Intestinal Bile Acid Transport Research Articles

Hypothalamic SLC7A14 accounts for aging-reduced lipolysis in white adipose tissue of male mice

The central nervous system has been implicated in the age-induced reduction in adipose tissue lipolysis. However, the underlying mechanisms remain unclear. Here, we show the expression of SLC7A14 is reduced in proopiomelanocortin (POMC) neurons of aged mice. Overexpression of SLC7A14 in POMC neurons alleviates the aging-reduced lipolysis, whereas SLC7A14 deletion mimics the age-induced lipolysis impairment. Metabolomics analysis reveals that POMC SLC7A14 increased taurochenodeoxycholic acid (TCDCA) content, which mediates the SLC7A14 knockout- or age-induced WAT lipolysis impairment. Furthermore, SLC7A14-increased TCDCA content is dependent on intestinal apical sodium-dependent bile acid transporter (ASBT), which is regulated by intestinal sympathetic afferent nerves. Finally, SLC7A14 regulates the intestinal sympathetic afferent nerves by inhibiting mTORC1 signaling through inhibiting TSC1 phosphorylation. Collectively, our study suggests the function for central SLC7A14 and an upstream mechanism for the mTORC1 signaling pathway. Moreover, our data provides insights into the brain–gut–adipose tissue crosstalk in age-induced lipolysis impairment.

Increased intestinal bile acid absorption contributes to age-related cognitive impairment

Cognitive impairment in the elderly is associated with alterations in bile acid (BA) metabolism. In this study, we observe elevated levels of serum conjugated primary bile acids (CPBAs) and ammonia in elderly individuals, mild cognitive impairment, Alzheimer's disease, and aging rodents, with a more pronounced change in females. These changes are correlated with increased expression of the ileal apical sodium-bile acid transporter (ASBT), hippocampal synapse loss, and elevated brain CPBA and ammonia levels in rodents. Invitro experiments confirm that a CPBA, taurocholic acid, and ammonia induced synaptic loss. Manipulating intestinal BA transport using ASBT activators or inhibitors demonstrates the impact on brain CPBA and ammonia levels as well as cognitive decline in rodents. Additionally, administration of an intestinal BA sequestrant, cholestyramine, alleviates cognitive impairment, normalizing CPBAs and ammonia in aging mice. These findings highlight the potential of targeting intestinal BA absorption as a therapeutic strategy for age-related cognitive impairment.

Apple-derived extracellular vesicles modulate the expression of human intestinal bile acid transporter ASBT/SLC10A2 via downregulation of transcription factor RARα

PurposePlant-derived extracellular vesicles (EVs) have been reported to exert biological activity on intestinal tissues by delivering their contents into intestinal cells. We previously reported that ASBT/SLC10A2 mRNA was downregulated by apple-derived extracellular vesicles (APEVs). ASBT downregulation is effective in the treatment of cholestasis and chronic constipation, similar to the beneficial effects of apples. Therefore, this study aimed to establish the mechanism of ASBT downregulation by APEVs, focusing on microRNAs present in APEVs. ResultsAPEVs downregulated the expression of ASBT, but no significant effect on SLC10A2-3′UTR was observed. Proteomics revealed that APEVs decreased the expression of RARα/NR1B1. The binding of RARα to SLC10A2 promoter was also decreased by APEVs. The stability of NR1B1 mRNA was attenuated by APEVs and its 3′UTR was found to be a target for APEVs. Apple microRNAs that were predicted to interact with NR1B1-3′UTR were present in APEVs, and their mimics suppressed NR1B1 mRNA expression. ConclusionsSuppression of ASBT by APEVs was indirectly mediated by the downregulation of RARα, and its stability was lowered by microRNAs present in APEVs. This study suggested that macromolecules in food directly affect intestinal function by means of EVs that stabilize them and facilitate their cellular uptake.

