An elevated deoxycholic acid level induced by high-fat feeding damages intestinal stem cells by reducing the ileal IL-22.

Long-term high-fat diet (HFD) destroys the intestinal mucosal barrier by damaging intestinal stem cells (ISCs). A HFD can increase the concentration of intestinal deoxycholic acid (DCA) and decrease the secretion of interleukin-22 (IL-22), which plays an important role in the proliferation, repair and regeneration of ISCs. We hypothesized that increased level of intestinal DCA induced by a HFD leads to ISC dysfunction by reducing the IL-22 levels in intestinal tissues. In this study, 2 weeks of a DCA diet or a HFD damaged ileal ISC and its proliferation and differentiation, resulting in a decrease in Paneth cells and goblet cells. Importantly, 2 weeks of a DCA diet or a HFD also reduced ileal IL-22 concentration, accompanied by a decreased number of group 3 innate lymphoid cells in ileal mucosa, which produce IL-22 after intestinal injury. Concurrent feeding with bile acid binder cholestyramine prevented all these changes induced by a HFD. In addition, in vitro study further confirmed that exogenous IL-22 reversed the decline in the proliferation and differentiation of ileal ISCs induced by DCA stimulation. Collectively, these results revealed that the decrease in intestinal IL-22 induced by DCA may be a novel mechanism by which HFD damages ISCs. The administration of IL-22 or a bile acid binder may provide novel therapeutic targets for the metabolic syndrome caused by a HFD.

Decrease in major secondary bile acid, hyodeoxycholic acid, was the main alteration in hepatic bile acid compositions in a hypertensive nonalcoholic fatty liver disease model.

BACKGROUND: Previous findings on hepatic bile acid compositions in nonalcoholic fatty liver disease (NAFLD) have been inconsistent and complicated. The aim of this study was to investigate the effects of steatosis on hepatic bile acid composition in a hypertensive NAFLD model without obesity and diabetes mellitus and compare hepatic bile acid composition between hypertensive rats with and without steatosis. METHODS: Two groups of hypertensive rats were studied: spontaneously hypertensive rats (SHR) fed with a normal diet (SHR-N) or a choline-deficient diet (SHR-CD). Two groups of normotensive rats were studied: Wistar Kyoto rats (WKY) fed a normal diet (WKY-N) or a choline-deficient diet (WKY-CD). Hepatic bile acid analysis was performed using liquid chromatography-electrospray ionization-tandem mass spectrometry. RESULTS: Regarding bile acid composition, the hyodeoxycholic acid (HDCA) species in the SHR-CD group showed the largest change in bile acid composition, significantly decreasing to 21.9% of that found in the SHR-N group. In the WKY-CD group, no reduction of HDCA species was observed. CONCLUSIONS: We demonstrated that the decrease in HDCA species was the main alteration in a hypertensive NAFLD model. It was suggested that the decrease in HDCA species in the SHR-CD group was caused by dysbiosis.

Bioproduction of bile acids and the glycine conjugates by Penicillium fungus.

It has been found that the Penicillium endophytic filamentous fungus with the young stems of Scurrula atroprupurea (Loranthaceae) produces cholic acid, deoxycholic acid and the glycine conjugates.

Biosynthesis of bile acids in a variety of marine bacterial taxa.

Several marine bacterial strains, which were isolated from seawater off the island Dokdo, Korea, were screened to find new bioactive compounds such as antibiotics. Among them, Donghaeana dokdonensis strain DSW-6 was found to produce antibacterial agents, and the agents were then purified and analyzed by LC-MS/MS and 1D- and 2D-NMR spectrometries. The bioactive compounds were successfully identified as cholic acid and glycine-conjugated glycocholic acid, the 7alpha-dehydroxylated derivatives (deoxycholic acid and glycodeoxycholic acid) of which were also detected in relatively small amounts. Other masine isolates, taxonomically different from DSW-6, were also able to produce the compounds in a quite different production ratio from DSW-6. As far as we are aware of, these bile acids are produced by specific members of the genus Streptomyces and Myroides, and thought to be general secondary metabolites produced by a variety of bacterial taxa that are widely distributed in the sea.

Deoxycholic acid formation in gnotobiotic mice associated with human intestinal bacteria.

