Development of extrahepatic bile duct excluding gall bladder in human fetuses: histological, histochemical, and immunohistochemical analysis.

BACKGROUND: The fetal development of extrahepatic bile ducts (EBD) is unkown. MATERIALS AND METHODS: Development of EBD was examined by immunohistochemistry in 16 fetuses of 7-40 gestational week (GW). Gall bladder (GB) was not investigated. RESULTS: At seven GW, a hepato-pancreatic bud (HPB) was seen near the hepatic hilus. At eight GW, embryonic EBD, GB and pacreas developed from HPB. Portal veins (PV) and hepatic arteries (HAs) were present in EBD at eight GW. Liver parenchyma was already present in seven GW. At eight GW, EBD at porta hepatis (PH) was already established; PH EBD was derived from ductal plate (DP). The distal and middle EBD gradually develeped and took shape of EBD at nine GW. In PH, cystic and hepatic ducts developed from DP at eight GW. EBD developed further, accompanying many nerve fibers (NF) at PH and distal and middle EBD. Apparent PV and HA were seen around 12 GW. Around 20 GW, HA and capillaries proliferated, giving rise to peribiliary capillary plexus (PCP) in all parts of EBD. EBD grew gradually further, and around 30 GW extrahepatic peribiliary glands (EPG) emerged from EBD but not from cystic duct. Around 36 GW, exocrine pancreatic acinar cells emerged from remodeled DP at PH. At term (40 GW), EBD was established but was as yet immature. Numerous NF were present around EBD. Histochemically, EBD epithelium had no mucins at 7-12 GW but contained neutral and acidic mucins at 23-40 GW. EPG had abundant neutral and acidic mucins. Immunohistochemically, alpha-fetoprotein (AFP) was consistently positive in the epithelial and mesenychyma. The NF and muscles of HPB present at seven GW were positive for neural cell adhesion molecule (NCAM), neuron-specific enolase (NSE), platelet-derived growth factor receptor-α (PDGFRA), and KIT, but they disappeared in nine GW. Expressions of cytokeratin (CK) seven and CK19 in EBD and EPG were slight or none, while expression of CK8 was moderate, and that of CK18 was strong. NF were positive for NCAM, NSE, synaptophysin, and chromogranin, and PDGFRA. MUC1 and MUC6 apomucins were noted in EBD and EPG. EPG contained numerous endocrine cells positive for chromogranin, synaptophysin, NCAM and NSE. A few endocrine cells positive for these antigens were seen in EBD. Numeous KIT-positive stem cells (SC) were seen in PH, EBD, PV, HA, PCP, and EPG. NCAM-positive and bcl-2-positive SC were also located in these structures. Epithelial cells of EBD and EPG showed expressions of MET, PDGFRA, CA19-9, MUC1, MUC2, MUC6, KIT, bcl-2, and ErbB2. No expressions of HepPar1, carcinoembryonic antigen (CEA), and epithelial membrane antigen (EMA) were noted. CONCLUSIONS: Although the findings have limitatios because this study of humans are descriptive one, the present data suggest that the processes of the development and differentiation of EBD system may be associated with EBD SC, CK prolifes, SFC/KIT signaling, HGF/MET signaling, PDGRa/PDGFRA signaling, fibroblast growth factor/ErbB2 signaling, neuroendocrine lineage, NF differentiation, pancreatic aninar cell differentiation, PCP differentiation, MUC apomucins differentiation, and expressions of AFP and CA19-9. HepPar1, EMA and CEA were not involved in them.

Reovirus type-2-triggered autoimmune cholangitis in extrahepatic bile ducts of weanling DBA/1J mice.