In vitro models to measure effects on intestinal deconjugation and transport of mixtures of bile acids

Bile acid metabolism and transport are critical to maintain bile acid homeostasis and host health. In this study, it was investigated if effects on intestinal bile acid deconjugation and transport can be quantified in vitro model systems using mixtures of bile acids instead of studying individual bile acids. To this end deconjugation of mixtures of selected bile acids in anaerobic rat or human fecal incubations and the effect of the antibiotic tobramycin on these reactions was studied. In addition, the effect of tobramycin on the transport of the bile acids in isolation or in a mixture across Caco-2 cell layers was characterized. The results demonstrate that both the reduction of bile acid deconjugation and transport by tobramycin can be adequately detected in vitro systems using a mixture of bile acids, thus eliminating the need to characterize the effects for each bile acid in separate experiments. Subtle differences between the experiments with single or combined bile acids point at mutual competitive interactions and indicate that the use of bile acid mixtures is preferred over use of single bile acid given that also in vivo bile acids occurs in mixtures.

Apple juice relieves loperamide-induced constipation in rats by downregulating the intestinal apical sodium-dependent bile acid transporter ASBT.

Apples are known to exhibit various beneficial effects on human health. In the present study, we investigated the effect of continuous intake of apple juice (AJ) on constipation status. A single dose of loperamide in rats as the constipation model markedly decreased the weight and number of fecal pellets compared to saline-administered rats as a control. After the administration of AJ twice a day for seven days, recovery of defecation close to that of the control was observed in loperamide-treated rats. In addition, the total bile acid content in the feces increased from day 4 after the administration of AJ. Among hepatic and intestinal transporters and enzymes that regulate bile acids, the mRNA expression of the apical sodium-dependent bile acid transporter (Asbt, slc10a2) was decreased by AJ in rats. Furthermore, the Asbt-mediated bile acid transport activity in the rat ileum decreased after AJ administration. Moreover, in human colonic cancer-derived Caco-2 cells, AJ exposure for 24 and 48 h decreased the expressions of ASBT mRNA and protein, and the uptake activity of taurocholic acid in both 7- and 21-d cultures. Several components of AJ, such as procyanidins, decreased the expression of ASBT in Caco-2 cells. In conclusion, ASBT downregulation is a possible mechanism responsible for the constipation-relieving effect of apples, and procyanidins may play a role in downregulating ASBT, which leads to the beneficial effects of apples against constipation. Although it is generally agreed that the common dietary compositions play a role in constipation relief, the novel specific mechanism of apples found in this study would facilitate understanding food functions.

Biomimetic and temporal-controlled nanocarriers with ileum transporter targeting for achieving oral administration of chemotherapeutic drugs

BackgroundOral chemotherapy is preferred for patients with cancer owing to its multiple advantages, including convenience, better patient compliance, and improved safety. Nevertheless, various physical barriers exist in this route that hamper the development of oral chemotherapeutic formulations, including destruction of drugs in the gastrointestinal tract (GIT), low permeability in enterocytes, and short residence time in the intestine. To overcome these limitations, it is necessary to design an efficient oral drug delivery system with high efficacy and improved safety.ResultsHerein, we designed novel glycocholic acid (GCA)-functionalized double layer nanoparticles (GCA-NPs), which can act via an endogenous pathway and in a temporally controlled manner in the intestine, to enhance the oral bioavailability of hydrophobic chemotherapeutic drugs such as paclitaxel (PTX). GCA-NPs were composed of quercetin (Qu)-modified liposomes (QL) coated with GCA-chitosan oligosaccharide conjugate (GCOS). The GCA-NPs thus prepared showed prolonged intestinal retention time and good GIT stability due to the presence of chitosan oligosaccharide (COS) and enhanced active transportation via intestinal apical sodium-dependent bile acid transporter (ASBT) due to the presence of GCA. GCA-NPs also efficiently inhibited intestinal P-gp induced by Qu. PTX-loaded GCA-NPs (PTX@GCA-NPs) had a particle size of 84 nm and an entrapment efficiency of 98% with good stability. As a result, the oral bioavailability of PTX was increased 19-fold compared to that of oral Taxol® at the same dose. Oral PTX@GCA-NPs displayed superior antitumor efficacy and better safety than Taxol® when administered intravenously.ConclusionsOur novel drug delivery system showed remarkable efficacy in overcoming multiple limitations and is a promising carrier for oral delivery of multiple drugs, which addresses several challenges in oral delivery in the clinical context.Graphical

Effects of Intestinal FXR-Related Molecules on Intestinal Mucosal Barriers in Biliary Tract Obstruction.

