Search

Centenarians have a decreased susceptibility to ageing-associated illnesses, chronic inflammation and infectious diseases1–3. Here we show that centenarians have a distinct gut microbiome that is enriched in microorganisms that are capable of generating unique secondary bile acids, including various isoforms of lithocholic acid (LCA): iso-, 3-oxo-, allo-, 3-oxoallo- and isoallolithocholic acid. Among these bile acids, the biosynthetic pathway for isoalloLCA had not been described previously. By screening 68 bacterial isolates from the faecal microbiota of a centenarian, we identified Odoribacteraceae strains as effective producers of isoalloLCA both in vitro and in vivo. Furthermore, we found that the enzymes 5α-reductase (5AR) and 3β-hydroxysteroid dehydrogenase (3β-HSDH) were responsible for the production of isoalloLCA. IsoalloLCA exerted potent antimicrobial effects against Gram-positive (but not Gram-negative) multidrug-resistant pathogens, including Clostridioides difficile and Enterococcus faecium. These findings suggest that the metabolism of specific bile acids may be involved in reducing the risk of infection with pathobionts, thereby potentially contributing to the maintenance of intestinal homeostasis. The microbiota of centenarians (aged 100 years and older) comprise gut microorganisms that are capable of generating unique secondary bile acids, including isoallolithocholic acid, a bile acid with potent antimicrobial effects against Gram-positive—but not Gram-negative—multidrug-resistant pathogens.

Organic anion transporting polypeptide (OATP) 1B1 (gene, solute carrier organic anion transporter family member 1B1 [SLCO1B1]) and OATP1B3 (SLCO1B3) serve as transporters for hepatic uptake of important endogenous substances and several commonly prescribed drugs. Inactivation of both proteins together causes Rotor syndrome. How this OATP1B1/1B3 defect disturbs bile acid (BA) metabolism is largely unknown. In this study, we performed detailed BA analysis in 3 patients with genetically diagnosed Rotor syndrome. We found that BAs glucuronidated at the C‐3 position (BA‐3G) accounted for 50% or more of total BAs in these patients. In contrast but similarly to healthy controls, only trace amounts of BA‐3G were detected in patients with constitutional indocyanine green excretory defect (OATP1B3 deficiency) or sodium‐taurocholate cotransporting polypeptide (NTCP; gene, solute carrier family 10 member 1 [SLC10A1]) deficiency. Therefore, substantial amounts of BA‐3G are synthesized in hepatocytes. The cycling pathway of BA‐3G, consisting of excretion from upstream hepatocytes and uptake by downstream hepatocytes by OATP1B1/1B3 may exist to reduce the burden on upstream hepatocytes. Conclusion: Detailed BA analysis revealed glucuronidated bile acidemia in patients with Rotor syndrome. Further exploration of the physiologic role of glucuronidated BAs is necessary.

BACKGROUND Elobixibat, a novel inhibitor of apical sodium-dependent bile acid transporter for treating chronic constipation, increases colonic bile acid concentrations, stimulating bowel function. However, it is not clear which bile acids are altered, or whether altered gut microbiota are associated with functional effects that may alter bowel function. AIMS To investigate the effects of elobixibat on changes in the faecal concentrations of total and individual bile acids and in faecal microbiota. METHODS This was a prospective, single-centre study. After baseline period, patients received 10 mg daily of elobixibat for 2 weeks. We evaluated the effects on bowel function, changes in faecal bile acid concentrations and composition of gut bacteria, before and after elobixibat administration. RESULTS In the 30 patients analysed, the frequency of pre- and post-treatment bowel movements per fortnight was 7 and 10 (P

Background: Postnatal growth restriction in very-preterm infants (VPIs) may have long-lasting effects. Recent evidence suggests that developmental problems in VPIs are related to abnormalities in intestinal microbial communities. Objective: To investigate the effect on growth outcomes in VPIs of supplementation with Bifidobacterium along with mother’s colostrum and breast milk. Methods: A randomized controlled study was performed on 35 VPIs, born between 24 and 31 weeks of gestation with birth weights Bifidobacterium breve supplementation (Bifid group) or vehicle supplement only (placebo group). Parenteral nutrition was initiated with glucose, amino acids, and fatty acids for all of the infants soon after birth. Each infant received their own mother’s colostrum within 24 h of birth, and breast milk on subsequent days. Fecal bacteria, organic acids, pH, bile acids, and plasma fatty acids were analyzed. Results: Seventeen infants were allocated to the Bifid group and 18 to the placebo group; the birth weights and gestational ages did not differ significantly between the two groups. Compared to the placebo group, the Bifid group showed significantly greater and earlier weight gain by 8 weeks; significantly higher total fecal bacterial counts, including bifidobacteria; higher levels of total fecal short-chain fatty acids and nominally (but not significantly) higher concentrations of plasma n–3 fatty acids; and lower levels of total fecal bile acid. Conclusions: Bifidobacterial supplementation of maternal colostrum and breast milk yielded the establishment of a beneficial microbiota profile, leading to favorable metabolic responses that appeared to provide improved growth in VPIs.