In humans and animals, intestinal flora is indispensable for bile acid transformation. The goal of our study was to establish gnotobiotic mice with intestinal bacteria of human origin in order to examine the role of intestinal bacteria in the transformation of bile acids in vivo using the technique of gnotobiology. Eight strains of bile acid-deconjugating bacteria were isolated from ex-germ-free mice inoculated with a human fecal dilution of 10(-6), and five strains of 7alpha-dehydroxylating bacteria were isolated from the intestine of limited human flora mice inoculated only with clostridia. The results of biochemical tests and 16S rDNA sequence analysis showed that seven out of eight bile acid-deconjugating strains belong to a bacteroides cluster (Bacteroides vulgatus, B. distasonis, and B. uniformis), and one strain had high similarity with Bilophila wadsworthia. All five strains that converted cholic acid to deoxycholic acid had greatest similarity with Clostridium hylemonae. A combination of 10 isolated strains converted taurocholic acid into deoxycholic acid both in vitro and in the mouse intestine. These results indicate that the predominant bacteria, mainly Bacteroides, in human feces comprise one of the main bacterial groups for the deconjugation of bile acids, and clostridia may play an important role in 7aplha-dehydroxylation of free-form primary bile acids in the intestine although these strains are not predominant. The gnotobiotic mouse with bacteria of human origin could be a useful model in studies of bile acid metabolism by human intestinal bacteria in vivo.

Octreotide induced prolongation of colonic transit increases faecal anaerobic bacteria, bile acid metabolising enzymes, and serum deoxycholic acid in patients with acromegaly.

BACKGROUND: Acromegalic patients have slow colonic transit, increased rates of deoxycholic acid formation, and an increased prevalence of cholesterol gall stones, especially during long term octreotide treatment. However, the effects of this prolonged large bowel transit time on the numbers of faecal anaerobes and the activities of the enzyme systems which biotransform conjugated cholic acid into unconjugated deoxycholic acid (cholylglycine hydrolase and 7alpha-dehydroxylase) are unknown. METHODS: Therefore, in 10 non-acromegalic controls, 11 acromegalic patients not treated with octreotide, and 11 acromegalics on long term (8-48 months) octreotide (100-200 mug three times daily subcutaneously), we measured large bowel transit time and, in freshly voided faeces, the activities of the two bile acid metabolising enzymes, and related the results to the proportion of deoxycholic acid in fasting serum. Moreover, in patients with acromegaly, we measured quantitative bacteriology in faeces. RESULTS: Mean large bowel transit time in acromegalics not treated with octreotide (35 (SEM 6.5) hours) was 66% longer than that in non-acromegalic controls (21 (3.1) hours; NS) and became further prolonged during octreotide treatment (48 (6.6) hours; p<0.001). These octreotide induced changes in transit were associated, in acromegalic patients, with more total (15.0 (2.5) v 6.3 (1.3)x10(9) colony forming units (cfu)/g; p<0.05) and Gram positive (6.3 (2.3) v 3.2 (1.0)x10(9) cfu/g; p<0.05) faecal anaerobes. Mean faecal cholylglycine hydrolase activity in the long term octreotide group (22.0 (6.0)x10(-2) U/mg protein) was 138% greater than that in non-acromegalic controls (12.0 (6.0)x10(-2); p<0.01). Similarly, mean 7alpha-dehydroxylase activity in octreotide treated acromegalics (11.1 (1.18)x10(-4) U/mg protein) was 78% greater than that in patients not receiving long term octreotide (6.3 (0.5)x10(-4); p<0.001). The mean proportion of deoxycholic acid in fasting serum also increased from 18.0 (2.88)% in the untreated group to 29.6 (2.3)% during long term octreotide (p<0.05). There were significant linear relationships between large bowel transit time and: (i) faecal 7alpha-dehydroxylase activity; and (ii) the proportion of deoxycholic acid in fasting serum and between 7alpha-dehydroxylase activity and the proportion of deoxycholic acid in serum.Summary/interpretation: These data suggest that increased deoxycholic acid formation seen in acromegalics during octreotide treatment is due not only to the greater numbers of faecal anaerobes but also to increased activity of the rate limiting enzyme pathway (7alpha-dehydroxylation) converting cholic acid to deoxycholic acid.

Bile acids are new products of a marine bacterium, Myroides sp. strain SM1.

