Bile Composition Research Articles

Rapid quantification of conjugated and unconjugated bile acids and C27 precursors in dried blood spots and small volumes of serum

The aim of the study was to develop a method for fast and reliable diagnosis of peroxisomal diseases and to facilitate differential diagnosis of cholestatic hepatopathy. For the quantification of bile acids and their conjugates as well as C(27) precursors di- and trihydroxycholestanoic acid (DHCA, THCA), in small pediatric blood samples we combined HPLC separation on a reverse-phase C18 column with ESI-MS/MS analysis in the negative ion mode. Analysis was done with good precision (CV 3,7%-11.1%) and sufficient sensitivity (LOQ: 11-91 nmol/L) without derivatization. Complete analysis of 17 free and conjugated bile acids from dried blood spots and 10 microL serum samples, respectively, was performed within 12 min. Measurement of conjugated primary bile acids plus DHCA and THCA as well as ursodeoxycholic acid was done in 4.5 min. In blood spots of healthy newborns, conjugated primary bile acids were found in the range of 0.01 to 2.01 micromol/L. Concentrations of C(27) precursors were below the detection limit in normal controls. DHCA and THCA were specifically elevated in cases of peroxysomal defects and one Zellweger patient.

Abstracts

Background: Bile salts have been shown to contribute to bile duct injury after orthotopic liver transplantation (OLT). Cholangiocytes modify bile composition by reabsorption of bile salts (cholehepatic shunt) and contribute to bile flow by active secretion of sodium and water via cystic fibrosis transmembrane conductance regulator (CFTR). We hypothesized that changes in these transporters may be associated with the development of bile duct injury after OLT. Aim: To examine the expression of cholangiocyte transporters in liver grafts and to correlate this with the development of bile duct injury. Methods: In 37 adult liver transplant recipients, liver biopsies were taken during and after OLT. Changes in the apical sodium-dependent bile salt transporter (ASBT), the organic solute transporters (OSTalpha and beta), and CFTR were assessed using real-time RT-PCR and immunofluorescence staining. Gene expressions were correlated with bile salt concentration as well as with the histological degree of bile duct injury. Results: Compared to normal controls, OSTalpha expression was significantly down-regulated at the end of cold storage and 3 hours after graft reperfusion, but levels normalized at one week after OLT. OSTbeta expression was more than 8-fold up-regulated at the time of transplantation and levels increased further during the first week. There were no major changes in the expression of CFTR. Expression of these transporters did not correlate with changes in bile salt concentration or with the development of bile duct injury after OLT. Conclusion: Liver transplantation is associated with changes in the expression of the bile salt transporters OST alpha and OST beta, while CFTR expression remains stable. Changes in OST alpha/beta do not correlate with bile salt concentration or the degree of bile duct injury, suggesting that the cholehepatic shunt does not play a major role in the development of bile salt induced bile duct injury after OLT.

High incidence of cholesterol gallstone disease in type 1 Gaucher disease: characterizing the biliary phenotype of type 1 Gaucher disease

In Gaucher disease (GD), lysosomal glucocerebrosidase deficiency results in glucosylceramide accumulation in macrophage lysosomes. Hepatocytes do not accumulate glucosylceramide due in part to biliary secretion. Although gallstones (GS) occur in type 1 Gaucher disease (GD1), the chemical nature of stones, their association with metabolic parameters, and whether bile composition is altered are not understood. We assessed the prevalence of GS, their chemical composition, biliary lipids, and associated metabolic factors. The study cohort comprised 417 patients comprehensively evaluated for GD1 severity. Ascertainment of GS, fasting lipoprotein profile, and bile lipid analyses were performed. The prevalence of GS in GD1 was 32%. Compared with men, the prevalence of GS was higher in women, increasing from 4.2% and 11.8% at age 20-29 years to 71% and 60% at age >70 years, respectively. Patients with GS were more likely to be asplenic (p < 0.0001), older (p < 0.0001), have higher low-density lipoprotein (LDL) cholesterol (p = 0.002), and more severe GD1 disease compared with those without GS. On multiple logistic regression analysis, factors associated with GS were age (p < 0.001), female sex (p = 0.03), and splenectomy (p = 0.005). Compared with the general population, prevalence of GS was approximately 5-fold higher. Bile lipid analyses revealed cholesterol stones in five patients and pigment stones in one. Bile lipid composition was abnormal and contained glucosylceramide. Our results point to a metabolic syndrome in GD1 consisting of a propensity to cholesterol GS, low high-density lipoprotein (HDL) cholesterol, LDL cholesterol, and body mass index (BMI) associated with abnormal biliary lipid secretion.

