Biliary Secretion Research Articles

Cystic fibrosis rabbits develop spontaneous hepatobiliary lesions and CF-associated liver disease (CFLD)-like phenotypes.

Cystic fibrosis (CF) is an autosomal recessive genetic disease affecting multiple organs. Approximately 30% CF patients develop CF-related liver disease (CFLD), which is the third most common cause of morbidity and mortality of CF. CFLD is progressive, and many of the severe forms eventually need liver transplantation. The mechanistic studies and therapeutic interventions to CFLD are unfortunately very limited. Utilizing the CRISPR/Cas9 technology, we recently generated CF rabbits by introducing mutations to the rabbit CF transmembrane conductance regulator (CFTR) gene. Here we report the liver phenotypes and mechanistic insights into the liver pathogenesis in these animals. CF rabbits develop spontaneous hepatobiliary lesions and abnormal biliary secretion accompanied with altered bile acid profiles. They exhibit nonalcoholic steatohepatitis (NASH)-like phenotypes, characterized by hepatic inflammation, steatosis, and fibrosis, as well as altered lipid profiles and diminished glycogen storage. Mechanistically, our data reveal that multiple stress-induced metabolic regulators involved in hepatic lipid homeostasis were up-regulated in the livers of CF-rabbits, and that endoplasmic reticulum (ER) stress response mediated through IRE1α-XBP1 axis as well as NF-κB- and JNK-mediated inflammatory responses prevail in CF rabbit livers. These findings show that CF rabbits manifest many CFLD-like phenotypes and suggest targeting hepatic ER stress and inflammatory pathways for potential CFLD treatment.

Gallstones: The thing in itself.

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S2901 Afferent Limb Syndrome Treated With Non-Cautery-Enhanced Lumen Apposing Metal Stent: An Alternate Approach

Introduction: Afferent limb syndrome (ALS) is a known complication after gastrojejunostomies from mechanical obstruction of the afferent limb (AL). Recently endoscopic ultrasound (EUS) guided gastroenterostomy placement has provided an alternate to surgical correction. However this approach is not always technically possible. We describe a case successfully treated with endoluminal non-cautery enhanced lumen apposing metal stent (LAMS) placement. Case Description/Methods: A 63 year old woman with a history of ileocolonic Crohns disease (diagnosed at age 25), status post multiple surgeries including ileocolic resections and Billroth II gastrectomy, was referred with persistent abdominal pain. She was recently diagnosed with acute pancreatitis with CT concerning for dilated afferent jejunal limb, a luminal stricture near the biliary anastomosis and dilated intra and extra hepatic bile ducts (Figure a). She underwent two deep enteroscopies revealing a stricture in the AL from intra-abdominal adhesion. This was dilated with minimal improvement in symptoms and imaging. After multi-disciplinary discussion it was decided to attempt EUS guided drainage of the AL to promote drainage of pancreatico- biliary secretions. During the procedure a significant amount of food residue was identified in the gastric remnant. The dilated afferent limb was visualized endosonographically adjacent to the posterior stomach at an acute angle. Considering large amount of food in the stomach, difficult angulation to deploy the AXIOS stent and concern for food entering the already obstructed limb after stent deployment it was decided to reconsider this approach. The AL was then explored with a therapeutic upper endoscope with stenosis at 10 cm from the anastomosis. A wire was passed through the stenosis under fluoroscopy. It was decided to place a stent using the 15 x 10 mm LAMS stent system considering short (< 1 cm) stricture. Using non cautery technique both flanges were deployed successfully across the stricture under fluoroscopic and endoscopic guidance with decompression of the AL (Figure b,c). 1 month follow up imaging demonstrated continued decompression of the AL. Discussion: EUS guided gastrojejunostomy has been increasingly used for ALS. However it can be challenging in situations where a good window cannot be identified. In such settings using a traditional luminal route to reach the stenosis and placing a non-cautery enhanced LAMS can provide a viable alternative.Figure 1.: (a) Dilated afferent limb with stricture (arrows), (b) post non-cautery enhanced LAMS deployment, (c) Fluoroscopy image post LAMS deployment

S1907 A Rare Case of Type 1 Choledochal Cyst With Type C Anomalous Pancreaticobiliary Maljunction in a 52-Year-Old Patient