BACKGROUND: Reovirus is a proposed cause of infantile biliary atresia. However, mechanistic insight regarding Reo-2 as a potential cholangiotropic virus is lacking. Furthermore, it is unknown whether Reo-2 infection can induce autoimmune-mediated bile duct injury. METHODS: Lesions of bile ducts in newborn DBA/1J mice infected with Reo-2 were analyzed immunopathologically. RESULTS: Damage to biliary epithelia occurs after Reo-2 infection. In addition, nonsuppurative cholangitis with fibrosis in extrahepatic (especially septal) bile ducts developed following complete viral clearance from the liver. At the inflamed ducts, major histocompatibility complex class I expressing((+)) and FAS(+) cholangiocytes were associated with FAS ligand(+) lymphocytes and tumor necrosis factor-α(+) mononuclear cells (macrophages and lymphocytes). These cholangiocytes were apoptotic and necrotic. Moreover, affected ducts were infiltrated by CD3(+), CD4(+), CD8(+), IFN-γ(+), and FAS(+) lymphocytes. Analysis of blood from Reo-2-infected mice revealed that they developed anticholangiocyte cytoplasm antibodies and had high serum IFN-γ concentration. Notably, there was no increase in Foxp3(+) lymphocytes at inflamed ducts, lymph nodes, and thymi. CONCLUSION: Reo-2 infection induced T-helper cell type 1-dependent injury to bile ducts in weanling mice. The lesions observed in mice may be analogous to those associated with human infantile biliary atresia, which are caused by an autoimmune-mediated process.

Anatomic variations of extrahepatic bile ducts and evaluation of the length of ducts composing the cystohepatic triangle / Variaciones anatómicas de los ductos biliares extrahepáticos y longitud de los ductos del trígono hepato-cístico

It is of paramount importance for surgeons to have a thorough knowledge of the normal anatomy of the extrahepatic bile ducts and its variations due to the high frequency with which they perform in this anatomic site. The cystohepatic triangle, or Calot's Triangle, is bound by the cystic duct, common hepatic duct, and the hepatic border; therefore, its surface area depends on the conformation of these ducts and is closely linked to surgical procedures performed in this region. It has been reported that the length and the position of these ducts may be related to the formation of bile duct stones, Mirizzi's syndrome, and bile duct cancer. Thus, the present work aims to analyze the configuration of the extrahepatic biliary tree and its possible variations, as well as measure the components that make up the cystohepatic triangle. For this task 41 samples from fixated human cadavers were analyzed, with 25 consisting of anatomic parts (liver and biliary tree) and 16 in situ samples. The extrahepatic biliary trees were dissected in order to measure the length of the common hepatic and cystic ducts with a digital caliper, and all anatomic variations were registered. The length of the common hepatic duct varied between 4.18 mm and 50.64 mm, with an average of 21.76 +/- 9.51 mm. The length of the cystic duct varied between 7.28 and 38.88 mm, with an average of 19.11 +/- 6.77 mm. Anatomic variations were found in 3 samples (7.3 percent): in one of them the cystic duct connected to the left hepatic duct; in another, the cystic duct connected to the right hepatic duct; in the third, there was a triple confluence of hepatic ducts (two right ducts and one left duct). The results are a contribution to the clinical and surgical anatomy of this region.

El conocimiento de la anatomía normal de las vías biliares extrahepáticas y sus variaciones es fundamental para los cirujanos digestivos debido a la frecuencia con que se actúa en esa región. El trígono hepato-cístico o de Calot es delimitado por el ducto cístico, ducto hepático común y el margen del hígado. De este modo, su área depende de la conformación de esos ductos y está íntimamente relacionada a procedimientos quirúrgicos efectuados en esa zona. Se ha señalado que la longitud y la disposición de esos ductos estarían involucradas en la formación de cálculos biliares, síndrome de Mirizzi y neoplasias de vías biliares. Así, el presente estudio tuvo como objetivo analizar la configuración de las vías biliares extrahepáticas y sus posibles variaciones, además de registrar parámetros métricos de los componentes del sistema biliar que integran el trígono hepato-cístico. Para el estudio se utilizaron 41 muestras de cadáveres formolizados de individuos adultos, siendo 25 piezas anatómicas (de hígado y vías biliares) y 16 in situ. Las vías biliares fueron disecadas, esquematizadas y fotografiadas, se registró la longitud del ducto hepático común y cístico con un caliper digital. La longitud promedio del ducto hepático común fue de 21,76 +/- 9,51 mm, variando de 4,18 mm a 50,64 mm; la longitud promedio del ducto cístico fue de 19,11 +/- 6,77 mm, variando de 7,28 a 38,88 mm. Se observaron variaciones en 3 muestras (7,3 por ciento), en una de ellas, el ducto cístico se unió al ducto hepático izquierdo, en otra, el ducto cístico se unió al ducto hepático derecho y en la otra muestra, se presentó una confluencia triple de ductos hepáticos, dos derechos y uno izquierdo. Los resultados obtenidos son un aporte a la anatomía clínica y quirúrgica de la región.