Cholestatic Liver Disease in Children

Inherited syndromes of intrahepatic cholestasis and biliary atresia are the most common causes of chronic liver disease and the prime indication for liver transplantation in children. Our understanding of the pathogenesis of these diseases has increased substantially by the discovery of genetic mutations in children with intrahepatic cholestasis and the findings that inflammatory circuits are operative at the time of diagnosis of biliary atresia. Building on this solid foundation, recent studies provide new insight into genotype-phenotype relationships and how mutations produce altered bile composition and cholestasis. New evidence exists that although liver transplantation is curative for patients with end-stage liver disease owing to cholestasis, some patients may develop recurrence of cholestasis because of the emergence of autoantibodies that disrupt canalicular function in the new graft. Progress is also evident in biliary atresia, with recent studies identifying candidate modifier genes and directly implicating lymphocytes and inflammatory signals in the pathogenesis of bile duct injury and obstruction.

Utility of Choledochoscopy/EUS to Diagnose and Treat Choledocholithiasis in Pregnancy

Purpose: The risk of choledocholithiasis is higher during pregnancy due to alteration in bile composition and biliary stasis that occurs during gestation. Treatment of these patients during pregnancy is challenging due to the risks from sedation and fluoroscopy. This is an interesting case of application of new technique to diagnose and treat choledocholithiasis. A twenty-six year old pregnant female at 28 weeks gestation presented with right upper quadrant pain, progressive jaundice, and low-grade fever. Her past medical history was significant for sickle cell disease, preeclampsia and choledocholithiasis. She had undergone a previous ERCP for stone removal after her last pregnancy. Upon presentation to the hospital, her total bilirubin was 6.8, AST 139, ALT 113, ALP 128 and WBC count was 16,500. Transabdominal ultrasound revealed a 9-mm common bile duct with possible filling defect. In order to avoid radiation exposure to fetus by fluoroscopy, EUS and choledochoscopy were used to examined the bile duct and achieve complete clearance. The linear echoendoscope revealed multiple hyperechoic well-rounded stones throughout the common bile duct. A prior sphincterotomy had been performed, so deep cannulation of the bile duct using the SpyGlass choledochoscope (Boston Scientific) was easily achieved. The SpyScope was useful in determining the exact location of the stones. A guidewire was advanced through the SpyScope into the left intrahepatic system. A 10 mm extraction balloon was used to extract multiple yellow pigment stones from CBD. The SpyScope was then reinserted and complete clearance of the CBD was seen. A 10 Fr 5-cm plastic biliary stent was placed to ensure drainage and prevent recurrent cholangitis during the remainder of her pregnancy. The linear echoendoscope was used to confirm adequate placement of the biliary stent. Subsequently, the patient had complete resolution of her symptoms and improvement in her LFT. The incidence of gallstones during pregnancy is estimated to be between 3% and 12%. Symptomatic gallstone disease is the second most common abdominal emergency in pregnant women and may require surgical intervention. In addition to the baseline risks of ERCP, fluoroscopy poses additional risk to the fetus. Various publications have reported using ERCP during pregnancy without fluoroscopy by using bile aspiration for confirmation of CBD cannulation, sphincterotomy and balloon extraction of stones. All these approaches are not perfect with regard to ensuring complete CBD clearance. This case highlights how choledochoscopy and endoscopic ultrasound are safe alternatives to fluoroscopy for evaluation of biliary disorders during pregnancy.