The formation of secondary bile acids by gut microbes is a current topic of considerable biomedical interest. However, a detailed understanding of the biology of anaerobic bacteria in the genus Clostridium that are capable of generating secondary bile acids is lacking. We therefore sought to determine the transcriptional responses of two prominent secondary bile acid producing bacteria, Clostridium hylemonae and Clostridium hiranonis to bile salts (in vitro) and the cecal environment of gnotobiotic mice. The genomes of C. hylemonae DSM 15053 and C. hiranonis DSM 13275 were closed, and found to encode 3,647 genes (3,584 protein-coding) and 2,363 predicted genes (of which 2,239 are protein-coding), respectively, and 1,035 orthologs were shared between C. hylemonae and C. hiranonis. RNA-Seq analysis was performed in growth medium alone, and in the presence of cholic acid (CA) and deoxycholic acid (DCA). Growth with CA resulted in differential expression (>0.58 log2FC; FDR

The aim of this paper was to more completely study the mitochondrial CYP27A1 initiated acidic pathway of cholesterol metabolism. The mitochondrial CYP27A1 initiated pathway of cholesterol metabolism (acidic pathway) is known to synthesize two well-described vital regulators of cholesterol/lipid homeostasis, (25R)-26-hydroxycholesterol (26HC) and 25-hydroxycholesterol (25HC). Both 26HC and 25HC have been shown to be subsequently 7α-hydroxylated by Cyp7b1; reducing their regulatory abilities and furthering their metabolism to chenodeoxycholic acid (CDCA). Cholesterol delivery into the inner mitochondria membrane, where CYP27A1 is located, is considered the pathway's only rate-limiting step. To further explore the pathway, we increased cholesterol transport into mitochondrial CYP27A1 by selectively increased expression of the gene encoding the steroidogenic acute transport protein (StarD1). StarD1 overexpression led to an unanticipated marked down-regulation of oxysterol 7α-hydroxylase (Cyp7b1), a marked increase in 26HC, and the formation of a third vital regulatory oxysterol, 24(S)-hydroxycholesterol (24HC), in B6/129 mice livers. To explore the further metabolism of 24HC, as well as, 25HC and 26HC, characterizations of oxysterols and bile acids using three murine models (StarD1 overexpression, Cyp7b1-/-, Cyp27a1-/-) and human Hep G2 cells were conducted. This report describes the discovery of a new mitochondrial-initiated pathway of oxysterol/bile acid biosynthesis. Just as importantly, it provides evidence for CYP7B1 as a key regulator of three vital intracellular regulatory oxysterol levels.

Diagnosis of biliary atresia (BA) can involve uncertainties. In the present prospective multicenter study, we considered whether urinary oxysterols represent a useful marker for diagnosis of BA in Japanese children. Subjects under 6 months old at 7 pediatric centers in Japan were prospectively enrolled, including patients with cholestasis and healthy controls (HC) without liver disease. Patients with cholestasis constituted 2 groups representing BA patients and others with cholestasis from other causes (non-BA). We quantitatively analyzed 7 oxysterols including 4β-, 20(S)-, 22(S)-, 22(R)-, 24(S)-, 25-, and 27-hydroxycholesterol by liquid chromatography/electrospray ionization-tandem mass spectrometry. Enrolled subjects included 14 with BA (median age 68 days; range 26–170) and 10 non-BA cholestatic controls (59; 14–162), as well as 10 HC (57; 25–120). Total urinary oxysterols were significantly greater in BA (median, 153.0 μmol/mol creatinine; range 24.1–486.7; P P P

Centenarians, or individuals who have lived more than a century, represent the ultimate model of successful longevity associated with decreased susceptibility to ageing-associated illness and chronic inflammation. The gut microbiota is considered to be a critical determinant of human health and longevity. Here we show that centenarians (average 107 yo) have a distinct gut microbiome enriched in microbes capable of generating unique secondary bile acids, including iso-, 3-oxo-, and isoallo-lithocholic acid (LCA), as compared to elderly (85-89 yo) and young (21-55 yo) controls. Among these bile acids, the biosynthetic pathway for isoalloLCA had not been described previously. By screening 68 bacterial isolates from a centenarian’s faecal microbiota, we identified Parabacteroides merdae and Odoribacteraceae strains as effective producers of isoalloLCA. Furthermore, we generated and tested mutant strains of P. merdae to show that the enzymes 5α-reductase (5AR) and 3β-hydroxysteroid dehydrogenase (3βHSDH) were responsible for isoalloLCA production. This secondary bile acid derivative exerted the most potent antimicrobial effects among the tested bile acid compounds against gram-positive (but not gram-negative) multidrug-resistant pathogens, including Clostridioides difficile and vancomycin-resistant Enterococcus faecium. These findings suggest that specific bile acid metabolism may be involved in reducing the risk of pathobiont infection, thereby potentially contributing to longevity.