Interstitial cells of Cajal are present in human extrahepatic bile ducts.

BACKGROUND AND AIMS: Interstitial cells of Cajal (ICC) are distributed with smooth muscle throughout the gastrointestinal tract and are involved in regulating motility. ICC were recently discovered in the wall of the human gallbladder. This study sought to determine whether ICC are present in human bile ducts. METHODS: Biliary tract samples were obtained from several sources: surgical specimens (n = 16, 11 women, mean age 61 years); archival post-mortem specimen (n = 1, 86 years, man); and cadavers (n = 2, 68 and 80 years, men). Paraffin-embedded sections (3 microm) from the gallbladder (fundus, body and neck) and both extrahepatic and intrahepatic bile ducts were investigated. A double immunofluorescence protocol using polyclonal and monoclonal c-kit antibodies and mast cell tryptase was used to distinguish c-kit-positive cells with typical ICC morphology from c-kit-positive mast cells. Small bowel samples were used as positive controls. ICC in the gallbladder were confirmed by ultrastructural study. RESULTS: c-kit-positive cells with characteristic ICC morphology were identified in the subepithelial and muscular layers of the gallbladder and extrahepatic bile ducts. They were most prominent within the muscle layer of the extrahepatic bile ducts where they were organized into loosely arranged laminae running parallel to circular smooth muscle fibers. ICC were not found in intrahepatic bile ducts. CONCLUSION: This study demonstrates for the first time that ICC are present in human extrahepatic bile ducts where they are more densely aggregated than in the gallbladder. This cellular network is likely to be involved in biliary tract motility and its related disorders.

Beta cells occur naturally in extrahepatic bile ducts of mice.

Insulin-secreting beta cells were thought to reside only in the pancreas. Here, we show that beta cells are also present in the extra-hepatic bile ducts of mice. They are characterised by insulin and C-peptide content, the presence of secretory granules that are immunoreactive for insulin, and the ducts exhibit glucose-stimulated insulin secretion. Genetic lineage labelling shows that these beta cells arise from the liver domain rather than the pancreas and, by histological study, they appear to be formed directly from the bile duct epithelium in late embryogenesis. Other endocrine cell types (producing somatostatin and pancreatic polypeptide) are also found in close association with the bile-duct-derived beta cells, but exocrine pancreatic tissue is not present. This discovery of beta cells outside the mammalian pancreas has implications for regenerative medicine, indicating that biliary epithelium might offer a new source of beta cells for the treatment of diabetes. The finding also has evolutionary significance, because it is known that certain basal vertebrates usually form all of their beta cells from the bile ducts. The mammalian bile-duct-derived beta cells might therefore represent an extant trace of the evolutionary origin of the vertebrate beta cell.

Immunohistochemical and ultrastructural study of ito cells (fat-storing cells) in response to extrahepatic bile duct ligation in broiler chickens.

The Ito cell (fat-storing cell) lies in perisinusoidal space of liver and has a variety of functions. We investigated the immunohistochemistry and ultrastructure of Ito cells in normal and cholestatic livers of broiler chickens. Immunohistochemistry demonstrated that Ito cells expressed HHF35 muscle actin, vimentin, desmin, glial fibrillary acidic protein (GFAP), neuron-specific enolase (NSE), chromogranin A and cytokeratins in normal livers. These cells were diffusely scattered throughout the lobules. Livers treated with extrahepatic bile duct ligation (BDL) showed cholestasis, fibrosis, proliferation of biliary ductules and Ito cells. The Ito cells were frequently found in fibrotic areas and were larger in size with more extensive immunoreactivity than those of normal livers. Ultrastructural study demonstrated that Ito cells were closely associated with the production of collagen fibers in BDL livers. These findings suggest that Ito cells actively react against hepatocytic injuries and play a major role in the hepatic fibrogenesis of cholestatic livers of chickens.