Impact of experimental colitis on hepatobiliary transporter expression and bile duct injury in mice

The pathogenetic link between ulcerative colitis and sclerosing cholangitis (SC) is unclear. We hypothesized that colitis induces changes in bile composition via inflammation-induced reduction of hepatobiliary transporter gene expression, ultimately resulting in cholestasis and bile duct injury. Alterations in transporter expression and bile secretion in acute dextran sulphate sodium (DSS)-induced colitis were compared with lipopolysaccharide (LPS)-treated mice serving as positive control. Whether chronic DSS-colitis elicits cholangitis in genetically predisposed animals was studied in heterozygous multidrug resistance gene 2 knockout mice (Mdr2(+/-)). LPS but not DSS-colitis changed major hepatobiliary transporters (Ntcp, Bsep, Mrp2-4, Ostalpha/beta, Abcg5/8, Oatp1-4, Mdr1b and Mdr2), enzymes (Cyp3a11 and Cyp7a1), nuclear receptors (RXRalpha, FXR, CAR and PXR) and proinflammatory mediators (tumour necrosis factor alpha and inducible nitric oxide synthase). Formation of toxic bile reflected by an increased bile acid/phospholipid ratio was observed neither in acute nor in chronic colitis, although heterozygous Mdr2(+/-) mice developed mild portal inflammation after chronic colitis. In contrast to LPS, DSS-colitis has a minor impact on hepatobiliary gene expression and bile secretion. Therefore, intestinal inflammation-associated changes of hepatobiliary transporter expression do not play a pathogenetic role in SC.

The role of the hepatocyte cytokeratin network in bile formation and resistance to bile acid challenge and cholestasis in mice #

The intermediate filament cytoskeleton of hepatocytes is composed of keratin (K) 8 and K18 and has important mechanical and nonmechanical functions. However, the potential role of the K8/K18 network for proper membrane targeting of hepatocellular adenosine triphosphate-binding cassette transporters and bile formation is unknown. We therefore designed a comparative study in K8 and K18 knockout mice and respective wild-type controls to test the hypothesis that intermediate filaments of hepatocytes play a role in normal bile formation. In addition, we challenged mice either with a 1% cholic acid-supplemented diet or a diet containing the porphyrinogenic xenobiotic 3,5-diethoxycarbonyl-1,4-dihydrocollidine to determine the effect of K8/K18 loss on bile flow/composition and liver injury under different physiological and toxic stress stimuli. Protein expression levels and membrane localization of various transporters and anion exchangers were compared using western blotting and immunofluorescence microscopy, respectively, and bile flow and composition were determined under various experimental conditions. Our results demonstrate that loss of the intermediate filament network had no significant effect on bile formation and composition, as well as expression levels and membrane targeting of key hepatobiliary transporters under baseline and stress conditions. However, loss of K8 significantly increased liver injury in response to toxic stress. The intermediate filament network of hepatocytes is not specifically required for proper bile formation in mice.

Relation of Gallbladder Motility to Viscosity and Composition of Gallbladder Bile in Patients with Cholesterol Gallstones