Centenarians have a decreased susceptibility to ageing-associated illnesses, chronic inflammation and infectious diseases

Background Berberine (BBR) is a plant-based nutraceutical that has been used for millennia to treat diarrheal infections and in contemporary medicine to improve patient lipid profiles. Reduction in lipids, particularly cholesterol, is achieved partly through up-regulation of bile acid synthesis and excretion into the gastrointestinal tract (GI). The efficacy of BBR is also thought to be dependent on structural and functional alterations of the gut microbiome. However, knowledge of the effects of BBR on gut microbiome communities is currently lacking. Distinguishing indirect effects of BBR on bacteria through altered bile acid profiles is particularly important in understanding how dietary nutraceuticals alter the microbiome. Results Germfree mice were colonized with a defined minimal gut bacterial consortium capable of functional bile acid metabolism (Bacteroides vulgatus, Bacteroides uniformis, Parabacteroides distasonis, Bilophila wadsworthia, Clostridium hylemonae, Clostridium hiranonis, Blautia producta; B4PC2). Multi-omics (bile acid metabolomics, 16S rDNA sequencing, cecal metatranscriptomics) were performed in order to provide a simple in vivo model from which to identify network-based correlations between bile acids and bacterial transcripts in the presence and absence of dietary BBR. Significant alterations in network topology and connectivity in function were observed, despite similarity in gut microbial alpha diversity (P = 0.30) and beta-diversity (P = 0.123) between control and BBR treatment. BBR increased cecal bile acid concentrations, (P < 0.05), most notably deoxycholic acid (DCA) (P +/H+ antiporter, cell wall synthesis/repair, carbohydrate metabolism and amino acid metabolism. Bile acid concentrations in the GI tract increased significantly during BBR treatment and developed extensive correlation networks with expressed genes in the B4PC2 community. Conclusions This work has important implications for interpreting the effects of BBR on structure and function of the complex gut microbiome, which may lead to targeted pharmaceutical interventions aimed to achieve the positive physiological effects previously observed with BBR supplementation.

NAFLD is an important public health issue closely associated with the pervasive epidemics of diabetes and obesity. Yet, despite NAFLD being among the most common of chronic liver diseases, the biological factors responsible for its transition from benign nonalcoholic fatty liver (NAFL) to NASH remain unclear. This lack of knowledge leads to a decreased ability to find relevant animal models, predict disease progression, or develop clinical treatments. In the current study, we used multiple mouse models of NAFLD, human correlation data, and selective gene overexpression of steroidogenic acute regulatory protein (StarD1) in mice to elucidate a plausible mechanistic pathway for promoting the transition from NAFL to NASH. We show that oxysterol 7α-hydroxylase (CYP7B1) controls the levels of intracellular regulatory oxysterols generated by the “acidic/alternative” pathway of cholesterol metabolism. Specifically, we report data showing that an inability to upregulate CYP7B1, in the setting of insulin resistance, results in the accumulation of toxic intracellular cholesterol metabolites that promote inflammation and hepatocyte injury. This metabolic pathway, initiated and exacerbated by insulin resistance, offers insight into approaches for the treatment of NAFLD.

Bile acid compositions are known to change dramatically after birth with aging. However, no reports have described the transition of conjugated urinary bile acids from the neonatal period to adulthood, and such findings would noninvasively offer insights into hepatic function. The aim of this study was to investigate differences in bile acid species, conjugation rates, and patterns, and to pool characteristics for age groups. We measured urinary bile acids in spot urine samples from 92 healthy individuals ranging from birth to 58 years old using liquid chromatography tandem mass spectrometry (LC/ESI-MS/MS). Sixty-six unconjugated and conjugated bile acids were systematically determined. After birth, urinary bile acids dramatically changed from fetal (i.e., Δ4-, Δ5-, and polyhydroxy-bile acids) to mature (i.e., CA and CDCA) bile acids. Peak bile acid excretion was 6–8 days after birth, steadily decreasing thereafter. A major change in bile acid conjugation pattern (taurine to glycine) also occurred at 2–4 months old. Our data provide important information regarding transitions of bile acid biosynthesis, including conjugation. The data also support the existence of physiologic cholestasis in the neonatal period and the establishment of the intestinal bacterial flora in infants.