Biliary tract of the rat as observed by scanning electron microscopy of cast samples.

The three-dimensional distribution of the biliary tract in the rat was studied by scanning electron microscopy of biliary casts. The casts were prepared by a retrograde infusion of a low viscosity or monomeric methacrylate resin mixture into the common bile duct. No resin flow from the bile canaliculi to sinusoidal capillaries was ever noted. Bile canaliculi formed intricate meshworks and drained via the Hering's canals into the bile ductules. The bile canalicular meshworks of adjacent lobules intercommunicated with each other. The bile ductules formed a marked periportal plexus around the portal vein branch, and drained into the intrahepatic bile duct running along the portal vein branch. The junctional zone of the Hering's canal and bile ductule usually showed an ampullary dilation. When the Hering's canal directly drained into a thick bile ductule or into a periportal plexus of bile ductules, such an ampullary dilation at the origin of the bile ductule was never replicated. The extrahepatic bile duct protruded many crypt-like projections which presumably corresponded to parietal glands. It is suggested that the periportal plexus of bile ductules may store the bile as a substitute for the gallbladder.

Centrosome abnormalities in human carcinomas of the gallbladder and intrahepatic and extrahepatic bile ducts.

During mitosis, 2 centrosomes ensure accurate assembly of bipolar spindles and fidelity of the chromosomal segregation. The presence of more than 2 copies of centrosomes during mitosis can result in the formation of multipolar spindles, unbalanced chromosome segregation, and aneuploidy. Recent studies have provided evidence that centrosome hyperamplification plays a pivotal role in carcinogenesis. Using immunofluorescence analysis with gamma-tubulin and pericentrin antibodies, paraffin-embedded sections from 40 malignant biliary diseases including gallbladder cancers (GC; n = 13), intrahepatic cholangiocellular carcinoma (CCC; n = 19), and extrahepatic bile duct cancers (BDC; n = 8) were examined. Thirty-seven benign biliary diseases including chronic cholecystitis, gallbladder adenoma, hepatolithiasis, and choledochal cyst were included as benign controls. The frequencies of the centrosome abnormalities were 70% for GC, 58% for CCC, and 50% for BDC, respectively. The frequencies of centrosome abnormalities in malignant biliary diseases were significantly higher than in their benign counterparts (GC, CCC, BDC; P =.001,.002, and.001, respectively). The results of current study also indicated that biliary malignancy in the advanced stage (III-IV) displayed a higher frequency of centrosome abnormalities than in the early stage (I-II) (P <.001). We conclude that abnormalities in size, number, and shape of the centrosome are frequently observed in biliary tract malignancy. Centrosome abnormalities started to occur in the early stage of biliary malignancy and became very frequent in the advanced stage. This implies that centrosome abnormality might relate to the transition from early to advanced malignancy in biliary malignancy.

Establishment of a new extrahepatic bile duct carcinoma cell line, TFK-1.

A new human extrahepatic bile duct carcinoma cell line (TFK-1) was established from a surgically resected tumor specimen, which was histologically diagnosed as partly papillary adenocarcinoma and partly differentiated tubular adenocarcinoma. The tumor cells cultured in RPMI-1640 medium supplemented with 10% FBS grew as monolayers showing epithelial-like morphology with a population doubling time of 37 hr during exponential growth at passage 40. Chromosome number was distributed in the range of 72 to 76, with a modal number of 73. Tumor markers (CEA, CA19-9, ST-439, DUPAN-2) were negative in culture supernatant and plasma of SCID mice grafted with TFK-1 cells. Though no point mutation at 12 codon of K-ras was detected, expression of c-erb B-2 product and MUC1 antigen was positive. TFK-1 is the third cell line established from extrahepatic bile duct carcinomas in the world literature, and should provide useful information on various aspects of this type of neoplasm.

The progressive degeneration of interlobular bile ducts in biliary atresia: an ultrastructural study.