Introduction: Choledochal cysts are rare anomalies of the bile duct that generally present in childhood but can rarely be delayed until adulthood in 20% of cases. They are often diagnosed incidentally or present with acute pancreatitis or cholangitis. We report a case of a type 1 choledochal cyst (T1CC) and type C anomalous pancreaticobiliary maljunction (PBM), successfully managed by a multidisciplinary approach. Case Description/Methods: A 52-year-old woman presented with right upper quadrant pain, jaundice and vomiting. Labs revealed elevated lipase, bilirubin, AST, ALT and alkaline phosphatase. A CT of the abdomen showed no gallstones but a marked dilation of extrahepatic bile duct with a transition point in the distal intrapancreatic portion (Figure a). An endoscopic retrograde cholangiopancreaticogram (ERCP) showed fusiform dilation of the CBD consistent with T1CC (Figure b). Spyglass cholangioscopy revealed focal stenosis in the distal CBD joining above the ventral pancreatic duct opening within the long common channel, suggestive of type C anomalous PBM (Figure c). A plastic biliary stent was placed into the CBD. EUS guided fine needle biopsy of the stricture showed no evidence of malignancy. Her clinical course was complicated by multiple admissions due to stent occlusion. After multidisciplinary discussion, the patient underwent choledochal cyst resection with hepaticojejunostomy, cholecystectomy and biliary reconstruction. Intraoperative gallbladder specimen revealed poorly differentiated adenocarcinoma with plan for chemotherapy. Discussion: Choledochal cysts are a rare congenital anomaly characterized by enlargement of the CBD. There are five types of choledochal cysts (Table). Etiology remains unclear and while some believe these cysts are congenital in nature, others propose they arise from anomalous PBM. PBM is defined as congenital malformation in which the pancreatic and bile duct join outside the duodenal wall forming a long common channel. This leads to reflux of biliary secretions leading to chronic inflammation, increasing the risk of developing cholangiocarcinoma and gallbladder cancer, as seen in our patient. Placement of biliary stents can temporarily alleviate symptoms, but surgical excision is the definitive treatment. Our case highlights the importance of including type 1 choledochal cyst in the differential diagnosis even in adult patients with a significantly dilated CBD. A thorough endoscopic and radiological work up should be done to prevent progression to malignancy.Figure 1.: A. CT of the abdomen showing marked dilatation of extrahepatic bile duct with a transition point in the distal intrapancreatic portion. B. Cholangiogram showing Type 1 Choledochal Cyst and Type C Anomalous Pancreaticobiliary Maljunction. C. Spyglass cholangioscopy showing JagwireTM within the CBD (yellow arrow) revealing focal stenotic CBD joining the common channel just above the ventral pancreatic duct (blue arrow). Table 1. - Types of Choledochal Cysts Type Description Type I Fusiform dilation of the CBD Type II Bile duct diverticulum Type III Saccular dilation of the intraduodenal portion of the CBD Type IVa Multiple cysts at intra and extrahepatic ducts Type IVb Multiple cysts at extrahepatic ducts only Type V Fusiform or saccular cystic dilation of the intrahepatic bile duct

S3221 Adult Onset Progressive Familial Intrahepatic Cholestasis: A Rare Presentation

Introduction: Progressive familial intrahepatic cholestasis (PFIC) is a heterogeneous group of autosomal recessive childhood genetic disorders that disrupt intrahepatic biliary secretion. PFIC accounts for 10 to 15% of children's cholestatic diseases and liver transplantation. Although all patients will require liver transplantation at a certain point, early diagnosis and intervention can delay complications and transplantation. Case Description/Methods: A 41-year-old male presented with cryptogenic cirrhosis and a MELD score of 35 for transplantation evaluation. He had a childhood history of abnormal liver enzymes with a mixed pattern and cholecystectomy due to recurrent gallstones. Prior liver biopsy showed autoimmune features; however, there was no significant response to the conventional therapy with prednisone and azathioprine. However, a re-evaluation of his liver biopsy favored biliary disease: subsequent MRCP with normal bile ducts. Given the insidious onset of his cirrhosis without an identifiable etiology and the suggestion of biliary cause, PFIC was considered during pre-transplantation workup. Genetic testing for ABCB4 confirmed PFIC type 3; subsequently, he underwent liver transplantation with an unremarkable post-operative course. Discussion: This case highlights the impact of exploring a diagnosis of PFIC in adults referred for liver transplantation with long-standing cholestatic liver disease and negative initial workup. While currently, liver transplantation is the primary treatment option for end-stage liver disease from PFIC, advances in gene therapy have led to potentially curative treatments. As such, improved provider awareness and having a lower threshold for diagnostic testing may lead to an earlier diagnosis which is critical for optimal management, delaying the onset of end-stage liver disease, and offering to screen for family members.