Strain SM1 was isolated as a biosurfactant-producing microorganism from seawater and presumptively identified as Myroides sp., based on morphology, biochemical characteristics and 16S rDNA sequence. The strain produced surface-active compounds in marine broth, which were purified, using emulsification activity for n-hexadecane as an indicator. The purified compounds were identified by thin-layer chromatography, (1)H- and (13)C-NMR spectra and fast atom bombardment mass spectrometry as cholic acid, deoxycholic acid and their glycine conjugates. Type strains of the genus Myroides, M. odoratus JCM7458 and M. odoramitimus JCM7460, also produced these compounds. Myroides sp. strain SM1 possessed a biosynthetic route to cholic acid from cholesterol. Thus, bile acids were found as new products of prokaryotic cells, genus Myroides.

Mechanism for the transit-induced increase in colonic deoxycholic acid formation in cholesterol cholelithiasis.

BACKGROUND & AIMS: Many patients with cholesterol gallbladder stones (GBS) have a high percentage of deoxycholic acid (DCA) in gallbladder bile (all of which are in the conjugated form), probably as a result of prolonged large bowel transit times (LBTT). However, whether the prolonged LBTT increases DCA formation, solubilization, or absorption (or all 3) is not known. METHODS: In 40 subjects (20 with GBS; age range, 24-74 years), we measured LBTT using radiopaque markers, and intestinal luminal pH by radiotelemetry. We also measured quantitative anaerobic bacteriology and the activities of 2 bile acid-metabolizing enzymes in fresh cecal aspirates obtained during clinically indicated unprepared colonoscopy, and related these results to the percentage of DCA in fasting serum measured by gas chromatography-mass spectrometry. RESULTS: Compared with controls, GBS patients had longer LBTT (mean 23.1 +/- SEM 2.8 h vs. 36.5 +/- 3.3 h; P < 0.01); more total (2.7 +/- 0.6 x 10(9) vs. 5.9 +/- 1.5 x 10(9) cfu/mL) and Gram-positive (9.5 +/- 3.1 x 10(8) vs. 18.0 +/- 4.1 x 10(8) cfu/mL; P < 0.05) anaerobes; and greater 7alpha-dehydroxylating (7alpha-DH) activity (3.39 +/- 0.59 vs. 10.37 +/- 1.15 x 10(-4) U/mg protein) in the cecal aspirates. They also had higher intracolonic pH values (P < 0.02) and increased percentages of DCA in fasting serum (13.4% +/- 1.52% vs. 21.8% +/- 2. 19%; P < 0.005). Results of univariate and multivariate analyses confirmed that LBTT was critical in determining the percentage of DCA in serum and showed that 7alpha-DH activity and apparent distal colonic pH were also significant independent variables. CONCLUSIONS: Slow colonic transit (more time), increased Gram-positive anaerobes (more bacteria), and greater 7alpha-DH activity (more enzyme) favor enhanced DCA formation; transit-induced increases in distal colonic luminal pH favor enhanced DCA solubilization/bioavailability; and increases in LBTT (more time) again favor DCA absorption.

Dietary psyllium hydrocolloid and pectin increase bile acid pool size and change bile acid composition in rats.

Bile acid composition, synthetic rate and pool size were determined in rats fed diets containing 5 g cellulose, 5 g pectin or 5 g psyllium hydrocolloid/100 g diet using the washout technique. Bile acid pool sizes were similar for pectin- and psyllium-fed rats, and both were higher than the pool size for rats fed cellulose (5.48 +/- 1.22, 4.70 +/- 0.71 and 1.77 +/- 0.41 mumol/100 g body wt, respectively). Bile acid secretion rates showed a similar pattern [1730 +/- 496, 1551 +/- 252 and 572 +/- 88 nmol/(h.100 g body wt)], as did basal synthetic rates [129 +/- 25, 126 +/- 42 and 87 +/- 18 pmol/(h.100 g body wt)]. Individual and total bile acid pool sizes were generally higher in animals fed the pectin- or psyllium-supplemented diet compared with rats fed cellulose. Pectin or psyllium consumption resulted in a lower hydrophobicity of the bile acid pool and a lower ratio of circulating 12 alpha-hydroxylated to non-12 alpha-hydroxylated bile acids compared with cellulose consumption. This reduced hydrophobicity has been shown to reduce feedback inhibition of bile acid synthesis and may be responsible for the larger bile acid pool size. Changes in the location of reabsorption of bile acids may also be responsible for changes in the pool size and composition. These changes are characteristic of greater sterol excretion.