Background: The farnesoid X receptor (FXR) is a key factor regulating hepatic bile acid synthesis and enterohepatic circulation. Repression of bile acid synthesis by the FXR is a potential strategy for treating cholestatic liver disease. However, the role of intestinal FXR on the intestinal barrier and intestinal microbiota needs further investigation. Materials: Intestinal tissues were collected from patients with biliary atresia or without hepatobiliary disease. Then, intestinal mRNA levels of FXR-related molecules were determined. To investigate the effect of FXR activation, bile-duct-ligation rats were treated with obeticholic acid [OCA (5 mg/kg/day)] or vehicle (0.5% methyl cellulose) per oral gavage for 14 days. The mRNA levels of intestinal FXR, SHP, TNF-α, FGF15 and bile acid transporter levels were determined. In addition, the intestinal permeability, morphologic changes, and composition of the intestinal microbiota were evaluated. Gut Microbiome was determined by 16S rDNA MiSeq sequencing, and functional profiling of microbial communities was predicted with BugBase and PICRUSt2. Finally, the role of OCA in injured intestinal epithelial cell apoptosis and proliferation was examined by pretreatment with lipopolysaccharide (LPS) in Caco-2 cells. Results: The downstream of the FXR in ileum tissues was inhibited in biliary obstruction. Activation of the FXR signaling pathway by OCA significantly reduced liver fibrosis and intestinal inflammation, improved intestinal microbiota, and protected intestinal mucosa in BDL rats. OCA also altered the functional capacities of ileum microbiota in BDL rats. Significant differences existed between the controls and BDL rats, which were attenuated by OCA in the alpha diversity analysis. Principal coordinates analysis showed that microbial communities in BDL rats clustered separately from controls, and OCA treatment attenuated the distinction. Bugbase and PICRUSt2 analysis showed that OCA changed the composition and structure of the intestinal microbiota and improved the metabolic function of the intestinal microbiota by increasing the relative abundance of beneficial bacteria and reducing the relative abundance of harmful bacteria. Moreover, OCA reduced the apoptosis induced by LPS in Caco-2 cells. Conclusion: The FXR agonist, OCA, activates the intestinal FXR signaling pathway and improves the composition and structure of the intestinal microbiota and intestinal barrier in BDL rats.

Impact of long-term administration of maralixibat on children with cholestasis secondary to Alagille syndrome.

There is growing interest in, but limited data about, intestinal bile acid transport inhibitors as treatment for cholestatic liver disease. The current analyses combine two similar randomized placebo‐controlled trials with subsequent extension phases investigating the impact of maralixibat in children with severe cholestasis secondary to Alagille Syndrome (n = 57). The primary outcomes were measures of pruritus (ItchRO[Obs]) and clinician scratch scale (CSS), both increasing in severity from 0 to 4) and quality of life (QoL) (Parent PedsQL and Multidimensional Fatigue Scale module [MFS] scaled 0–100 with increased QoL) at week 48 of the extension phase relative to the baseline of the placebo‐controlled trials (week 13). Secondary assessments included other clinical and biochemical parameters assessed in participants at week 72 or end of treatment (after week 48). At week 48, statistically and clinically significant least square mean (95% CI) improvements in pruritus and QoL were observed (ItchRO[Obs] −1.59 [−1.81, −1.36], CSS −1.36 [−1.67, −1.05], PedsQL +10.17 [4.48, 15.86], and multidimension fatigue [MFS] +13.97 [7.85, 20.08]). At week 48, serum bile acids, platelet count, and cholesterol decreased, whereas alanine aminotransferase (ALT) increased and total bilirubin (TB) and albumin were stable. Changes were durable at week 72 and end of treatment. There were no deaths; 2 participants underwent liver transplantation. Study drug was discontinued in 9 participants after treatment‐emergent adverse events, 6 of which were events of increased ALT or TB. Conclusion: Maralixibat administration was associated with marked improvement in pruritus and QoL. Interpretation of these findings is complicated by the complex natural history of severe cholestasis in Alagille syndrome.

Use of funded multicenter prospective longitudinal databases to inform clinical trials in rare diseases-Examination of cholestatic liver disease in Alagille syndrome.