In Vitro Rescue of the Bile Acid Transport Function of ABCB11 Variants by CFTR Potentiators

ABCB11 is responsible for biliary bile acid secretion at the canalicular membrane of hepatocytes. Variations in the ABCB11 gene cause a spectrum of rare liver diseases. The most severe form is progressive familial intrahepatic cholestasis type 2 (PFIC2). Current medical treatments have limited efficacy. Here, we report the in vitro study of Abcb11 missense variants identified in PFIC2 patients and their functional rescue using cystic fibrosis transmembrane conductance regulator potentiators. Three ABCB11 disease-causing variations identified in PFIC2 patients (i.e., A257V, T463I and G562D) were reproduced in a plasmid encoding an Abcb11-green fluorescent protein. After transfection, the expression and localization of the variants were studied in HepG2 cells. Taurocholate transport activity and the effect of potentiators were studied in Madin–Darby canine kidney (MDCK) clones coexpressing Abcb11 and the sodium taurocholate cotransporting polypeptide (Ntcp/Slc10A1). As predicted using three-dimensional structure analysis, the three variants were expressed at the canalicular membrane but showed a defective function. Ivacaftor, GLP1837, SBC040 and SBC219 potentiators increased the bile acid transport of A257V and T463I and to a lesser extent, of G562D Abcb11 missense variants. In addition, a synergic effect was observed when ivacaftor was combined with SBC040 or SBC219. Such potentiators could represent new pharmacological approaches for improving the condition of patients with ABCB11 deficiency due to missense variations affecting the function of the transporter.

A missense mutation in the proprotein convertase gene furinb causes hepatic cystogenesis during liver development in zebrafish.

Hepatic cysts are fluid‐filled lesions in the liver that are estimated to occur in 5% of the population. They may cause hepatomegaly and abdominal pain. Progression to secondary fibrosis, cirrhosis, or cholangiocarcinoma can lead to morbidity and mortality. Previous studies of patients and rodent models have associated hepatic cyst formation with increased proliferation and fluid secretion in cholangiocytes, which are partially due to impaired primary cilia. Congenital hepatic cysts are thought to originate from faulty bile duct development, but the underlying mechanisms are not fully understood. In a forward genetic screen, we identified a zebrafish mutant that developed hepatic cysts during larval stages. The cyst formation was not due to changes in biliary cell proliferation, bile secretion, or impairment of primary cilia. Instead, time‐lapse live imaging data showed that the mutant biliary cells failed to form interconnecting bile ducts because of defects in motility and protrusive activity. Accordingly, immunostaining revealed a disorganized actin and microtubule cytoskeleton in the mutant biliary cells. By whole‐genome sequencing, we determined that the cystic phenotype in the mutant was caused by a missense mutation in the furinb gene, which encodes a proprotein convertase. The mutation altered Furinb localization and caused endoplasmic reticulum (ER) stress. The cystic phenotype could be suppressed by treatment with the ER stress inhibitor 4‐phenylbutyric acid and exacerbated by treatment with the ER stress inducer tunicamycin. The mutant liver also exhibited increased mammalian target of rapamycin (mTOR) signaling. Treatment with mTOR inhibitors halted cyst formation at least partially through reducing ER stress. Conclusion: Our study has established a vertebrate model for studying hepatic cystogenesis and illustrated the contribution of ER stress in the disease pathogenesis.