Background/Aims: Increased viscosity and supersaturation of cholesterol in gallbladder bile, as well as an impaired motility of the gallbladder, are considered to be important factors in the pathogenesis of cholesterol gallstones. However, the relation of these parameters has not yet been determined. Material and Methods: Bile viscosity (mPa·s) was measured by rotation viscosimetry and the composition of gallbladder bile was determined using standard methodology. Gallbladder motility was calculated as ejection fraction in percent of total volume 45 min after a test meal using ultrasonography in patients with gallstones prior to elective cholecystectomy. Results: The study included 35 patients with cholesterol gallstones. Viscosity of gallbladder bile ranged between 0.9 and 12.5 mPa·s (median 2.2 mPa·s) and an ejection fraction of the gallbladder of 55.4 ± 18.3% (mean ± SD) was determined. No significant correlation (r = 0.19, p < 0.2) between the 2 parameters could be calculated. Analysis of the composition of gallbladder bile revealed a positive correlation of all components to biliary viscosity but not to the motility of the gallbladder, with the exceptions of a negative correlation (r = 0.39, p < 0.02) between mucin concentration and the ejection fraction at 45 min after the test meal. Conclusions: The motility of the gallbladder appears to be unrelated to the viscosity of gallbladder bile or gallbladder bile composition. The negative correlation between the ejection fraction of the gallbladder and mucin concentration of gallbladder bile suggests that chronic inflammation of the gallbladder wall is associated with both an impaired motility of the gallbladder and increased mucin release into gallbladder bile.

Drug exposure and perceived adverse drug events reported by liver-transplant patients

Posttransplant patient outcome and quality of life are affected by different factors, such as post-graft context, psychological state, and polymedication. Many surveys have been carried out to study immunosuppressant ADRs, and have mainly used a questionnaire completed by patients, but few have asked patients about their drug exposure. The aim of this study is to describe drug exposure and adverse drug events (ADEs) reported by liver-transplant patients (LTP). This observational, retrospective study assessed questionnaires from LTPs concerning demographic data, drug exposure, and ADEs. 118 LTPs exposed to 5.9 (+/- 2.8) drugs with immunosuppressive regimens, consisting mainly of tacrolimus (79.3%), cyclosporine (18.1%), or sirolimus (2.6%), were also exposed to antihypertensive drugs (43.2%), protonpump inhibitors (30.5%), statins (28.8%), drugs acting on bile composition (26.3%), and diuretics (19.5%). 1,389 ADEs were reported: 30.1% neurological, 13.4% cutaneous, 12.4% hematological, 11.1% digestive, 10.1% osteomuscular, 6.6% cardiovascular, and 16.3% others. Significantly more ADEs were reported by patients exposed to cyclosporine than those receiving tacrolimus (p < 0.05). Patients with a transplant for < 18 months had more tremors and those with a transplant for > 79 months reported more hirsutism, gingival hypertrophia, and arterial hypertension. This study shows the value of patient-reporting via structured interviews for both drug exposure and ADEs, and the importance of this approach to complement total data collection.

Endogenous Bile Salts Are Associated With Bile Duct Injury in the Rat Liver Transplantation Model

Nonanastomotic biliary strictures are a serious complication after orthotopic liver transplantation (OLT) and are difficult to cure. We examined the role of endogenous bile salt toxicity in the pathogenesis of bile duct injury after OLT. A total of 90 Sprague-Dawley rats were divided into three groups: Sham-operated group (Sham, n=30), OLT group with 1 hr donor liver preservation (OLT-1h, n=30), and OLT group with 12 hr donor liver preservation (OLT-12h, n=30). Bile was collected and analyzed biochemically. The histopathologic study was performed to determine the intrahepatic bile duct morphologic changes. Hepatic expressions of bile transporters Ntcp, Bsep, Mdr2, and Oatps were detected. During the first 2 weeks after transplantation, bile salt secretion was not parallel with phospholipid secretion, resulting in high biliary bile salt-to-phospholipid (BS:PL) ratio. The expression of bile transporters was consistent with the change of bile composition. Bile duct injury correlated significantly with bile salt secretion and BS:PL ratio. Moreover, OLT group with longer donor liver preservation time (OLT-12h) had significantly lower Mdr2 messenger RNA/protein level, higher BS:PL ratio, and better correlation between BS:PL ratio and bile duct injury compared with those of OLT-1h. The unparallel secretion of bile salts and phospholipids results in cytotoxic bile formation with high BS:PL ratio after liver transplantation. Longer donor liver preservation time will increase graft bile cytotoxicity. The results of this study suggest that endogenous bile salts play a role in the pathogenesis of bile duct injury after OLT.