Few reports describe oxysterols in healthy children or in children with liver disease. We aimed to determine whether developmental changes in urinary and serum oxysterols occur during childhood, and to assess whether oxysterols might be biomarkers for pediatric liver disease. Healthy children enrolled as subjects (36 and 35 for urine and serum analysis, respectively) included neonates, infants, preschoolers, and school-age children, studied along with 14 healthy adults and 8 children with liver disease. We quantitated 7 oxysterols including 4β-, 20(S)-, 22(S)-, 22(R)-, 24(S)-, 25-, and 27-hydroxycholesterol using liquid chromatography/electrospray ionization-tandem mass spectrometry. Urinary total oxysterols were significantly greater in neonates than in infants (P 0.05), preschoolers (P 0.001), school-age children (P 0.001), or adults (P 0.001), declining with age. Serum total oxysterols in neonates were significantly lower than in infants (P 0.05), preschoolers (P 0.001), school-age children (P 0.05), or adults (P 0.01). Compared with healthy children, total oxysterols and 24(S)-hydroxycholesterol in liver disease were significantly increased in both urine (P 0.001 and P 0.001, respectively) and serum (P 0.001 and P 0.05, respectively). Oxysterols in liver disease, particularly 24(S)-hydroxycholesterol, were greater in urine than serum. Oxysterols change developmentally and might serve as a biomarker for pediatric liver disease. To our knowledge, this is the first such report.

Patients with cirrhosis are often exposed to antibiotics that can lead to resistance and fungal overgrowth. The role of fecal microbial transplant (FMT) in restoring gut microbial function is unclear in cirrhosis. In a Food and Drug Administration-monitored phase 1 clinical safety trial, patients with decompensated cirrhosis on standard therapies (lactulose and rifaximin) were randomized to standard-of-care (SOC, no antibiotics/FMT) or 5 days of broad-spectrum antibiotics followed by FMT from a donor enriched in Lachnospiraceae and Ruminococcaceae. Microbial composition (diversity, family-level relative abundances), function (fecal bile acid [BA] deconjugation, 7α-dehydroxylation, short-chain fatty acids [SCFAs]), and correlations between Lachnospiraceae, Ruminococcaceae, and clinical variables were analyzed at baseline, postantibiotics, and 15 days post-FMT. FMT was well tolerated. Postantibiotics, there was a reduced microbial diversity and autochthonous taxa relative abundance. This was associated with an altered fecal SCFA and BA profile. Correlation linkage changes from beneficial at baseline to negative after antibiotics. All of these parameters became statistically similar post-FMT to baseline levels. No changes were seen in the SOC group. Conclusion In patients with advanced cirrhosis on lactulose and rifaximin, FMT restored antibiotic-associated disruption in microbial diversity and function. (Hepatology 2018; 00:000-000).

Cerebrotendinous xanthomatosis (CTX) is a rare, autosomal recessive, inborn disruption in bile acid synthesis characterized by severe systemic xanthomas, cataracts and neurological injuries occurring before adolescence without elevation of the serum cholesterol or triglyceride levels. CTX is caused by a deficiency of the mitochondrial enzyme sterol 27-hydroxylase, which is encoded by the CYP27A1 gene. We herein report a 50-year-old Japanese woman with late-onset CTX who had no relevant symptoms before the development of bilateral Achilles tendon xanthomas in middle age. A genetic analysis revealed a compound heterozygous mutation in the CYP27A1 gene with a previously known missense mutation (NM_000784.3:c.1421 G>A) and a novel frame shift mutation of NM_000784.3:c.1342_1343insCACC.

Background Cirrhosis and alcohol can independently affect the gut–liver axis with systemic inflammation. However, their concurrent impact in humans is unclear. Methods Our aim was to determine the effect of continued alcohol misuse on the gut-liver axis in cirrhotic patients. Age- and MELD-balanced cirrhotic patients who were currently drinking (Alc) or abstinent (NAlc) and healthy controls underwent serum and stool collection. A subset underwent upper endoscopy and colonoscopy for biopsies and duodenal fluid collection. The groups were compared regarding (i) inflammation/intestinal barrier: systemic tumor necrosis factor levels, intestinal inflammatory cytokine (duodenum, ileum, sigmoid), and ileal antimicrobial peptide expression; (ii) microbiota composition: 16SrRNA sequencing of duodenal, ileal, and colonic mucosal and fecal microbiota; and (iii) microbial functionality: duodenal fluid and fecal bile acid (BA) profile (conjugation and dehydroxylation status), intestinal BA transporter (ASBT, FXR, FGF-19, SHP) expression, and stool metabolomics using gas chromatography/mass spectrometry. Results Alc patients demonstrated a significant duodenal, ileal, and colonic mucosal and fecal dysbiosis, compared to NAlc and controls with lower autochthonous bacterial taxa. BA profile skewed toward a potentially toxic profile (higher secondary and glycine-conjugated BAs) in duodenal fluid and stool in Alc patients. Duodenal fluid demonstrated conjugated secondary BAs only in the Alc group. There was a greater expression of all ileal BA transporters in Alc patients. This group also showed higher endotoxemia, systemic and ileal inflammatory expression, and lower amino acid and bioenergetic-associated metabolites, without change in antimicrobial peptide expression. Conclusions Despite cirrhosis, continued alcohol misuse predisposes patients to widespread dysbiosis with alterations in microbial functionality such as a toxic BA profile, which can lead to intestinal and systemic inflammation.