Intrahepatic and Extrahepatic Cholestatic Jaundice Caused by Adenocarcinoma Pancreas with Hepatitis B Infection: A Case Report

Background: Cholestasis is defined as the stagnation of biliary duct secretion. Intrahepatic jaundice cholestasis is caused by inflammation that happens intrahepatic from an infection such as hepatitis. Meanwhile, extrahepatic cholestasis is caused by obstruction of the biliary duct outside the liver. Stagnation of the bile flow due to these couple of conditions occurring concurrently has been rarely reported. Biliary duct stagnation that occurs with an infection of hepatocytes will cause more serious and progressive effects.&#x0D; Case presentation: A 73 years old woman with extrahepatic and intrahepatic cholestasis caused by pancreatic adenocarcinoma along with chronic hepatitis B. The diagnosis is made based on clinical findings, lab findings, and radiology findings. Quick action is mandatory to prevent complications caused by hyperbilirubinemia and slows the progression of hepatitis B-induced liver fibrosis. A Whipple procedure was performed on this patient to treat the biliary duct obstruction. The patient's condition improved after surgery. The patient is then planned to be given adjuvant chemotherapy, preceded by the administration of antivirals for hepatitis.&#x0D; Conclusion: Cholestasis jaundice causes hyperbilirubinemia that presents with clinical complaints such as jaundice, itchy and could cause several complications due to damage to the liver tissue whose clinical manifestations are related to impaired liver function caused by hyperbilirubinemia in the form of hypoalbumin, hypercoagulation, and in advanced stages can end up with liver cirrhosis. The principle management for cholestatic jaundice is bile duct decompression which is expected to rule out the etiology of the blockage. Tenofovir was given before chemotherapy in these patients to prevent hepatitis flare due to hepatitis B.

Identification of the chloride channel, leucine-rich repeat-containing protein 8, subfamily a (LRRC8A), in mouse cholangiocytes.

Chloride (Cl- ) channels in the apical membrane of biliary epithelial cells (BECs), also known as cholangiocytes, provide the driving force for biliary secretion. Although two Cl- channels have been identified on a molecular basis, the Cystic Fibrosis Transmembrane Conductance Regulator and Transmembrane Member 16A, a third Cl- channel with unique biophysical properties has been described. Leucine-Rich Repeat-Containing Protein 8, subfamily A (LRRC8A) is a newly identified protein capable of transporting Cl- in other epithelium in response to cell swelling. The aim of the present study was to determine if LRRC8A represents the volume-regulated anion channel in mouse BECs. Studies were performed in mouse small (MSC) and large (MLC) cholangiocytes. Membrane Cl- currents were measured by whole-cell patch-clamp techniques and cell volume measurements were performed by calcein-AM fluorescence. Exposure of either MSC or MLC to hypotonicity (190 mOsm) rapidly increased cell volume and activated Cl- currents. Currents exhibited outward rectification, time-dependent inactivation at positive membrane potentials, and reversal potential at 0 mV (ECl ). Removal of extracellular Cl- or specific pharmacological inhibition of LRRC8A abolished currents. LRRC8A was detected in both MSC and MLC by reverse transcription polymerase chain reaction and confirmed by western blot. Transfection with LRRC8A small interfering RNA decreased protein levels by >70% and abolished volume-stimulated Cl- currents. These results demonstrate that LRRC8A is functionally present in mouse BECs, contributes to volume-activated Cl- secretion, and, therefore, may be a target to modulate bile formation in the treatment of cholestatic liver disorders.

The Effects of Taurocholic Acid on Biliary Damage and Liver Fibrosis Are Mediated by Calcitonin-Gene-Related Peptide Signaling.