The conduct of long‐term conventional randomized clinical trials in rare diseases is very difficult, making evidenced‐based drug development problematic. As a result, real‐world data/evidence are being used more frequently to assess new therapeutic approaches in orphan diseases. In this investigation, inclusion and exclusion criteria from a published trial of maralixibat in Alagille syndrome (ALGS, ITCH NCT02057692) were applied to a prospective longitudinal cohort of children with cholestasis (LOGIC NCT00571272) to derive contextual comparator data for evolving clinical trials of intestinal bile acid transport inhibitors in ALGS. A natural history/clinical care cohort of 59 participants who met adapted inclusion and exclusion criteria of ITCH was identified from 252 LOGIC participants with ALGS with their native liver. Frequency weighting was used to match the age distribution of ITCH and yielded a cohort (Alagille Syndrome Natural History [ALGS NH]) that was very similar to the baseline status of ITCH participants. During a 2‐year prospective follow‐up there was a significant reduction in pruritus in the weighted ALGS NH cohort as assessed by the clinician scratch score (−1.43 [0.28] −1.99, −0.87; mean [SEM] 95% confidence interval). During the same time period, the total bilirubin, albumin, and alanine aminotransferase levels were unchanged, whereas platelet count dropped significantly (−65.2 [16.2] −98.3, −32.1). Weighted survival with native liver was 91% at 2 years in the ALGS NH. These investigations provide valuable real‐world data that can serve as contextual comparators to current clinical trials, especially those without control populations, and highlight the value and importance of funded multicenter, prospective, natural history studies.

Hepatitis D Virus Entry Inhibitors Based on Repurposing Intestinal Bile Acid Reabsorption Inhibitors.

Identification of Na+/taurocholate co-transporting polypeptide (NTCP) as high-affinity hepatic entry receptor for the Hepatitis B and D viruses (HBV/HDV) opened the field for target-based development of cell-entry inhibitors. However, most of the HBV/HDV entry inhibitors identified so far also interfere with the physiological bile acid transporter function of NTCP. The present study aimed to identify more virus-selective inhibitors of NTCP by screening of 87 propanolamine derivatives from the former development of intestinal bile acid reabsorption inhibitors (BARIs), which interact with the NTCP-homologous intestinal apical sodium-dependent bile acid transporter (ASBT). In NTCP-HEK293 cells, the ability of these compounds to block the HBV/HDV-derived preS1-peptide binding to NTCP (virus receptor function) as well as the taurocholic acid transport via NTCP (bile acid transporter function) were analyzed in parallel. Hits were subsequently validated by performing in vitro HDV infection experiments in NTCP-HepG2 cells. The most potent compounds S985852, A000295231, and S973509 showed in vitro anti-HDV activities with IC50 values of 15, 40, and 70 µM, respectively, while the taurocholic acid uptake inhibition occurred at much higher IC50 values of 24, 780, and 490 µM, respectively. In conclusion, repurposing of compounds from the BARI class as novel HBV/HDV entry inhibitors seems possible and even enables certain virus selectivity based on structure-activity relationships.

Identification of Triterpene Acids in Poria cocos Extract as Bile Acid Uptake Transporter Inhibitors.

Literature reports that Poria cocos reduces blood lipid levels; however, the underlying mechanism remains unclear. Blood lipid levels are closely related to the enterohepatic circulation of bile acids, where uptake transporters playing a significant role. P. cocos extract is commonly used in traditional prescriptions and food supplements in China. We investigated the effects of P. cocos and its five triterpene acids on bile acid uptake transporters, including intestinal apical sodium-dependent bile acid transporter (ASBT) and hepatic sodium/taurocholate cotransporting polypeptide (NTCP). Triterpene acids were fingerprinted by high-performance liquid chromatography-TripleTOF and quantified by ultraperformance liquid chromatography/tandem mass spectrometry. The inhibitory effect of P. cocos and its five major representative triterpene acids on ASBT and NTCP was investigated by in vitro assays using Xenopus oocytes expressing ASBT and NTCP. P. cocos extract exhibited significant inhibitory effects with half-maximum inhibition constants of 5.89 µg/ml and 14.6 µg/ml for NTCP and ASBT, respectively. Among five triterpene acids, poricoic acid A, poricoic acid B, and polyporenic acid C significantly inhibited NTCP function. Poricoic acid A, poricoic acid B, and dehydrotumulosic acid significantly inhibited ASBT function. The representative triterpene acid, poricoic acid A, was identified as a competitive inhibitor of NTCP with an inhibitory constant of 63.4 ± 18.7 µM. In conclusion, our results indicate that both P. cocos extract and its major triterpenes are competitive inhibitors of ASBT and NTCP. Accordingly, it was suggested that competitive inhibition of these bile acid transporters is one of the underlying mechanisms for the hypolipidemic effect of P. cocos. SIGNIFICANCE STATEMENT: Poria cocos, a commonly used Chinese herbal medicine and food supplement, demonstrates significantly inhibitory effects on the function of apical sodium-dependent bile acid transporter and sodium/taurocholate cotransporting polypeptide. P. cocos has potential to reduce the blood lipid through inhibition of these uptake transporters in enterohepatic circulation of bile acid.