Background: We encountered 7 Japanese patients with bile acid synthesis disorders (BASD) over 21 years between 1996 and 2017. Diagnoses were made by bile acid and genetic analyses. We gave low-dose, long-term chenodeoxycholic acid (CDCA) treatment to 5 of the patients, who had 3β-hydroxy-Δ 5 -C 27 -steroid dehydrogenase/isomerase (3β-HSD) deficiency (n=3) or Δ 4 -3-oxosteroid 5β-reductase (5β-reductase) deficiency (n=2). Another patient with the latter diagnosis whose bile acid analyses had mitigating features was maintained on ursodeoxycholic acid according to parental preferences and now remains healthy after discontinuation of treatment. A patient with oxysterol 7α-hydroxylase deficiency developed liver failure and fully recovered after successful liver transplantation. We used clinical records to clarify long-term outcome and value of CDCA in the other patients. Efficacy of CDCA treatment was evaluated in the 5 patients given a low dose (5 to 10 mg/kg/day) for a long term. Results: Medians with ranges of current patient ages and duration of CDCA treatment are10 years (8 to 43) and 10 years (8 to 21), respectively. All 7 patients, who had homozygous or compound heterozygous mutations in the HSD3B7 , SRD5B1 , or CYP7B1 gene, are currently in good health without liver dysfunction. In the 5 patients with CDCA treatment, hepatic function gradually improved following initiation. No adverse effects were noted. Conclusions: We concluded that low-dose CDCA treatment is effective in 3β-HSD deficiency and 5β-reductase deficiency, as cholic acid has been in other countries. BASD carry a good prognosis following early diagnosis and initiation of long-term, low-dose CDCA treatment.

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.

BACKGROUND. Hepatic encephalopathy (HE) is associated with poor outcomes. A prior randomized, pilot trial demonstrated safety after oral capsular fecal microbial transplant (FMT) in HE, with favorable changes in microbial composition and cognition. However, microbial functional changes are unclear. The aim of this study was to determine the effect of FMT on the gut-brain axis compared with placebo, using microbial function based on bile acids (BAs), inflammation (serum IL-6, LPS-binding protein [LBP]), and their association with EncephalApp. METHODS. Twenty cirrhotic patients were randomized 1:1 into groups that received 1-time FMT capsules from a donor enriched in Lachnospiraceae and Ruminococcaceae or placebo capsules, with 5-month follow-up for safety outcomes. Stool microbiota and BA; serum IL-6, BA, and LBP; and EncephalApp were analyzed at baseline and 4 weeks after FMT/placebo. Correlation networks among microbiota, BAs, EncephalApp, IL-6, and LBP were performed before/after FMT. RESULTS. FMT-assigned participants had 1 HE recurrence and 2 unrelated infections. Six placebo-assigned participants developed negative outcomes. FMT, but not placebo, was associated with reduced serum IL-6 and LBP and improved EncephalApp. FMT-assigned participants demonstrated higher deconjugation and secondary BA formation in feces and serum compared with baseline. No change was seen in placebo. Correlation networks showed greater complexity after FMT compared with baseline. Beneficial taxa, such as Ruminococcaceae, Verrucomicrobiaceae, and Lachnospiraceae, were correlated with cognitive improvement and decrease in inflammation after FMT. Fecal/serum secondary/primary ratios and PiCRUST secondary BA pathways did not increase in participants who developed poor outcomes. CONCLUSION. Gut microbial function in cirrhosis is beneficially affected by capsular FMT, with improved inflammation and cognition. Lower secondary BAs in FMT recipients could select for participants who develop negative outcomes. TRIAL REGISTRATION. Clinicaltrials.gov NCT03152188. FUNDING. National Center for Advancing Translational Sciences NIH grant R21TR002024, VA Merit Review grant 2I0CX001076, the United Kingdom National Institute for Health Research Biomedical Facility at Imperial College London, the British Heart Foundation, Wellcome Trust, and King’s College London.