Background & aims: Cholangiocytes are the target cells of liver diseases that are characterized by biliary senescence (evidenced by enhanced levels of senescence-associated secretory phenotype, SASP, e.g., TGF-β1), and liver inflammation and fibrosis accompanied by altered bile acid (BA) homeostasis. Taurocholic acid (TC) stimulates biliary hyperplasia by activation of 3′,5′-cyclic cyclic adenosine monophosphate (cAMP) signaling, thereby preventing biliary damage (caused by cholinergic/adrenergic denervation) through enhanced liver angiogenesis. Also: (i) α-calcitonin gene-related peptide (α-CGRP, which activates the calcitonin receptor-like receptor, CRLR), stimulates biliary proliferation/senescence and liver fibrosis by enhanced biliary secretion of SASPs; and (ii) knock-out of α-CGRP reduces these phenotypes by decreased cAMP levels in cholestatic models. We aimed to demonstrate that TC effects on liver phenotypes are dependent on changes in the α-CGRP/CALCRL/cAMP/PKA/ERK1/2/TGF-β1/VEGF axis. Methods: Wild-type and α-CGRP−/− mice were fed with a control (BAC) or TC diet for 1 or 2 wk. We measured: (i) CGRP levels by both ELISA kits in serum and by qPCR in isolated cholangiocytes (CALCA gene for α-CGRP); (ii) CALCRL immunoreactivity by immunohistochemistry (IHC) in liver sections; (iii) liver histology, intrahepatic biliary mass, biliary senescence (by β-GAL staining and double immunofluorescence (IF) for p16/CK19), and liver fibrosis (by Red Sirius staining and double IF for collagen/CK19 in liver sections), as well as by qPCR for senescence markers in isolated cholangiocytes; and (iv) phosphorylation of PKA/ERK1/2, immunoreactivity of TGF-β1/TGF- βRI and angiogenic factors by IHC/immunofluorescence in liver sections and qPCR in isolated cholangiocytes. We measured changes in BA composition in total liver by liquid chromatography/mass spectrometry. Results: TC feeding increased CALCA expression, biliary damage, and liver inflammation and fibrosis, as well as phenotypes that were associated with enhanced immunoreactivity of the PKA/ERK1/2/TGF-β1/TGF-βRI/VEGF axis compared to BAC-fed mice and phenotypes that were reversed in α-CGRP−/− mice fed TC coupled with changes in hepatic BA composition. Conclusion: Modulation of the TC/ α-CGRP/CALCRL/PKA/ERK1/2/TGF-β1/VEGF axis may be important in the management of cholangiopathies characterized by BA accumulation.

Hepatitis A: Current view of an ancient disease

Hepatitis A: Current view of an ancient disease

Abstract 439: Abcg5 Abcg8 Are Essential To Maintain Sterol Balance When Challenged With Dietary Cholesterol In Mice

Background: ABCG5/ABCG8 (G5/G8) is the primary biliary cholesterol transporter and accounts for &gt;70% of biliary cholesterol secretion. Mice and humans maintain fecal cholesterol excretion under a variety of conditions that disrupt biliary cholesterol secretion, indicating the presence of a non-biliary route for cholesterol elimination. Cholesterol can be actively secreted from the intestine, a pathway termed transintestinal cholesterol excretion (TICE). The present study investigates rates of intestinal cholesterol secretion in mice lacking G5/G8 to determine if adaptive responses within the gut facilitate cholesterol excretion in the absence of the primary biliary cholesterol transporter. Methods: Wild-type (WT) and whole body Abcg5/Abcg8 deficient mice (G5/G8 KO) were maintained on a plant sterol free (PSF) diet to prevent the development of experimental sitosterolemia and potential confounding effects of biologically active phytosterols. Mice were anesthetized, the gall bladder cannulated, and the intestine perfused with model bile to simultaneously determine biliary and intestinal cholesterol secretion rates. Sterol balance studies were conducted in mice sequentially maintained on PSF diet, a PSF diet supplemented with 0.2% cholesterol, and standard rodent chow. Mice were allowed to acclimate to each diet for 14 days, then placed on wire-bottom cages for fecal collection. At termination of the experiment, basal bile, plasma, and tissues were collected. Fecal neutral sterols (FNS) were analyzed by GC-MS. Results: Despite a substantial reduction in biliary cholesterol secretion in G5/G8 KO mice, differences in intestinal cholesterol secretion rates were only observed in males, with KO mice excreting less cholesterol into the intestinal lumen. FNS did not differ between genotypes when maintained on the PSF diet. On chow diet, FNS increased 2x in both genotypes, but did not differ across genotypes. When PSF diet was supplemented with 0.2% cholesterol, FNS increased by 3x in WT mice, but were unchanged in G5/G8 KO mice, relative to PSF diet levels. Conclusions: Abcg5/Abcg8 is essential to maintain sterol balance when cholesterol is present in the diet. TICE is unable to compensate for Abcg5/Abcg8 deficiency in cholesterol fed mice.