MECHANISMS IN ENDOCRINOLOGY: FXR signalling: a novel target in metabolic diseases.

During the last decades, it has become clear that the gastrointestinal tract plays a pivotal role in the regulation of glucose homeostasis. More than 40 hormones originate from the gastrointestinal tract and several of these impact glucose metabolism and appetite regulation. An astonishing example of the gut's integrative role in glucose metabolism originates from investigations into bile acid biology. From primary animal studies, it has become clear that bile acids should no longer be labelled as simple detergents necessary for lipid digestion and absorption but should also be recognised as metabolic regulators implicated in lipid, glucose and energy metabolism. The nuclear farnesoid X receptor (FXR) is a part of an exquisite bile acid-sensing system that among other things ensures the optimal size of the bile acid pool. In addition, intestinal and hepatic FXR also impact the regulation of several metabolic processes such as glucose and lipid metabolism. Accordingly, natural and synthetic FXR agonists and certain FXR-regulated factors (i.e. fibroblast growth factor 19 (FGF19)) are increasingly being evaluated as treatments for metabolic diseases such as type 2 diabetes and non-alcoholic fatty liver disease (and its inflammatory version, non-alcoholic steatohepatitis). Interestingly, decreased FXR activation also benefits glucose metabolism. This can be obtained by reducing bile acid absorption using bile acid sequestering agents (approved for the treatment of type 2 diabetes) or inhibitors of intestinal bile acid transporters,that is the apical sodium-dependent bile acid transporter (ASBT). This article discusses recent clinical trials that provide insights about the role of FXR-FGF19-targetted therapy for the treatment of metabolic diseases.

S-acylation status of bile acid transporter hASBT regulates its function, metabolic stability, membrane expression, and phosphorylation state

The human apical sodium-dependent bile acid transporter (hASBT, SLC10A2) is the rate-limiting step of intestinal bile acid absorption in the enterohepatic circulation system of bile acids. Therefore, the regulation and stability of hASBT is vital in maintaining bile acid and cholesterol homeostasis and may serve as a potential target for cholesterol-related disorders. We hypothesized that post-translational mechanisms that govern hASBT function and regulation will provide novel insight on intestinal bile acid transport and homeostasis. In this study, we confirm the S-acylation status of hASBT via acyl biotin exchange in COS-1 cells and its impact on hASBT expression, function, kinetics, and protein stability. Using the acylation inhibitor, 2-bromopalmitate, we show that S-acylation is an important modification which modulates the function, surface expression, and maximal transporter flux (Jmax) of hASBT. By means of proteasome inhibitors, S-acylated hASBT was found to be cleared via the proteasome whereas a reduction in the palmitoylation status of hASBT resulted in rapid proteolytic degradation compared to the unmodified transporter. Screening of cysteine mutants in and or near transmembrane domains, some of which are exposed to the cytosol, confirmed Cys314 to be the predominate S-acylated residue. Lastly, we show that S-acylation was reduced in a mutant form of hASBT devoid of cytosolic facing tyrosine residues, suggestive of crosstalk between acylation and phosphorylation post-translational modification mechanisms.

TGR5 signaling mitigates parenteral nutrition-associated liver disease.