The formation of secondary bile acids by gut microbes is a current topic of considerable biomedical interest. However, a detailed understanding of the biology of anaerobic bacteria in the genus Clostridium that are capable of generating secondary bile acids is lacking. We therefore sought to determine the transcriptional responses of two prominent secondary bile acid producing bacteria, Clostridium hylemonae and Clostridium hiranonis to bile salts (in vitro) and the cecal environment of gnotobiotic mice. The genomes of C. hylemonae DSM 15053 and C. hiranonis DSM 13275 were closed, and found to encode 3,647 genes (3,584 protein-coding) and 2,363 predicted genes (of which 2,239 are protein-coding), respectively, and 1,035 orthologs were shared between C. hylemonae and C. hiranonis. RNA-Seq analysis was performed in growth medium alone, and in the presence of cholic acid (CA) and deoxycholic acid (DCA). Growth with CA resulted in differential expression (>0.58 log(2)FC; FDR

Chemotherapeutic agents can reduce bone marrow (BM) activity, causing myelosuppression, a common life-threatening complication of cancer treatment. It is challenging to predict the patients in whom prolonged myelosuppression will occur, resulting in a delay or discontinuation of the treatment protocol. An early indicator of recovery from myelosuppression would thus be highly beneficial in clinical settings. In this study, bile acids (BAs) were highly increased in the systemic circulation as a natural response during recovery from myelosuppression, supporting regeneration of BM cells. BA levels in the blood of pediatric cancer patients and mice treated with chemotherapeutic agents were increased, in synchrony with early proliferation of BM cells and recovery from myelosuppression. In a mouse model of altered BA composition, Cyp8b1 knockout mice, a subset of mice recovered poorly after chemotherapy. The poor recovery correlated with low levels and changes in composition of BAs in the liver and systemic circulation. Conversely, BA supplementation in chemotherapy-treated wild-type mice resulted in significantly improved recovery. The results suggest that part of the mechanism by which BAs support recovery is the suppression of endoplasmic reticulum stress pathways in expanding and recovering hematopoietic cells. The findings propose a novel role of BAs as early markers of recovery and active components of the recovery process after chemotherapy.

BACKGROUND In pediatric patients with cholestasis of unknown cause, inborn errors of bile acid (BA) synthesis (IEBAS) may be considered. For the initial screening for IEBAS, clarification of the urine BA profile is essential. The transportation of urine in a frozen state via air delivery, however, is laborious and costly. This study assessed the feasibility of using dried urine spots (DUS) to establish a more convenient and affordable method of IEBAS screening. METHODS We created DUS using urine samples from patients with 3β-hydroxy-Δ5-C27-steroid dehydrogenase/isomerase deficiency (3β-HSD) and Δ4-3-oxo-steroid 5β-reductase deficiency as standard preparations. We started accepting DUS specimens by regular mail. RESULTS The ratio of unusual to usual BA is essential for the initial detection of IEBAS, and the recovery rates of abnormal BA were acceptable. The recovery rate of Δ4-BA on day 28 decreased to 31.8% at 25°C, and to 19.6% at 37°C. Therefore, the sending of DUS should be avoided under conditions of high temperature. Of a total of 49 children with cholestasis, eight new patients were diagnosed with IEBAS using this screening method. CONCLUSION The mailing screening system is expected to facilitate the shipment, from regions outside of Japan, of samples for IEBAS screening.

AIM: To better understand the effect of partial external biliary diversion (PEBD) in patients with progressive familial intrahepatic cholestasis 1 (PFIC1). MATERIALS AND METHODS: We analyzed quantitative changes in the concentrations of individual bile acids in serum, urine, and biliary bile, and concomitant changes in symptoms such as pruritus, in 2 patients whose PFIC1 clinically evolved to resemble the course of benign recurrent intrahepatic cholestasis (BRIC) upon treatment with PEBD. RESULTS: Patient 1 showed good therapeutic effect after PEBD including improved liver function test results and decreased total bile acids in serum, as well as decreased pruritus. The cholic acid (CA)/chenodeoxycholic acid (CDCA) ratio in biliary bile also decreased. Patient 2, however, experienced little decrease in pruritus after PEBD, and showed no clear change in the CA/CDCA ratio. Nonetheless, this patient’s course did not rapidly worsen. CONCLUSION: In PFIC1, increased bile secretion and a decreased CA/CDCA ratio in biliary bile may be the most important responses after PEBD. Relief of pruritus is also an important sign of therapeutic effect. Moreover, progression of symptoms in PFIC1 sometimes can be delayed by PEBD, resulting in a relatively mild BRIC-like course, even when the CA/CDCA ratio and pruritus show little change. However, mechanisms underlying clinical benefits remain obscure.