Side chain structure determines unique physiologic and therapeutic properties of norursodeoxycholic acid in Mdr2 −/− mice†‡§

24-norursodeoxycholic acid (norUDCA), a side chain-modified ursodeoxycholic acid derivative, has dramatic therapeutic effects in experimental cholestasis and may be a promising agent for the treatment of cholestatic liver diseases. We aimed to better understand the physiologic and therapeutic properties of norUDCA and to test if they are related to its side chain length and/or relative resistance to amidation. For this purpose, Mdr2(-/-) mice, a model for sclerosing cholangitis, received either a standard diet or a norUDCA-, tauro norursodeoxycholic acid (tauro- norUDCA)-, or di norursodeoxycholic acid (di norUDCA)-enriched diet. Bile composition, serum biochemistry, liver histology, fibrosis, and expression of key detoxification and transport systems were investigated. Direct choleretic effects were addressed in isolated bile duct units. The role of Cftr for norUDCA-induced choleresis was explored in Cftr(-/-) mice. norUDCA had pharmacologic features that were not shared by its derivatives, including the increase in hepatic and serum bile acid levels and a strong stimulation of biliary HCO(3)(-)-output. norUDCA directly stimulated fluid secretion in isolated bile duct units in a HCO(3)(-)-dependent fashion to a higher extent than the other bile acids. Notably, the norUDCA significantly stimulated HCO(3)(-)-output also in Cftr(-/-) mice. In Mdr2(-/-) mice, cholangitis and fibrosis strongly improved with norUDCA, remained unchanged with tauro- norUDCA, and worsened with di norUDCA. Expression of Mrp4, Cyp2b10, and Sult2a1 was increased by norUDCA and di norUDCA, but was unaffected by tauro- norUDCA. The relative resistance of norUDCA to amidation may explain its unique physiologic and pharmacologic properties. These include the ability to undergo cholehepatic shunting and to directly stimulate cholangiocyte secretion, both resulting in a HCO(3)(-)-rich hypercholeresis that protects the liver from cholestatic injury.

Very high aquaporin-1 facilitated water permeability in mouse gallbladder

Water transport across gallbladder epithelium is driven by osmotic gradients generated from active salt absorption and secretion. Aquaporin (AQP) water channels have been proposed to facilitate transepithelial water transport in gallbladder and to modulate bile composition. We found strong AQP1 immunofluorescence at the apical membrane of mouse gallbladder epithelium. Transepithelial osmotic water permeability (Pf) was measured in freshly isolated gallbladder sacs from the kinetics of luminal calcein self-quenching in response to an osmotic gradient. Pf was very high (0.12 cm/s) in gallbladders from wild-type mice, cAMP independent, and independent of osmotic gradient size and direction. Although gallbladders from AQP1 knockout mice had similar size and morphology to those from wild-type mice, their Pf was reduced by approximately 10-fold. Apical plasma membrane water permeability was greatly reduced in AQP1-deficient gallbladders, as measured by cytoplasmic calcein quenching in perfluorocarbon-filled, inverted gallbladder sacs. However, neither bile osmolality nor bile salt concentration differed in gallbladders from wild-type vs. AQP1 knockout mice. Our data indicate constitutively high water permeability in mouse gallbladder epithelium involving transcellular water transport through AQP1. The similar bile salt concentration in gallbladders from AQP1 knockout mice argues against a physiologically important role for AQP1 in mouse gallbladder.