Abstract 437: Carnitine Palmitoyltransferase 1a Modulates Lipoprotein Metabolism

Background: Nonalcoholic fatty liver disease (NAFLD) affects almost 1 billion people worldwide and is associated with risk factors such as obesity and dyslipidemia. Studies show linked variants and methylation status of carnitine palmitoytransferase 1a (CPT1a) to disrupt very low-density lipoprotein (VLDL) cholesterol and triglyceride (TG) levels. We exhibit liver-specific deletion of CPT1a in mice lowers plasma cholesterol and TG while exacerbating NAFLD and inflammation. Methods: Eight-week old Cpt1a floxed mice with the human apoB100 transgene (Cpt1a fl/fl /B100 Tg ) were administered control adenoassociated virus (AAV) or AAV encoding Cre-recombinase under control of a liver specific promoter (TBG-Cre). Control and Cpt1a liver-specific knock out (LKO) mice were placed on low-fat control or western-type diet (WTD; 42% kcal fat, 0.2% cholesterol) for 16 weeks. Body weights were recorded weekly and body composition by MRI was performed at the study midpoint and end. Tissues and plasma were collected and analyzed for lipid composition and gene and protein expression by QPCR and immunoblotting, respectively. Results: CPT1a LKO mice had lower plasma cholesterol and TG irrespective of diet. The reduction in plasma cholesterol was limited to the LDL pool in FPLC-fractionated plasma. Hepatic TG was elevated in mice fed WTD and LKO Cpt1a mice. Loss of hepatic Cpt1a had no effect on hepatic cholesterol in male mice, but increased total and free cholesterol by 2- and 2.5-fold, respectively, in females. The rise in hepatic free cholesterol in female Cpt1a LKO mice associated with increases in Kupffer cell ( Clec4f ) and collagen ( Col1a1 ) gene expression. Free cholesterol levels were not related to differences in transcripts for enzymes involved in cholesterol synthesis or secretion ( Srebp2, Hmgcr, Hmgcs, Soat2, Mttp ), hepatic clearance ( Ldlr ), metabolism ( Cpy7a1, Cyp8b1 ), or biliary secretion ( Abcg5, Abcg8 ). Conclusions: Liver-specific deletion of CPT1a reduces plasma LDL-cholesterol and increases cholesterol levels in female, but not male mice. The increase in hepatic free cholesterol did not alter expression of cholesterol-responsive genes, suggesting cholesterol accumulation outside of the regulatory pool.

Abstract 528: Characterization Of Sitosterolemia-causing Mutations In The Extracellular Domain Of Abcg8

Background: ABCG5 and ABCG8 form a heterodimer that promotes biliary secretion and opposes intestinal absorption of cholesterol and phytosterols. Whole-genome and exome sequencing has revealed over 2000 variants of ABCG5 and ABCG8, many of which are predicted to be pathogenic or likely pathogenic. Missense mutations in clinically confirmed cases of Sitosterolemia are of particular interest for their potential to reveal structure-function relationships in ABCG5 ABCG8 and other ABC transporter family members. Mutations in ABCG5 ABCG8 with clinically confirmed Sitosterolemia generally cluster within the ATP binding cassette and transmembrane spanning segments that form the substrate-binding domain. However, four disease-causing mutations occur in the short extracellular loop in ABCG8, suggesting that these mutations may reveal novel elements of the ABCG5 ABCG8 function. Methods: Mutants of ABCG8 were generated by site-directed mutagenesis. Plasmids encoding normal ABCG5 and normal or mutant ABCG8s were co-transfected into human Huh7 hepatocytes, lysates prepared, and analyzed by SDS-PAGE and immunoblot analysis. Maturation and trafficking of the ABCG5 ABCG8 heterodimer beyond the endoplasmic reticulum was assessed by the appearance of the higher molecular weight, mature form of each glycoprotein. Results: Mutants G574R, G575D, and L596R failed to support trafficking of ABCG5 ABCG8 and were designated as Class II mutants causative in Sitosterolemia. Only G574E supported maturation of the ABCG5 ABCG8 transporter in cultured human hepatocytes and remains unclassified. Conclusion: Formation and trafficking of the ABCG5 ABCG8 heterodimer is dependent on key residues within the third extracellular loop in ABCG8. Maturation of ABCG5 ABCG8 is tolerant of substitutions in glycine 574, including the addition of a larger and negatively charged residue. However, neither substitution forms a functional ABCG5 ABCG8 transporter.