Liver transplantation (LT) improves daily function and ameliorates gut microbial composition. However, the effect of LT on microbial functionality, which can be related to overall patient benefit, is unclear and could affect the post-LT course. The aims were to determine the effect of LT on gut microbial functionality focusing on endotoxemia, bile acid (BA), ammonia metabolism, and lipidomics. We enrolled outpatient patients with cirrhosis on the LT list and followed them until 6 months after LT. Microbiota composition (Shannon diversity and individual taxa) and function analysis (serum endotoxin, urinary metabolomics and serum lipidomics, and stool BA profile) and cognitive tests were performed at both visits. We enrolled 40 patients (age, 56 ± 7 years; mean Model for End-Stage Liver Disease score, 22.6). They received LT 6 ± 3 months after enrollment and were re-evaluated 7 ± 3 months after LT with a stable course. A significant improvement in cognition with increase in microbial diversity, increase in autochthonous and decrease in potentially pathogenic taxa, and reduced endotoxemia were seen after LT compared with baseline. Stool BAs increased significantly after LT, and there was evidence of greater bacterial action (higher secondary, oxo and iso-BAs) after LT although the levels of conjugated BAs remained similar. There was a reduced serum ammonia and corresponding rise in urinary phenylacetylglutamine after LT. There was an increase in urinary trimethylamine-N-oxide, which was correlated with specific changes in serum lipids related to cell membrane products. The ultimate post-LT lipidomic profile appeared beneficial compared with the profile before LT. In conclusion, LT improves gut microbiota diversity and dysbiosis, which is accompanied by favorable changes in gut microbial functionality corresponding to BAs, ammonia, endotoxemia, lipidomic, and metabolomic profiles. Liver Transplantation 24 752-761 2018 AASLD.

Background & Aims The 7α-dehydroxylation of primary bile acids (BAs), chenodeoxycholic (CDCA) and cholic acid (CA) into the secondary BAs, lithocholic (LCA) and deoxycholic acid (DCA), is a key function of the gut microbiota. We aimed at studying the linkage between fecal BAs and gut microbiota in cirrhosis since this could help understand cirrhosis progression. Methods Fecal microbiota were analyzed by culture-independent multitagged-pyrosequencing, fecal BAs using HPLC and serum BAs using LC–MS in controls, early (Child A) and advanced cirrhotics (Child B/C). A subgroup of early cirrhotics underwent BA and microbiota analysis before/after eight weeks of rifaximin. Results Cross-sectional: 47 cirrhotics (24 advanced) and 14 controls were included. In feces, advanced cirrhotics had the lowest total, secondary, secondary/primary BA ratios, and the highest primary BAs compared to early cirrhotics and controls. Secondary fecal BAs were detectable in all controls but in a significantly lower proportion of cirrhotics ( p Enterobacteriaceae (potentially pathogenic) but lower Lachonospiraceae , Ruminococcaceae and Blautia (7α-dehydroxylating bacteria) abundance. CDCA was positively correlated with Enterobacteriaceae (r=0.57, p Ruminococcaceae were positively correlated with DCA (r=0.4, p Ruminococcaceae and DCA/CA (r=0.82, p Blautia with LCA/CDCA (r=0.61, p Veillonellaceae and in secondary/primary BA ratios. Conclusions Cirrhosis, especially advanced disease, is associated with a decreased conversion of primary to secondary fecal BAs, which is linked to abundance of key gut microbiome taxa.

A method for the determination of conjugated and unconjugated 3β-hydroxy-Δ 5 -bile acids and related bile acids in human urine and serum has been developed using high-performance liquid chromatography–electrospray ionization-tandem mass spectrometry. Calibration curves for the bile acids were linear over the range of 10–2000 pmol/mL, and the detection limit was less than 4 pmol/mL for all bile acids using selected reaction monitoring analysis. The bile acids in urine and serum samples from two patients with severe cholestatic liver disease were measured by this analytical method. Glycine-conjugated 3β-hydroxy-Δ 5 -bile acid 3-sulfates were determined to be the major bile acids in the urine and serum from patients with a 3β-hydroxy-Δ 5 -C 27 -steriod dehydrogenase/isomerase deficiency or dysfunction.

Inborn errors of bile acid synthesis are rare genetic disorders that can present as neonatal cholestasis and fat-soluble vitamin deficiency. Though rare, these diseases account for some patients with cholestasis of unknown etiology. Seven known defects of bile acid synthesis occur in children. These defects may be categorized as deficiencies in activity of enzymes catalyzing reactions affecting the steroid nucleus or the side chain. Defects in reactions involving the steroid nucleus include cholesterol 7α-hydroxylase deficiency, 3β-hydroxysteroid-Δ5-C27-steroid dehydrogenase/isomerase deficiency, Δ4-3-oxosteroid 5β-reductase deficiency, and oxysterol 7α-hydroxylase deficiency. Defects in reactions involving side-chain modification are sterol 27-hydroxylase deficiency, α-methyl-CoA racemase deficiency, disorders of peroxisomal β-oxidation, bile acid-CoA: amino acid N-acyltransferase deficiency, and bile acid-CoA ligase deficiency. Cholesterol 7α-hydroxylase deficiency and disorders of peroxisomal β-oxidation are not considered here, since cholesterol 7α-hydroxylase deficiency is not known to occur in children and disorders of peroxisomal β-oxidation represent disease of peroxisomes. When identified early, many patients with inborn errors of bile acid synthesis have a favorable clinical response to oral primary bile acid therapy. These inborn errors characteristically result in elevated serum bilirubin and aminotransferase concentrations but produce no abnormalities of serum γ-glutamyltransferase or of total bile acid concentrations detectable by enzymatic methods. Screening for inborn errors of bile acid synthesis using fast atom bombardment ionization mass spectrometry, gas chromatography-mass spectroscopy, and liquid chromatography-electrospray ionization tandem mass spectrometry is useful. Genetic analysis is available for a definitive diagnosis. Disorders of bile acid synthesis account for some 2–3% of screened cases of cholestatic liver disease in infants and children.