Effects of cholecystectomy on the kinetics of primary and secondary bile acids.

Removal of the gallbladder is thought to increase formation and pool size of secondary bile acids, mainly deoxycholic acid (DCA), by increased exposure of primary bile acids (cholic acid [CA], chenodeoxycholic acid [CDCA]) to bacterial dehydroxylation in the intestine. We have tested this hypothesis by simultaneous determination of pool size and turnover of DCA, CA, and CDCA in nine women before and at various intervals after removal of a functioning gallbladder. An isotope dilution technique using marker bile acids labeled with stable isotopes (2H4-DCA, 13C-CA, 13C-CDCA) was used. After cholecystectomy, concentration and output of bile acids relative to bilirubin increased (P less than 0.02) in fasting duodenal bile and cholesterol saturation decreased by 27% (P less than 0.05) consistent with enhanced enterohepatic cycling of bile acids. Three months after removal of the gallbladder bile acid kinetics were in a new steady state: pool size and turnover of CDCA were unchanged. Synthesis of CA, the precursor of DCA, was diminished by 37% (P = 0.05), probably resulting from feedback inhibition by continuous transhepatic flux of bile acids. The fraction of CA transferred after 7 alpha-dehydroxylation to the DCA pool increased from 46 +/- 16 to 66 +/- 32% (P less than 0.05). However, this enhanced transfer did not lead to increased input or size of the DCA pool, because synthesis of the precursor CA had decreased.

Total parenteral nutrition and the function of the intestinal microflora in Crohn's disease.

The effect of total parenteral nutrition (TPN) on the following six intestinal microflora-associated characteristics (MACs) was studied in patients with Crohn's disease: faecal tryptic activity (FTA), formation of coprostanol, urobilinogen, and deoxycholic acid, and degradation of mucin and beta-aspartylglycine. The FTA showed high levels before TPN, in accordance with previous findings, and decreased during TPN. Formation of coprostanol, urobilinogen, and deoxycholic acid was reduced in some patients, whereas no changes were found in the mucin and beta-aspartylglycine degradation.

Changes in bile salt composition after cholecystectomy and ileal resection.

The effect of ileal resection and cholecystectomy on bile salt metabolism was studied in female prairie dogs 4 weeks after either a sham laparotomy, cholecystectomy, ileal resection, or cholecystectomy and ileal resection. Bile was collected from a common bile duct cannula at hourly intervals for 12 hours. Pool sizes and synthetic rates of primary and secondary bile salts were determined from washout curves. Cholate, chenodeoxycholate, deoxycholate, and lithocholate levels were determined by gas chromatography from pooled collections of bile. After cholecystectomy and ileal resection, the pool sizes of primary and secondary bile salts were significantly reduced to amounts that were much less than the pool sizes after either procedure alone. Primary bile salt synthesis was significantly increased after combined cholecystectomy and ileal resection, to the same degree as cholecystectomy alone. After the combined procedures, there was a decrease in the proportion of cholate in hepatic bile associated with an increase in chenodeoxycholate, deoxycholate, and lithocholate levels. The data suggest that after the loss of both ileum and gallbladder the bile salt pool sizes are drastically reduced, the synthesis of primary bile salts is increased, and the proportion of secondary bile salts is increased. Cholecystectomy should be avoided, if possible, in patients with ileal resection in order to conserve the bile salt pool and prevent severe fat malabsorption.

The economy of the enterohepatic circulation of bile acids in the baboon. 2. Regulation of bile acid synthesis by enterohepatic circulation of bile acids.