Bile acid receptors regulate the metabolic and immune functions of circulating enterohepatic bile acids. This process is disrupted by administration of parenteral nutrition (PN), which may induce progressive hepatic injury for unclear reasons, especially in the newborn, leading to PN-associated liver disease. To explore the role of bile acid signaling on neonatal hepatic function, we initially observed that Takeda G protein receptor 5 (TGR5)-specific bile acids were negatively correlated with worsening clinical disease markers in the plasma of human newborns with prolonged PN exposure. To test our resulting hypothesis that TGR5 regulates critical liver functions to PN exposure, we used TGR5 receptor deficient mice (TGR5-/-). We observed PN significantly increased liver weight, cholestasis, and serum hepatic stress enzymes in TGR5-/- mice compared with controls. Mechanistically, PN reduced bile acid synthesis genes in TGR5-/-. Serum bile acid composition revealed that PN increased unconjugated primary bile acids and secondary bile acids in TGR5-/- mice, while increasing conjugated primary bile acid levels in TGR5-competent mice. Simultaneously, PN elevated hepatic IL-6 expression and infiltrating macrophages in TGR5-/- mice. However, the gut microbiota of TGR5-/- mice compared with WT mice following PN administration displayed highly elevated levels of Bacteroides and Parabacteroides, and possibly responsible for the elevated levels of secondary bile acids in TGR5-/- animals. Intestinal bile acid transporters expression was unchanged. Collectively, this suggests TGR5 signaling specifically regulates fundamental aspects of liver bile acid homeostasis during exposure to PN. Loss of TGR5 is associated with biochemical evidence of cholestasis in both humans and mice on PN.NEW & NOTEWORTHY Parenteral nutrition is associated with deleterious metabolic outcomes in patients with prolonged exposure. Here, we demonstrate that accelerated cholestasis and parental nutrition-associated liver disease (PNALD) may be associated with deficiency of Takeda G protein receptor 5 (TGR5) signaling. The microbiome is responsible for production of secondary bile acids that signal through TGR5. Therefore, collectively, these data support the hypothesis that a lack of established microbiome in early life or under prolonged parenteral nutrition may underpin disease development and PNALD.

Efficient reabsorption of transintestinally excreted cholesterol is a strong determinant for cholesterol disposal in mice

Transintestinal cholesterol excretion (TICE) is a major route for eliminating cholesterol from the body and a potential therapeutic target for hypercholesterolemia. The underlying mechanism, however, is largely unclear, and its contribution to cholesterol disposal from the body is obscured by the counteracting process of intestinal cholesterol reabsorption. To determine the quantity of TICE independent from its reabsorption, we studied two models of decreased intestinal cholesterol absorption. Cholesterol absorption was inhibited either by ezetimibe or, indirectly, by the genetic inactivation of the intestinal apical sodium-dependent bile acid transporter (ASBT; SLC10A2). Both ezetimibe treatment and Asbt inactivation virtually abrogated fractional cholesterol absorption (from 46% to 4% and 6%, respectively). In both models, fecal neutral sterol excretion and net intestinal cholesterol balance were considerably higher than in control mice (5- and 7-fold, respectively), suggesting that, under physiological conditions, TICE is largely reabsorbed. In addition, the net intestinal cholesterol balance was increased to a similar extent but was not further increased when the models were combined, suggesting that the effect on cholesterol reabsorption was already maximal under either condition alone. On the basis of these findings, we hypothesize that the inhibition of cholesterol (re)absorption combined with stimulating TICE will be most effective in increasing cholesterol disposal.

Defective FXR-FGF15 signaling and bile acid homeostasis in cystic fibrosis mice can be restored by the laxative polyethylene glycol.

