Histopathology of cholestasis.
ABSTRACT Cholestasis may be extrahepatic or intrahepatic in origin. The block in bile secretion may be complete or incomplete to variable extent. Complete cholestasis occurs in case of primary parenchymal disease (intrahepatic cholestasis) or total obstruction of extrahepatic bile ducts (extrahepatic cholestasis). Incomplete block in bile secretion is due to incomplete obstruction of intra- or extrahepatic bile ducts (intra- or extrahepatic cholestasis or both). Histologically, it is useful to distinguish between bilirubionstasis and cholate-stasis. Complete secretory block causes as early changers: bilirubinostasis (in hepatocytes, canaliculi and Kupffer cells) in acinar zone 3, and "ductular reaction" in acinar zone 1. The latter refers to an increase in periportal ductular profiles, associated with neutrophil infiltration. With longer duration of cholestasis, further lesions ensue: feathery degeneration of hepatocytes due to retention of detergent bile acids, cholestatic liver cell rosettes representing a shift from hepatocellular to biliary differentiation, xanthomatous cells reflecting hyperlipidemia, cholate stasis in acinar zone 1 due to overload of membrane-damaging bile acids, eventually paraportal bile infarcts, and progressive ductular reaction. The latter may be due to multiplication of pre-existing ductules, to metaplasia of periportal hepatocytes, or to activation of progenitor cells. It is invariably associated with periductular fibrosis: the pacemaker for increasing matrix deposition, resulting in biliary fibrosis and eventually in true biliary cirrhosis. Incomplete cholestasis (e.g. PBC, PSC) is characterized by absence of bilirubinostasis during long periods of time, whereas the afore mentioned features of chronic cholestasis do appear. Hence follows that the most reliable markers of chronic incomplete cholestasis are cholate stasis, cholestatic rosettes and ductular reaction. Bilirubinostasis is only a late and often ominous sign.
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ABSTRACT: Sandwich-cultured hepatocytes (SCH) are used commonly to investigate hepatic transport protein-mediated uptake and biliary excretion of substrates. However, little is known about the disposition of endogenous bile acids (BAs) in SCH. In this study, four endogenous conjugated BAs common to rats and humans [taurocholic acid (TCA), glycocholic acid (GCA), taurochenodeoxycholic acid (TCDCA), and glycochenodeoxycholic acid (GCDCA)], as well as two BA species specific to rodents (α- and β-tauromuricholic acid; α/β TMCA), were profiled in primary rat and human SCH. Using B-CLEAR® technology, BAs were measured in cells+bile canaliculi, cells, and medium of SCH by LC-MS/MS. Results indicated that, just as in vivo, taurine-conjugated BA species were predominant in rat SCH, while glycine-conjugated BAs were predominant in human SCH. Total intracellular BAs remained relatively constant over days in culture in rat SCH. Total BAs in control (CTL) cells+bile, cells, and medium were approximately 3.4, 2.9, and 8.3-fold greater in human than in rat. The estimated intracellular concentrations of the measured total BAs were 64.3±5.9 μM in CTL rat and 183±56 μM in CTL human SCH, while medium concentrations of the total BAs measured were 1.16±0.21 μM in CTL rat SCH and 9.61±6.36 μM in CTL human SCH. Treatment of cells for 24h with 10 μM troglitazone (TRO), an inhibitor of the bile salt export pump (BSEP) and the Na⁺-taurocholate cotransporting polypeptide (NTCP), had no significant effect on endogenous BAs measured at the end of the 24-h culture period, potentially due to compensatory mechanisms that maintain BA homeostasis. These data demonstrate that BAs in SCH are similar to in vivo, and that SCH may be a useful in vitro model to study alterations in BA disposition if species differences are taken into account.Toxicology and Applied Pharmacology 02/2012; 261(1):1-9. · 3.98 Impact Factor
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ABSTRACT: Abstract Background Extensive bile duct proliferation is a key feature of the tissue reaction to clinical and experimental forms of liver injury. Experimental infection of mice by Schistosoma mansoni is a well-studied model of liver fibrosis with bile duct hyperplasia. However, the regulatory mechanisms of bile duct changes are not well understood. In this study we report the reproducible isolation of long-term cultures of cholangiocytes from mice livers with schistosomal fibrosis. Methods We have isolated a cholangiocyte cell line from Schistosoma -induced liver granulomas using a combination of methods including selective adhesion and isopyknic centrifugation in Percoll. Results The cell line was characterized by morphological criteria in optical and transmission electron microscopy, ability to form well differentiated ductular structures in collagen gels and by a positive staining for cytokeratin 18 and cytokeratin 19. To our knowledge, this is the first murine cholangiocyte cell line isolated from schistosomal fibrosis reported in the literature. Conclusion After 9 months and 16 passages this diploid cell line maintained differentiated characteristics and a high proliferative capacity. We believe the method described here may be a valuable tool to study bile duct changes during hepatic injury.BMC Gastroenterology 01/2006; · 2.11 Impact Factor
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ABSTRACT: Culture of extrahepatic bile duct epithelial cells is a useful model to investigate physiology of extrahepatic bile duct epithelia and hepatobiliary disease mechanisms. The aim of this work was to establish and characterize a primary murine extrahepatic bile duct epithelial cell culture. Epithelial cells were isolated from extrahepatic bile ducts of BALB/c mice that were intraperitoneally injected with newborn bovine serum to induce the proliferation of extrahepatic bile ducts' epithelial cells and cultured on rat tail type I collagen-coated plastic culture flask containing DMEM/HamF12 with 10% FBS and 10 ng/ml epidermal growth factor at 37°C in an incubator with 5% humidified CO(2). The cells showed typical morphologic characteristics of epithelial phenotypes with cobblestone appearance in monolayer within 5-6 d after culture; they were positive against anticytokeratin-19 immunostaining. Transmission electron microscopy showed typical bile duct epithelia with microvilli on the cytomembrane, Golgi complex, massive mitochondria, and rough endoplasmic reticulum in the cytoplasmic. The growth curve of the epithelial cells was determined by a MTT assay which showed a normal sigmoidal growth curve. This culture technique might be a reliable method for isolation, purification, and primary culture of extrahepatic bile duct epithelial cells that can serve as a model for in vitro studies on the pathophysiology of hepatobiliary diseases as well as pharmacological and toxicological targets relevant to hepatobiliary diseases.In Vitro Cellular & Developmental Biology - Animal 12/2010; 46(10):820-3. · 1.29 Impact Factor