Isotope dilution within bile acid pools and radiochemical assessment of cholesterol oxidation to bile acids were methods used to measure short-term feedback regulation of bile acid synthesis in baboons with controlled enterohepatic circulations. Intraduodenal infusion of labeled endogenous bile acid pools into bile acid-depleted animals with enhanced bile acid synthesis showed that the rate of bile acid returned to the liver affected the degree of inhibition of bile acid synthesis. Infusion of prepared bile acid pools of varying composition resulted in a specific pattern of feedback inhibition of bile salt synthesis related to pool composition and mass. Individual bile salts inhibited their own synthesis more than that of other bile salts, and chenodeoxycholic and deoxycholic acids were found to have greater inhibitory effects than cholic acid. Glycine-conjugated cholic and chenodeoxycholic acids had greater inhibitory effects than did the respective free bile salts. Infusion of mixed bile acid pools showed that dihydroxy bile acids (chenodeoxycholic or deoxycholic) enhanced feedback inhibition of cholic acid. In all studies, inhibition of bile acid synthesis occurred twice as fast as its derepression.

Absence of transformation of beta-muricholic acid by human microflora implanted in the digestive tracts of germfree male rats.

Germfree rats biosynthetize cholic and beta-muricholic acids. The latter does not exist in humans. Germfree rats were given human fecal suspensions. These rats degraded cholic acid into deoxycholic acid but failed to metabolize beta-muricholic acid.

Cholesterol and bile acid metabolism in hypertensive arteriolipidosis-prone rats (ALR).

Bile flow and biliary bile acids were analyzed in arteriolipidosis-prone rats (ALR), the hypertensive model for lipidemic arterial lesions with reactive hypercholesterolemia and compared to those in spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto rats (WKY). Bile flow and bile acid secretion decreased in the order of WKY, SHR and ALR. When labelled cholesterol was given intraperitoneally, the biliary radioactivity secretion was significantly slow in ALR in comparison with that in WKY. The decay of radioactive cholesterol in serum after the injection was also delayed in ALR. Our data suggest that the abnormality in bile flow, biliary bile acid secretion and/or cholesterol turnover to bile acids may be pathogenically related to reactive hypercholesterolemia noted in the ALR.

Intestinal microflora and bile acids. In vitro cholic acid transformation by mixed fecal culture of rats.

In vitro cholic acid (CA) transformation by mixed fecal culture was investigated. Concentrations of glucose, peptone, and yeast extract in the medium and the initial pH of the medium markedly affected the CA transformation. Yeast extract enhanced the transformation, whereas high concentrations of glucose and peptone inhibited it. When the initial pH of the medium was below 6.5, CA was converted to 7-keto-deoxycholic acid (7KD), and formation of deoxycholic acid (DC) was not observed. In contrast, with an initial pH of 7.0, about 60% of the CA was converted to 7KD after 3 days of incubation, and then DC gradually formed after 4 days of incubation, following the disappearance of 7KD. The formation of DC in the cultured samples was paralleled in each case by disappearance of 7KD. In pure culture systems, Escherichia coli and some strains of Bacteroides formed 7KD from CA. No DC formation was observed in pure cultures of any of the strains examined.

Effect of pH on bile salt degradation by mixed fecal cultures.

Stoll specimens from 3 healthy volunteers were cultured under an-aerobic conditions in brain heart infusion broth with and without the addition of cholate, deoxycholate of chenodeoxycholate. The initial pH of the medium was adjusted to 5.5, 6.3, 7.3 (unadjusted), 8.0, and 9.0. Cell-free extracts prepared from the resulting bacterial growth contained increased levels of NAD- and NADP-dependent 3alpha-, 7alpha-, and 12alpha-hydroxysteroid oxidoreductases when the initial pH was 8.0 or 9.0 and depressed levels of these activities when the initial pH was 5.5 or 6.3 (as compared to control values obtained at 7.3). At pH 5,5 all activities except NAD-dependent 7alpha-hydroxysteroid oxidoreductase were absent. A powerful selective effect was imposed on NAD-dependent 7alpha-hydroxysteroid oxidoreductase when deoxycholate or chenodeoxycholate were incorporated into or chenodeoxycholate were incorporated into the medium. Thin-layer chromatography of either extracts of cholate-containing, acidified spent bacterial medium showed alkaline or neutral (optimal at pH 8). The precent hydroxyl group estimations at the 3alpha-, 7alpha-, and 12alpha-positions revealed an increase in disappearance of OH groups at all three positions with increasing initial pH value. The order of extent of bioconversion was 7alpha-OH greater than 3alpha-OH; at pH 8 AND 9, approximately 90% 7alpha-OH bioconversion was observed. Spent bacterial media and a number of commercial secondary bile salts were all negative in the Ames' assay for mutagenicity.