Proteomic analysis and 2-D-Gel electrophoresis of bile-duct bile: A new diagnostic approach

Aims: Extensive research has been performed on the lipid composition of bile and its mucin content, but not on differences in protein composition. Though this could identify new molecular players and markers of disease. Methods: Bile samples were acquired by ERC from patients suffering from gallstones, primary sclerosing cholangitis (PSC), liver transplantation (LTx), and/or malignancy of the pancreatico-biliary tract. Purified protein samples were analyzed by 1- and 2-D-Gel electrophoresis, DIGE and mass spectrometry. Results: We collected >800 bile samples from different cholestatic diseases. A new purification protocol was established removing albumin, IgG and lipids minimizing interference with further analysis and protein loss. 1-D- and 2-D-PAGE verified samples of highest quality and reproducibility. Protein concentrations from patients suffering from PSC or malignancy were markedly elevated. Protein patterns of bile revealed by 1-D and 2-D gel electrophoresis were quite similar within the gallstone group, but different to the majority of patients in the PSC and LTx group. Mass spectrometry of >200 protein spots yielded serum proteins as the major components in all groups, but a number of proteins could be detected with a higher frequency only within subgroups of cholestatic diseases. Protein composition of bile-duct bile is markedly different compared to reported gallbladder bile. Conclusion: This newly established protocol of bile-duct bile acquisition, storage, purification and analysis enables for a new diagnostic approach in cholestatic diseases. By applying state-of-the-art techniques we could already identify that: 1) Bile-duct bile has a protein composition different than gallbladder bile, 2) Protein composition of bile of patients suffering from different diseases is stable regarding major serum proteins, but variable regarding other secreted proteins. 3) Protein patterns in 2-D-gels are different among various cholestatic diseases.

Safety of endoscopic retrograde cholangiopancreatography during pregnancy

The risk of choledocholithiasis is expected to be higher during pregnancy. This is attributed to alteration in bile composition as well as biliary stasis that take place during gestation. There is significant concern regarding application of endoscopic procedures especially the more invasive ones for treatment of choledocholithiasis during pregnancy. Our aim was to provide an additional support to the efficacy and safety of endoscopic retrograde cholangiopancreatography (ERCP) in the management of biliary diseases during pregnancy. The medical records of 10 pregnant patients who underwent ERCP at King Abdullah University Hospital, during the period from 2002 to 2007 were reviewed. Pregnancy course and outcomes were followed up in all cases. Results were analysed and compared with published data on safety and efficacy of this procedure. The mean age for mothers was 24.3 years. The mean duration of gestation was 18.4 weeks. Two patients were in the first trimester, five were in their second trimester and another three in the third trimester. The main indication for ERCP was obstructive choledocholithiasis on ultrasound and liver function tests. Fetal radiation exposure was not routinely measured. During, or after, the procedure there was no need for tocolytic agents. Also there was no intrauterine fetal distress. Screening for congenital anomalies was negative in all cases. Major complications of biliary obstruction have been prevented through this procedure. Short-term follow up for all neonates whom mothers underwent ERCP during pregnancy supports its safety. However, specific long-term fetal complications of radiation exposure have not been investigated yet.

Bile composition in patients with noncirrhotic portal hypertension

Bile composition in patients with noncirrhotic portal hypertension

Bile composition in Alagille Syndrome and PFIC patients having Partial External Biliary Diversion.

BackgroundPartial External Biliary Diversion (PEBD) is a surgical intervention to treat children with Progressive Familial Intrahepatic Cholestasis (PFIC) and Alagille syndrome (AGS). PEBD can reduce disease progression, and examining the alterations in biliary lipid composition may be a prognostic factor for outcome.MethodsBiliary lipid composition and the clinical course of AGS and PFIC patients were examined before and after PEBD.ResultsPre-PEBD bile from AGS patients had greater chenodeoxycholic/cholic acid (CDCA/CA), bile salt, cholesterol and phospholipid concentrations than PFIC patients. AGS patients, and PFIC patients with familial intrahepatic cholestasis 1 (FIC1) genotype, responded better to PEBD than PFIC patients with bile salt export protein (BSEP) genotype. After successful PEBD, AGS patients have higher biliary lipid concentrations than PFIC patients and PEBD also increases biliary phospholipid concentrations in FIC1 patients.ConclusionBoth AGS and FIC1 patients can benefit from PEBD, and preserved biliary phospholipid concentrations may be associated with better outcomes post-PEBD.

