1. Volume 123, Issue 4, Pages 1238-1251 (October 2002)
Ursodeoxycholic acid aggravates bile infarcts in bile duct–ligated and Mdr2 knockout mice via disruption of cholangioles 
Peter Fickert, Gernot Zollner, Andrea Fuchsbichler, Conny Stumptner, Andreas H. Weiglein, Frank Lammert, Hanns–Ulrich Marschall, Oleksiy Tsybrovskyy, Kurt Zatloukal, Helmut Denk, Michael Trauner 
Gastroenterology 
Volume 123, Issue 4, Pages (October 2002)
DOI: /gast
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2. Fig. 1
UDCA aggravates liver injury in CBDL mice. (A, D) Feeding of hydrophilic UDCA (0.5% wt/wt for 10 days) in sham-operated controls did not result in significant changes of liver histology. (B, E) CBDL for 3 days results in elongated bile ducts with dilated lumina and irregular epithelium surrounded by edematous portal connective tissue infiltrated by neutrophils. In addition, bile infarcts (asterisks) predominantly in acinar zones 1 and 2 infiltrated by neutrophils are observed. Inserts (A–C) show portal tract changes in greater detail. (C, F) Mice were fed a UDCA supplemented diet (for 7 days) and then subjected to CBDL (for 3 days) and continued UDCA feeding. CBDL in UDCA-fed mice results in a marked increase in size and numbers of bile infarcts (asterisks). cv, central vein; pv, portal vein; arrowheads indicate portal tracts. (H&E staining; original magnifications 20× for A–C, 80× for D–F.)
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3. Fig. 2
UDCA feeding increases mortality in CBDL mice. CBDL in UDCA-fed mice resulted in significantly increased mortality in comparison with controls (P < 0.05). Surviving UDCA-prefed animals were sacrificed on day 3. In addition, UDCA feeding started after CBDL resulted in a 100% mortality rate after 5 days (6 of 6 vs. 0 of 6 in controls, P < 0.05).
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4. Fig. 3
UDCA-induced aggravation of liver injury in selective bile duct-ligated (SBDL) mice is confined to ligated lobes. SBDL of the left lobe was performed in mice fed control diet (upper panel) and mice fed UDCA (lower panel). Livers are viewed from top (facies visceralis). Gross inspection reveals liver necroses restricted to the ligated left lobes, whereas the nonligated right lobes (indicated by the closed lines) remain unchanged. Note the larger necrotic areas (arrowheads) in UDCA-fed SBDL mice in comparison with controls. Note also the significant enlargement of the gallbladder (asterisks) in UDCA-fed SBDL mice indicating increased bile flow and biliary pressure.
Gastroenterology  , DOI: ( /gast )
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5. Fig. 4
Bile infarcts communicate with the biliary tract. UDCA-fed mice were injected with India ink into either the common bile duct (A–C) or the portal vein (D) 6 hours after CBDL. (A, B) Injection of India ink into the bile duct results in dye deposition in the periportal area reflecting canals of Hering and also fills bile canaliculi (arrows). (C) India ink injected into the common bile duct enters the bile infarct and becomes distributed within the necrotic area (asterisk), indicating a connection between the biliary system and the bile infarct. (D) In contrast, injection of India ink into the portal vein does not reveal dye accumulation within the bile infarct (asterisk). cv, central vein; pv, portal vein. (H&E staining; original magnification 20× for A, 160× for B, 80× for C, D.)
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6. Fig. 5
Rupture of the cholangioles (canals of Hering) and hepatocyte necroses in UDCA-fed common bile duct-ligated (CBDL) mice. Electron microscopy was performed in UDCA-fed CBDL mice (A, B). Note dilated canaliculi (asterisks) with loss of microvilli. (A, C) In addition, cholangioles (stars) formed by 2 cholangiocytes (C) and 1 hepatocyte (H) are dilated. (B, D) Moreover, ruptured cholangioles (stars) are observed with surrounding necrotic hepatocytes, characterized by karyorrhexis, lysis of cell membranes, and invading neutrophil granulocytes (NG) and red blood cells (RBC). (D) Arrowheads indicate still intact parts of the wall of the cholangioles, whereas arrows indicate its rupture. Bar = 500 nm.
