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We determined whether extrahepatic biliary epithelial cells can differentiate into cells with phenotypic features of hepatocytes. Gallbladders were removed from transgenic mice expressing hepatocyte-specific [beta]-galactosidase ([beta]-Gal) and cultured under standard conditions and under experimental conditions designed to induce differentiation into a hepatocyte-like phenotype. Gallbladder epithelial cells (GBEC) cultured under standard conditions exhibited no [beta]-Gal activity. [beta]-Gal expression was prominent in 50% of cells cultured under experimental conditions. Similar morphological changes were observed in GBEC from green fluorescent protein transgenic mice cultured under experimental conditions. These cells showed higher levels of mRNA for genes expressed in hepatocytes, but not in GBEC, including aldolase B, albumin, hepatocyte nuclear factor-4[alpha], aldehyde dehydrogenase 1, and glutamine synthetase, and they synthesized bile acids. Additional functional evidence of a hepatocyte-like phenotype included LDL uptake and enhanced benzodiazepine metabolism. Connexin-32 expression was evident in murine hepatocytes and in cells cultured under experimental conditions, but not in cells cultured under standard conditions. Notch 1, 2, and 3 and Notch ligand Jagged 1 mRNAs were downregulated in these cells compared with cells cultured under standard conditions. CD34, [alpha]-fetoprotein, and Sca-1 mRNA were not expressed in cells cultured under standard conditions, suggesting that the hepatocyte-like cells did not arise from hematopoietic stem cells or oval cells. These results point to future avenues for investigation into the potential use of GBEC in the treatment of liver disease. stern cells; cell culture; transgenic mice; transdifferentiation; biliary epithelium

: ObjectivesOxysterols are ubiquitous in the body and are potential cytotoxic agents in addition to being metabolic regulators. Although bile contains high concentrations of cholesterol, oxysterol concentrations in bile and the effect of infection on oxysterol levels have not been measured, nor has their origin been studied. The purpose of this study was to determine if infection of the biliary tract was associated with increased concentrations of oxysterols in the bile and, if so, which oxysterols showed a significant change.: MethodsHepatic bile was obtained from eight patients with biliary tract disease by means of a naso-biliary catheter. Oxysterols were extracted and purified by solid-phase extraction, derivatized and measured by gas chromatography–mass spectrometry.: ResultsThe following were quantified in hepatic bile: 7-α-hydroxycholesterol, 7-β-hydroxycholesterol, cholestan-3-beta,5-alpha,6-β-triol, 25-hydroxycholesterol, 26-hydroxycholesterol, 7-ketocholesterol, and 7-α-hydroxy-4-cholesten-3-one. Total oxysterols in hepatic bile ranged from 0.133 μmol/L to 7.748 μmol/L (1.47 ± 2.55 μmol/L). Levels of 7-α-hydroxycholesterol and 7-β-hydroxycholesterol were increased in infected bile (14.2 ± 15.1 × 10−3% of cholesterol vs 1.9 ± 0.5 × 10−3% of cholesterol, p < 0.05, and 22.0 ± 25.0 × 10−3% of cholesterol vs 1.6 ± 1.2 × 10−3% of cholesterol, p < 0.05, respectively). Serum C-reactive protein levels correlated positively with biliary levels of 7-α-hydroxycholesterol (R = 0.948), 7-β-hydroxycholesterol (R = 0.976), cholestan-3-beta,5-alpha,6-β-triol (R = 0.823), 7-α-hydroxy-4-cholesten-3-one (R = 0.846,) and 7-ketocholesterol (R = 0.973). Different oxysterols were found in gallstones, chiefly 3-keto-cholest-4-ene (624 ± 316 parts per million [ppm] of dry weight), 3-keto-cholesta-4,6-diene (240 ± 329 ppm) and 7-keto-cholesterol (77 ± 81 ppm). Incubation of human leukocytes with model bile in the presence of bacterial lipopolysaccharide resulted in changes in sterol composition, including increases in oxysterols.: ConclusionsWe have identified and quantified oxysterols from uninfected and infected human hepatic bile and from gallstones and gallbladder bile. Biliary infection may be involved in the biogenesis of oxysterols in bile through the production of reactive oxygen species from activated leukocytes. [Copyright &y& Elsevier]

OBJECTIVE: Numerous investigators have proposed a role for bacteria in biliary lithogenesis. We hypothesized that bacterial DNA is present in gallstones, and that categorical differences exist between gallstone type and the frequency of bacterial sequences. METHODS: Polymerase chain reaction (PCR) was used to amplify bacterial 16S rRNA and uidA (encoding Escherichia coli [E. coli] β-glucuronidase) genes in different types of gallstones. PCR products were sequenced. RESULTS: Bacterial 16S rRNA and uidA DNA sequences in E. coli were detected in all brown pigment, common bile duct, and mixed cholesterol gallstones (n = 14). In contrast, only one (14%) of seven pure cholesterol gallstones yielded a PCR product. Most (88%) mixed cholesterol gallstones yielded PCR amplification products from their central, as well as their outer, portions. Sequenced products possessed 88-98% identity to 16S rRNA genes of E. coli and Pseudomonas species. CONCLUSIONS: Bacterial DNA sequences are usually present in mixed cholesterol (to 95% cholesterol content), brown pigment, and common bile duct, but rarely in pure cholesterol gallstones. The presence of bacterial β-glucuronidase is also suggested. The role of bacteria and their products in the formation of mixed cholesterol gallstones, which comprise the majority of cholesterol gallstones, warrants further study. [ABSTRACT FROM AUTHOR]

Bile acids are implicated as aetiological factors in many types of gastrointestinal tract cancer including cholangiocarcinoma (CCA). Alterations in bile acid concentrations may affect the pathogenesis of these different types of cancer. Our aim was to determine the bile acid profile in gallbladder bile from patients who underwent liver resection. Thirty-seven patients with cholangiocarcinoma, 5 with hepatocellular carcinoma, and 7 with benign biliary diseases were studied. High pressure liquid chromatography was used to analyze conjugated and unconjugated bile acids. CCA patients with low (≤ 2 mg/dl) and high (>2 mg/dl) levels of total serum bilirubin had significantly higher total bile acid and conjugated bile acid concentrations than the benign biliary disease group. Markedly elevated levels of cholic and chenodeoxycholic acid were found in CCA cases with high levels of total serum bilirubin. Concentrations of total bile acids and primary bile acid were correlated with serum cholesterol, bilirubin and ALP in CCA. Notably, correlation of the carcinoembryonic antigen, a tumor marker, was found with level of total bile acids and chenodeoxycholic acid. These findings suggest a different pattern of bile acid concentration in cancer patients compared to patients with benign biliary diseases. Thus, accumulation of certain bile acids may be involved in carcinogenesis.