Altered bile composition after liver transplantation is associated with the development of nonanastomotic biliary strictures

Nonanastomotic biliary strictures are troublesome complications after liver transplantation. The pathogenesis of NAS is not completely clear, but experimental studies suggest that bile salt toxicity is involved. In one hundred and eleven adult liver transplants, bile samples were collected daily posttransplantation for determination of bile composition. Expression of bile transporters was studied perioperatively. Nonanastomotic biliary strictures were detected in 14 patients (13%) within one year after transplantation. Patient and donor characteristics and postoperative serum liver enzymes were similar between patients who developed nonanastomotic biliary strictures and those who did not. Secretions of bile salts, phospholipids and cholesterol were significantly lower in patients who developed strictures. In parallel, biliary phospholipids/bile salt ratio was lower in patients developing strictures, suggestive for increased bile cytotoxicity. There were no differences in bile salt pool composition or in hepatobiliary transporter expression. Although patients who develop nonanastomotic biliary strictures are initially clinically indiscernible from patients who do not develop nonanastomotic biliary strictures, the biliary bile salts and phospholipids secretion, as well as biliary phospholipids/bile salt ratio in the first week after transplantation, was significantly lower in the former group. This supports the concept that bile cytotoxicity is involved in the pathogenesis of nonanastomotic biliary strictures.

Lessons from the toxic bile concept for the pathogenesis and treatment of cholestatic liver diseases

Alterations in bile secretion at the hepatocellular and cholangiocellular levels may cause cholestasis. Formation of 'toxic bile' may be the consequence of abnormal bile composition and can result in hepatocellular and/or bile duct injury. The canalicular phospholipid flippase (Mdr2/MDR3) normally mediates biliary excretion of phospholipids, which normally form mixed micelles with bile acids and cholesterol to protect the bile duct epithelium from the detergent properties of bile acids. Mdr2 knockout mice are not capable of excreting phospholipids into bile and spontaneously develop bile duct injury with macroscopic and microscopic features closely resembling human sclerosing cholangitis. MDR3 mutations have been linked to a broad spectrum of hepatobiliary disorders in humans ranging from progressive familial intrahepatic cholestasis in neonates to intrahepatic cholestasis of pregnancy, drug-induced cholestasis, intrahepatic cholelithiasis, sclerosing cholangitis and biliary cirrhosis in adults. Other examples for bile injury due to the formation of toxic bile include the cholangiopathy seen in cystic fibrosis, after lithocholate feeding (in mice) and vanishing bile duct syndromes induced by drugs and xenobiotics. Therapeutic strategies for cholangiopathies may target bile composition/toxicity and the affected bile duct epithelium itself, and ideally should also have anti-cholestatic, anti-fibrotic and anti-neoplastic properties. Ursodeoxycholic acid (UDCA) shows some of these properties, but is of limited efficacy in the treatment of human cholangiopathies. By contrast to UDCA, its side chain-shortened homologue norUDCA undergoes cholehepatic shunting leading to a bicarbonate-rich hypercholeresis. Moreover, norUDCA has anti-inflammatory, anti-fibrotic and anti-proliferative effects, and stimulates bile acid detoxification. Upcoming clinical trials will have to demonstrate whether norUDCA or other side chain-modified bile acids are also clinically effective in humans. Finally, drugs for the treatment of cholangiopathies may target bile toxicity via nuclear receptors (FXR, PPARalpha) regulating biliary phospholipid and bile acid excretion.