From Dietary Cholesterol to Blood Cholesterol, Physiological Lipid Fluxes, and Cholesterol Homeostasis.

Dietary cholesterol (C) is a major contributor to the endogenous C pool, and it affects the serum concentration of total C, particularly the low-density lipoprotein cholesterol (LDL-C). A high serum concentration of LDL-C is associated with an increased risk for atherosclerosis and cardiovascular diseases. This concentration is dependent on hepatic C metabolism creating a balance between C input (absorption and synthesis) and C elimination (conversion to bile acids and fecal excretion). The daily C absorption rate is determined by dietary C intake, biliary C secretion, direct trans-intestinal C excretion (TICE), and the fractional C absorption rate. Hepatic C metabolism coordinates C fluxes entering the liver via chylomicron remnants (CMR), LDL, high-density lipoproteins (HDL), hepatic C synthesis, and those leaving the liver via very low-density lipoproteins (VLDL), biliary secretion, and bile acid synthesis. The knowns and the unknowns of this C homeostasis are discussed.

Multidrug Resistance-Associated Protein 2 Deficiency Aggravates Estrogen-Induced Impairment of Bile Acid Metabolomics in Rats.

Multidrug resistance-associated protein 2 (Mrp2) mediates biliary secretion of anionic endobiotics and xenobiotics. Genetic alteration of Mrp2 leads to conjugated hyperbilirubinemia and predisposes to the development of intrahepatic cholestasis of pregnancy (ICP), characterized by increased plasma bile acids (BAs) due to mechanisms that are incompletely understood. Therefore, this study aimed to characterize BA metabolomics during experimental Mrp2 deficiency and ICP. ICP was modeled by ethinylestradiol (EE) administration to Mrp2-deficient (TR) rats and their wild-type (WT) controls. Spectra of BAs were analyzed in plasma, bile, and stool using an advanced liquid chromatography–mass spectrometry (LC–MS) method. Changes in BA-related genes and proteins were analyzed in the liver and intestine. Vehicle-administered TR rats demonstrated higher plasma BA concentrations consistent with reduced BA biliary secretion and increased BA efflux from hepatocytes to blood via upregulated multidrug resistance-associated protein 3 (Mrp3) and multidrug resistance-associated protein 4 (Mrp4) transporters. TR rats also showed a decrease in intestinal BA reabsorption due to reduced ileal sodium/bile acid cotransporter (Asbt) expression. Analysis of regulatory mechanisms indicated that activation of the hepatic constitutive androstane receptor (CAR)-Nuclear factor erythroid 2-related factor 2 (Nrf2) pathway by accumulating bilirubin may be responsible for changes in BA metabolomics in TR rats. Ethinylestradiol administration to TR rats further increased plasma BA concentrations as a result of reduced BA uptake and increased efflux via reduced Slco1a1 and upregulated Mrp4 transporters. These results demonstrate that Mrp2-deficient organism is more sensitive to estrogen-induced cholestasis. Inherited deficiency in Mrp2 is associated with activation of Mrp3 and Mrp4 proteins, which is further accentuated by increased estrogen. Bile acid monitoring is therefore highly desirable in pregnant women with conjugated hyperbilirubinemia for early detection of intrahepatic cholestasis.

Sexual dimorphism: increased sterol excretion leads to hypocholesterolaemia in female hyperbilirubinaemic Gunn rats.