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7. Fig. 6
Significantly increased biliary pressure in UDCA-fed mice after bile duct occlusion. Cholangiomanometry was performed in mice fed UDCA or control diet to determine whether the observed rupture of cholangioles (canals of Hering) is related to increased biliary pressure. (A) Schematic representation of experimental system for cholangiomanometry. After laparotomy and cholecystectomy (1), the common bile duct was ligated immediately before entering the duodenum (2). A polyethylene tube was then inserted into the common bile duct through a small incision and advanced over the junction of the superior pancreatic duct, secured with a silk tie, and connected to a pressure transducer (3). After equilibration, the common bile duct was occluded and intrabiliary pressure continuously measured for 10 minutes. (B) Typical manometric tracing in a UDCA- and control diet-fed mouse after common bile duct occlusion. Note the more rapid and pronounced increase of biliary pressure in the UDCA-fed mouse compared with control.
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8. Fig. 7
Absence of apoptosis in UDCA-fed common bile duct-ligated (CBDL) mice. Active caspase-3 immunohistochemistry (A, B) and double-immunofluorescence labeling for activated caspase-3 and cytokeratin (CK) 18 (C–F) were performed. (A) In Fas-activated mice (IP injection of the Fas agonist Jo2), activated caspase-3 (red) is detected in several hepatocytes. (C, E) Double-immunofluorescence labeling for CK 18 (green) and activated caspase-3 (red and yellow due to colocalization with CK 18) in Jo2-challenged mice reveals hepatocellular activation of caspase-3 in cells with CK intermediate filament breakdown and granular cytoplasmic condensation (characteristic of apoptotic epithelial cell death). (B) In contrast, immunoreaction for activated caspase-3 is negative in UDCA-fed CBDL mice. (D, F) In addition, CK intermediate filament alterations are rare (arrowhead). In particular, zones of bile infarcts (asterisks) and bile duct epithelial cells show no signs of apoptotic cell death.
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9. Fig. 8
Development of bile duct lesions in Mdr2 P-glycoprotein deficient (Mdr2−/−) mice. (A) Liver of 1-day (1d)-old Mdr2−/− mouse showing a dense neutrophil-granulocytic infiltrate (arrows) as well as proliferating fibroblasts and a ductular reaction in larger portal tracts. (B) One-week (1w)-old Mdr2−/− mouse liver showing qualitatively similar but somewhat more pronounced alterations. (C) Four-week (4w)-old Mdr2−/− mouse liver with periductal fibroblast proliferation and periductal fibrosis as well as granulocytic infiltration. Note also partial necrosis of bile duct and desquamation of the bile duct epithelium (arrowheads). (D) Eight-week (8w)-old Mdr2−/− mouse liver with periductal fibrosis of a medium-sized bile duct of onionskin type leading to fibrous scar formation with obliteration of bile duct lumen. (H&E staining; original magnification 160× for A–C and 80× for D.)
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10. Fig. 9
Mdr2 P-glycoprotein-deficient (Mdr2−/−) mice develop PSC-like cholangiopathy. Plastination of the bile duct system followed by maceration of the liver was performed in (A) wild-type and (B) Mdr2−/− mice. (A) The bile duct system was filled with plastogen G after cannulation of the gallbladder (left margin). In contrast to the delicately structured bile duct system in the wild-type mouse, the Mdr2−/− mouse (B) shows strictures and dilatations of extra- and intrahepatic bile ducts, leading to a beaded appearance.
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11. Fig. 10
UDCA decreases periductal fibrosis but induces bile infarcts in Mdr2 P-glycoprotein-deficient (Mdr2−/−) mice. Two-month-old Mdr2−/− mice were fed a control diet (A, C, E) or UDCA (0.5%, wt/wt)-supplemented diet (B, D, F) for 7 days. (A) Liver histology in Mdr2−/− mice reveals narrowed lumina of the larger and middle-sized bile ducts with pronounced periductal fibrosis (arrowheads). (C, E) In addition, a pronounced ductular reaction in tracts surrounded by neutrophils is observed. (B) UDCA feeding in Mdr2−/− mice results in decreased periductular fibrosis (arrowheads) but (D) only light reduction of the ductular reaction. (F) In addition, areas of liver cell necrosis (asterisk; identical to bile infarcts observed after CBDL) are noted in UDCA-fed Mdr2−/− mice. pv, portal vein. (H&E staining; original magnifications 80× for A, B, E, F and 120× for C, D.)
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12. Fig. 11
UDCA feeding aggravates bile infarcts in CBDL mice via disruption of cholangioles. Suggested sequence of events. (1) UDCA feeding leads to increased bile flow. (2) CBDL in the presence of increased bile flow results in increased intracanalicular and intraductal pressure. (3) Cholangiole rupture ensues because of increased biliary pressure. (4) Leakage of bile leads to necrosis of surrounding hepatocytes. pv, portal vein; bd, bile duct; ha, hepatic artery.
Gastroenterology  , DOI: ( /gast )
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