Glycine Protects Bile Physiology and Biliary-Specific Liver Cell Metabolism from Ischemia-Reperfusion Injury: A1H NMR Study

Liver ischemia-reperfusion (I/R) injury is a major complication of liver resection and transplantation. Cytokine release by activated Kupffer cells (KCs) play a central role in the inflammatory cascade of I/R injury. There is evidence that glycine may protect against liver I/R injury by inhibition of KC activity. However, its effect on bile flow, an established marker of hepatic function, and bile composition is not known. A rabbit model of hepatic lobar warm I/R was used. Under general anesthesia, the sham group (n = 6) underwent laparotomy alone for 7 h. The I/R group (n = 6) underwent 60 min of left and median lobe inflow occlusion and 6 h of reperfusion. The glycine + I/R group (n = 6) underwent a same procedure to controls after receiving glycine 5 mg/kg intravenous infusion for over 15 min. Bile flow was collected, measured, and analyzed by proton magnetic resonance spectroscopy. Glycine prevented the significant reduction in bile flow seen in I/R at 6 h reperfusion (145.0 ± 11.4 vs. 108.3 ± 28.2 μL/min/g, p = 0.011). It reduced phosphatidylcholine shedding (1.2 ± 0.8 vs. 3.0 ± 0.5 μmol/L, p = 0.001) and lactate surge (8.1 ± 4.3 vs. 26.3 ± 7.8 μmol/L, p < 0.001) compared to I/R. Glycine significantly stimulated bile acid (17.9 ± 2.8 vs. 8.9 ± 2.1 μmol/L, p = 0.001), pyruvate (1.3 ± 0.3 vs. 0.7 ± 0.1 μmol/L, p = 0.005), glucose (3.9 ± 0.9 vs. 1.6 ± 0.6 μmol/L, p = 0.007), and acetoacetate (0.7 ± 0.1 vs. 0.4 ± 0.1 μmol/L, p = 0.009) release in bile on reperfusion compared to I/R. Glycine induced a reduction of liver I/R results in increased bile flow and altered bile composition.

Ursodeoxycholic acid reduces lipid peroxidation and mucin secretagogue activity in gallbladder bile of patients with cholesterol gallstones

Recently it has been postulated that gallbladder mucin hypersecretion observed in the pathogenesis of cholesterol gallstone disease may be induced by biliary lipid peroxidation. Ursodeoxycholic acid treatment reduces mucin concentration and the formation of cholesterol crystals in the gallbladder bile of patients with cholesterol gallstones and this effect might be mediated by a decrease of biliary lipid peroxidation. In a double-blind, placebo-controlled trial patients with symptomatic cholesterol gallstones received either ursodeoxycholic acid (750 mg daily) (n = 10) or placebo (n = 12) 10-12 days prior to cholecystectomy. As a marker for lipid peroxidation malondialdehyde was measured in bile together with mucin concentration. In addition, the mucin secretagogue activity of the individual bile samples was assessed in cultured dog gallbladder epithelial cells. Ursodeoxycholic acid therapy resulted in a significant reduction of lipid peroxidation in bile as determined by the biliary malondialdehyde concentration (1.36 +/- 0.28 vs. 2.05 +/- 0.38 micromol L(-1); P < 0.005) and the malondialdehyde (micromol L(-1))/total bile acid (mmol L(-1)) ratio (0.02 +/- 0.005 vs. 0.06 +/- 0.01; P < 0.001). Furthermore, a decrease in mucin concentrations (0.7 +/- 0.3 vs. 1.3 +/- 0.5 mg mL(-1); P < 0.005) and of the mucin secretagogue activity of gallbladder bile (0.9 +/- 0.2 vs. 2.2 +/- 0.3 times control; P < 0.001) was observed. The reduction of lipid peroxidation and mucin secretagogue activity of gallbladder bile induced by ursodeoxycholic acid treatment may contribute to the beneficial effects of this drug on gallbladder bile composition and symptoms in cholesterol gallstone patients.