The gastrointestinal phenotype of cystic fibrosis (CF) features intestinal bile acid (BA) malabsorption, impaired intestinal farnesoid X receptor (FXR) activation, and consequently reduced fibroblast growth factor 19 (FGF19, FGF15 in mice) production. The osmotic laxative polyethylene glycol (PEG) has been shown to decrease intestinal mucus accumulation in CF mice and could, by doing so, improve BA reabsorption. Here we determined the effect of PEG on BA excretion and FXR-FGF15 signaling in CF mice. Male Cftr-/-tm1Unc (CF) and wild-type (WT) littermates were administered PEG 4000 in drinking water and fed either chow or a semisynthetic diet. PEG was withdrawn for 3 days before termination. Fecal BA excretion was measured at PEG dosages of 37 g/l (100%) and 0 g/l (0%). Ileal FXR activation was assessed by gene expression of its downstream targets Fgf15 and small heterodimer partner ( Shp). In CF mice, PEG withdrawal increased fecal BA excretion on either diet compared with full PEG dosage (chow, 2-fold, P = 0.06; semisynthetic, 4.4-fold, P = 0.007). PEG withdrawal did not affect fecal BA excretion in WT mice on either diet. After PEG withdrawal, gene expression levels of intestinal FXR target genes Fgf15 and Shp were decreased in CF mice but unaffected in WT littermates. PEG did not affect the gene expression of the main intestinal BA transporter apical sodium-dependent bile acid transporter (ASBT). PEG treatment ameliorates intestinal BA malabsorption in CF mice and restores intestinal FXR-FGF15 signaling, independent from Asbt gene expression. These findings highlight the potential of PEG in the prevention and treatment of the gastrointestinal phenotype of CF. NEW & NOTEWORTHY A gastrointestinal feature of cystic fibrosis is bile acid malabsorption and consequent impairment of farnesoid X receptor (FXR)-fibroblast growth factor 15 (FGF15) signaling. FXR-FGF15 signaling regulates various metabolic processes and could be implicated in metabolic and gastrointestinal complications of cystic fibrosis, such as diabetes and liver disease. In cystic fibrosis mice, treatment with the osmotic laxative polyethylene glycol is associated with decreased fecal bile acid loss and restoration of FXR-FGF15 signaling.

Placebo-Controlled Randomized Trial of an Intestinal Bile Salt Transport Inhibitor for Pruritus in Alagille Syndrome.

Medically refractory, severe, cholestasis‐induced pruritus in Alagille syndrome may be improved by surgical interruption of the enterohepatic circulation. This multicenter trial (NCT02057692) tested the hypothesis that the intestinal bile acid transport inhibitor maralixibat would similarly reduce pruritus in Alagille syndrome. Thirty‐seven children with Alagille syndrome were randomly assigned to double‐blinded administration of placebo, 70, 140, or 280 µg/kg/day of maralixibat for 13 weeks. Pruritus was assessed by caregiver (itch‐reported outcome instrument [ItchRO]) and clinician report (range, 0‐4 [severe]). Liver chemistries and serum bile acids were measured. The primary outcome was the change from baseline to week 13 in ItchRO relative to placebo. In the a priori first analysis of the primary efficacy endpoint, the mean adjusted difference between participants receiving 140 or 280 µg/kg/day and placebo was –0.47 (95% confidence interval [CI], –1.14, 0.20; P = 0.16). Statistically significant decreases were observed with doses of 70 and 140 µg/kg/day (mean adjusted difference, –0.89; 95% CI, –1.70, –0.08; P = 0.032; and mean adjusted difference, –0.91; 95% CI, –1.62, –0.19; P = 0.014) but not 280 µg/kg/day (mean adjusted difference, –0.04; 95% CI, –0.94, 0.86; P = 0.44) or all doses combined (mean adjusted difference, –0.61; 95% CI, –1.24, 0.20; P = 0.055). A 1‐point reduction in pruritus was more common in maralixibat‐treated versus placebo‐treated participants (caregiver ItchRO, 65% versus 25%; P = 0.06; clinician score, 76% versus 25%; P = 0.01). There were no significant changes in liver chemistries or bile acids relative to placebo. Adverse and serious adverse events were similar between maralixibat and placebo. Conclusion: Although the prespecified primary analyses of ItchRO were not all statistically significant, the data suggest that maralixibat is safe and may reduce pruritus in Alagille syndrome.

Effects of dietary pectin on the profile and transport of intestinal bile acids in young pigs.