Circulating bilirubin is associated with reduced serum cholesterol concentrations in humans and in hyperbilirubinaemic Gunn rats. However, mechanisms contributing to hypocholesterolaemia remain unknown. Therefore, this study aimed to investigate cholesterol synthesis, transport and excretion in mutant Gunn rats. Adult Gunn and control rats were assessed for daily faecal sterol excretion using metabolic cages, and water was supplemented with [1‐13C]‐acetate to determine cholesterol synthesis. Bile was collected to measure biliary lipid secretion. Serum and liver were collected for biochemical analysis and for gene/protein expression using RT‐qPCR and western blot, respectively. Additionally, serum was collected and analysed from juvenile rats. A significant interaction of sex, age and phenotype on circulating lipids was found with adult female Gunn rats reporting significantly lower cholesterol and phospholipids. Female Gunn rats also demonstrated elevated cholesterol synthesis, greater biliary lipid secretion and increased total faecal cholesterol and bile acid excretion. Furthermore, they possessed increased hepatic low‐density lipoprotein (LDL) receptor and SREBP2 expression. In contrast, there were no changes to sterol metabolism in adult male Gunn rats. This is the first study to demonstrate elevated faecal sterol excretion in female hyperbilirubinaemic Gunn rats. Increased sterol excretion creates a negative intestinal sterol balance that is compensated for by increased cholesterol synthesis and LDL receptor expression. Therefore, reduced circulating cholesterol is potentially caused by increased hepatic uptake via the LDL receptor. Future studies are required to further evaluate the sexual dimorphism of this response and whether similar findings occur in females with benign unconjugated hyperbilirubinaemia (Gilbert's syndrome).Key points Female adult hyperbilirubinaemic (Gunn) rats demonstrated lower circulating cholesterol, corroborating human studies that report a negative association between bilirubin and cholesterol concentrations.Furthermore, female Gunn rats had elevated sterol excretion creating a negative intestinal sterol balance that was compensated for by elevated cholesterol synthesis and increased hepatic low‐density lipoprotein (LDL) receptor expression.Therefore, elevated LDL receptor expression potentially leads to reduced circulating cholesterol levels in female Gunn rats providing an explanation for the hypocholesterolaemia observed in humans with elevated bilirubin levels.This study also reports a novel interaction of sex with the hyperbilirubinaemic phenotype on sterol metabolism because changes were only reported in females and not in male Gunn rats.Future studies are required to further evaluate the sexual dimorphism of this response and whether similar findings occur in females with benign unconjugated hyperbilirubinaemia (Gilbert's syndrome).

Effects of Coffee on the Gastro-Intestinal Tract: A Narrative Review and Literature Update.

The objective of the present research was to review the state of the art on the consequences of drinking coffee at the different levels of the gastrointestinal tract. At some steps of the digestive process, the effects of coffee consumption seem rather clear. This is the case for the stimulation of gastric acid secretion, the stimulation of biliary and pancreatic secretion, the reduction of gallstone risk, the stimulation of colic motility, and changes in the composition of gut microbiota. Other aspects are still controversial, such as the possibility for coffee to affect gastro-esophageal reflux, peptic ulcers, and intestinal inflammatory diseases. This review also includes a brief summary on the lack of association between coffee consumption and cancer of the different digestive organs, and points to the powerful protective effect of coffee against the risk of hepatocellular carcinoma. This review reports the available evidence on different topics and identifies the areas that would most benefit from additional studies.

Gut microbiota promotes cholesterol gallstone formation by modulating bile acid composition and biliary cholesterol secretion

Cholesterol gallstone disease is a worldwide common disease. Cholesterol supersaturation in gallbladder bile is the prerequisite for its pathogenesis, while the mechanism is not completely understood. In this study, we find enrichment of gut microbiota (especially Desulfovibrionales) in patients with gallstone disease. Fecal transplantation of gut microbiota from gallstone patients to gallstone-resistant strain of mice can induce gallstone formation. Carrying Desulfovibrionales is associated with enhanced cecal secondary bile acids production and increase of bile acid hydrophobicity facilitating intestinal cholesterol absorption. Meanwhile, the metabolic product of Desulfovibrionales, H2S increase and is shown to induce hepatic FXR and inhibit CYP7A1 expression. Mice carrying Desulfovibrionales present induction of hepatic expression of cholesterol transporters Abcg5/g8 to promote biliary secretion of cholesterol as well. Our study demonstrates the role of gut microbiota, Desulfovibrionales, as an environmental regulator contributing to gallstone formation through its influence on bile acid and cholesterol metabolism.

Melancholé: The Dark Side of Bile Acids and Its Cellular Consequences.

The first bile acid, cholic acid, was discovered in 1848 from ox gallbladder, and the bile acid structure solved by Otto Wieland was recognized with the 1927 Nobel Prize in Chemistry.1 Primary bile acids are synthesized in the liver from cholesterol and conjugated to taurine or glycine for biliary secretion. Bile acid synthesis is catalyzed by approximately 17 enzymes via the classic or alternative pathways. The classic bile acid synthesis pathway is initiated by cholesterol 7α-hydroxylase (CYP7A1), followed by sterol 12α-hydroxylase (CYP8B1).