A method has been developed for the measurement of transport activities in membrane vesicles obtained from Sf9 cells for 3β-hydroxy-Δ(5)-bile acids by high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry. Calibration curves for the bile acids were linear over the range of 10 to 2000 pmol/mL, and the detection limit was less than 1 pmol/mL for 3β-hydroxy-Δ(5)-bile acids using selected reaction monitoring analysis. The analytical method was applied to measurements of transport activities in membrane vesicles obtained from human multidrug resistance-associated protein 2-, 3-, and human bile salt export pump-expressing Sf9 cells for conjugated 3β-hydroxy-Δ(5)-bile acids. The present study demonstrated that human multidrug resistance-associated protein 3 vesicles accepted conjugated 3β-hydroxy-Δ(5)-bile acids along with common bile acids such as glycocholic acid and taurolithocholic acid 3-sulfate.

We previously reported that alcohol drinkers with and without cirrhosis showed a significant increase in fecal bile acid secretion compared to nondrinkers. We hypothesized this may be due to activation by alcohol of hepatic cyclic adenosine monophosphate responsive element-binding protein 3-like protein 3 (CREBH), which induces cholesterol 7α-hydroxylase (Cyp7a1). Alternatively, the gut microbiota composition in the absence of alcohol might increase bile acid synthesis by up-regulating Cyp7a1. To test this hypothesis, we humanized germ-free (GF) mice with stool from healthy human subjects (Ctrl-Hum), human subjects with cirrhosis (Cirr-Hum), and human subjects with cirrhosis and active alcoholism (Alc-Hum). All animals were fed a normal chow diet, and none demonstrated cirrhosis. Both hepatic Cyp7a1 and sterol 12α-hydroxylase (Cyp8b1) messenger RNA (mRNA) levels were significantly induced in the Alc-Hum and Ctrl-Hum mice but not in the Cirr-Hum mice or GF mice. Liver bile acid concentration was correspondingly increased in the Alc-Hum mice despite fibroblast growth factor 15, fibroblast growth receptor 4, and small heterodimer partner mRNA levels being significantly induced in the large bowel and liver of the Ctrl-Hum mice and Alc-Hum mice but not in the Cirr-Hum mice or GF mice. This suggests that the normal pathways of Cyp7a1 repression were activated in the Alc-Hum mice and Ctrl-Hum mice. CREBH mRNA was significantly induced only in the Ctrl-Hum mice and Alc-Hum mice, possibly indicating that the gut microbiota up-regulate CREBH and induce bile acid synthesis genes. Analysis of stool bile acids showed that the microbiota of the Cirr-Hum and Alc-Hum mice had a greater ability to deconjugate and 7α-dehydroxylate primary bile acids compared to the microbiota of the Cirr-Hum mice. 16S ribosomal RNA gene sequencing of the gut microbiota showed that the relative abundance of taxa that 7-α dehydroxylate primary bile acids was higher in the Ctrl-Hum and Alc-Hum groups. Conclusion: The composition of gut microbiota influences the regulation of the rate-limiting enzymes in bile acid synthesis in the liver. (Hepatology Communications 2017;1:61-70).

In two Japanese infants with neonatal cholestasis, 3-oxo-Δ(4)-steroid 5β-reductase deficiency was diagnosed based on mutations of the SRD5B1 gene. Unusual bile acids such as elevated 3-oxo-Δ(4) bile acids were detected in their serum and urine by gas chromatography-mass spectrometry. We studied effects of oral chenodeoxycholic acid treatment.SRD5B1 gene analysis used peripheral lymphocyte genomic DNA. Diagnosis and treatment of these two patients were investigated retrospectively and prospectively investigated.With respect to SRD5B1, one patient was heterozygous (R266Q, a novel mutation) while the other was a compound heterozygote (G223E/R261C). Chenodeoxycholic acid treatment was effective in improving liver function and decreasing unusual bile acids such as 7α-hydroxy- and 7α,12α-dihydroxy-3-oxo-4-cholen-24-oic acids in serum and urine.Primary bile acid treatment using chenodeoxycholic acid was effective for these patients treated in early infancy before the late stage of chronic cholestatic liver dysfunction.