Pectin has been known to lower circulating cholesterol by interacting with bile acid (BA) metabolism. The current study was aimed to investigate intestinal BA transport at the molecular level in a pig model. Twelve young pigs (11.05 ± 0.11 kg) were randomly divided into 2 groups and fed corn-soybean meal diets with either 5% pectin or cornstarch for 72 d. In pigs fed with pectin, total cholesterol and low-density lipoprotein cholesterol (LDL-C) were lowered but high-density lipoprotein (HDL-C) was increased (P < 0.05). Serum triglycerides tended to be lower in the pectin-fed animals (P = 0.093), whereas no change was noted in serum total bile acid. Along the length of the intestine, the size and composition of BA pools vary. The ratio of primary, secondary, taurine-conjugated, and glycine-conjugated BAs in the ileal pool was about 46:15:9:30, whereas it was 28:61:1:11 in the cecum and 22:65:3:9 in the colon (P < 0.05). In the feces, lithocholic acid and ursodeoxycholic acid (UDCA) made up of over 97% of the total BA pool. Overall, the ileum had the greatest expression of farnesoid X receptor (FXR) and apical sodium-coupled bile acid transporter (ASBT) than the duodenum, jejunum, cecum, and colon (P < 0.05), whereas organic solute transporters α/β (OSTα/β) gene expression was peaked in the ileum and jejunum (P < 0.05). Expression multidrug resistance protein 2 (MRP2) gradually decreased towards the end of the intestine (P < 0.05). Greater expression of G protein-coupled bile acid receptor and multidrug resistance protein 3 (MRP3) was found in the cecum and colon (P < 0.05). In pigs fed with 5% pectin, only cecal UDCA (P = 0.097) and hyocholic acid (P = 0.088) showed a decreasing tendency. But FXR, ASBT, and MRP2 were upregulated in the ileum and FXR, OSTα/β, MRP2, and MRP3 in the cecum of PEC-fed pigs (P < 0.05). Liver enzymes involved in BA biosynthesis (CYP7A1, CYP27A1, bile acid-CoA synthase, and bile acid-CoA:amino acid N acyltransferase) were not affected by pectin consumption. In conclusion, the abundant distribution of BA transporters and the greater BA pool size suggests the ileum as the major site for intestinal BA reabsorption in pigs. In the ileum, pectin increased in-and-out BA transport on the apical membrane by increasing ASBT and MRP2, but it increased the overall BA transport in the cecum by increasing OSTα/β and MRP3.

Continued Alcohol Misuse in Human Cirrhosis is Associated with an Impaired Gut-Liver Axis.

Cirrhosis and alcohol can independently affect the gut-liver axis with systemic inflammation. However, their concurrent impact in humans is unclear. 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. 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. 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.

Editor's Highlight: Clofibrate Decreases Bile Acids in Livers of Male Mice by Increasing Biliary Bile Acid Excretion in a PPARα-Dependent Manner.

Fibrates and their receptor, namely peroxisome proliferator-activated receptor α (PPARα), have been reported to regulate bile acid (BA) synthesis and transport. However, the effect of fibrate treatment and PPARα activation on BA homeostasis remains controversial. In this study, both wild-type (WT) and PPARα-null male mice were treated with clofibrate (CLOF) for 4 days to evaluate the effects of short-term PPARα activation on BA homeostasis. Although a decrease in total BAs (ΣBAs) was observed in livers of CLOF-treated WT mice, it was not observed in PPARα-null mice. CLOF-mediated decrease in ΣBAs in the liver was not likely due to the reduction in BA synthesis or BA uptake, as evidenced by an increase in the BA synthetic enzyme (Cyp7a1) and 2 BA uptake transporters (Na (+)-taurocholate cotransporting polypeptide [Ntcp] and organic anion transporting polypeptide [Oatp]1b2). Instead, the decrease in liver BAs by CLOF is largely a result of increased biliary excretion of BAs, which was associated with a significant induction of the canalicular efflux transporter (bile salt export pump [Bsep]) in the liver. The PPARα-mediated increase in Cyp7a1 in CLOF-treated WT mice was not due to farnesoid X receptor (Fxr)-small heterodimer partner (Shp) signaling in the liver, but due to suppression of Fxr- fibroblast growth factor15 signaling in the ileum. Additionally, CLOF also suppressed intestinal BA transporters (apical sodium-dependent bile acid transporter and organic solute transporterβ) and cholesterol efflux transporters (Abcg5 and Abcg8) in a PPARα-dependent manner. In summary, this study provides the first comprehensive analysis on the effect of a short-term CLOF treatment on BA homeostasis, and revealed an essential role of PPARα in regulating BA synthesis, transport and signaling.