in which the carboxyl functions was activated by formation of a mixed anhydride with pivalic anhydride afforded the cross coupled compounds, which were converted in novel side chain modified bile acids by one pot carbonyl reduction/removal of the protecting acetyl groups by Wolff–Kishner reduction. Unambiguous assignments of the NMR signals and crystal characterization of the heretofore unknown compounds are provided.