The ultrastructure of the interlobular bile ducts were observed in hepatic specimens obtained at surgery in ten patients with biliary atresia, in order to investigate the progress of their degeneration over time after the onset of the disease. As an index for the passage of time, the degree of fibrosis in the portal tracts was investigated. Then the ultrastructural features in the interlobular bile ducts were contrasted with the grading of portal fibrosis Generally, the ultrastructural changes of the interlobular bile ducts were more marked in the cases with severe portal fibrosis than that observed in the cases with mild fibrosis. This correlation suggests that the degeneration of the interlobular bile ducts progressively worsens over time after the obliteration of the extrahepatic bile duct.

[Electron microscopic changes in the liver of patients with calculosis of the extrahepatic bile ducts]. / Elektronnomikroskopski promeni v cherniia drob pri bolni s kalkuloza na ekstrakhepatalnite zhluchni putishta.

Thirty-four patients, operated for calculosis of extrahepatic bile ducts with obstructive jaundice, are studied. They are divided in two groups according to cholestasis duration-10 and 20 days respectively. Material from liver tissue, taken intraoperatively, is investigated light-and electron-microscopically. The statistical analysis shows a significant increase in cell count in either group, as compared to controls (2.0-controls, 4.2-group one, and 6.2-group two; P < 0.05). There is electron microscope evidence of perisinusoidal fibrosis periportally, ito cell's fibroblastic transformation, and collagenogenesis phase II and III. In group one fibrogenesis is moderate, and in group two-markedly expressed. It is established that parallel to increase in cholestasis duration, fibrogenesis in zone 1 and 2 of the liver lobule intensifies. This explains the necessity of undertaking early surgical intervention.

Effects of biliary endoprostheses on the extrahepatic bile ducts in relation to subsequent operation of the biliary tract.

Despite the widespread use of transpapillary biliary endoprostheses, little is known about their effect on the extrahepatic bile ducts. In an experimental study in dogs, we induced inflammatory changes in the bile ducts by stent insertion and studied the reversibility of these changes after stent removal. In addition, the consequences of a period of preoperative stenting for subsequent operation of the biliary tract and the eventual detrimental effects of stenting on the histologic factors of the liver were studied. Twenty-six mongrel dogs were randomly divided into four groups: group 1, stenting during four weeks; group 2, after four weeks stenting, construction of a hepaticojejunostomy; group 3, four days of common bile duct (CBD) ligation, four weeks stenting and hepaticojejunostomy, and group 4, four days of CBD ligation and hepaticojejunostomy. All dogs were sacrificed two months after the last procedure. Hepatic biopsies were obtained during each procedure and bile duct biopsies during hepaticojejunostomy and upon sacrifice. Four weeks of stenting of a normal or obstructed CBD resulted in fibrosed bile ducts, showing severe chronic inflammation with papillary hyperplasia of the epithelium. All bile cultures grew fecal bacteria. Two months after stent removal, inflammation was still present, albeit less severe. Stenting and subsequent surgical treatment resulted in a higher incidence of postoperative complications (54 percent) compared with the control group (14 percent), although this did not reach statistical significance. Hepatic histologic factors were not markedly changed after transpapillary endoprosthesis placement, but after hepaticojejunostomy cholangiolitis was observed. Whenever transpapillary biliary endoprostheses are used, the local effects on the extrahepatic bile ducts and the subsequent bacterial contamination of the bile should be considered.

Morphological observation on extrahepatic bile duct of golden hamsters fed a lithogenic diet: histochemical, ultrastructural and cell kinetic studies.

Cholelithiasis is often accompanied with disorders of the extrahepatic bile duct and pancreas. However, studies on changes of the extrahepatic bile duct in cholecystolithiasis have not shown this clearly. We therefore investigated sequential histologic changes, mucous secretion and DNA synthetic activity of the extrahepatic bile duct epithelium in cholecystolithiasis. Serial changes in the mucosal epithelial cells of the extrahepatic bile duct in golden hamsters treated with a lithogenic diet were examined by light and electron microscopy and an ultrastructural quantitative technique. In addition, epithelial cell kinetics were studied using bromodeoxyuridine (BrdU). After the 2nd week of diet, the extrahepatic bile duct showed an increase in goblet cells of the mucosal epithelium, a large number of secretory granules in the upper nuclear area of the epithelial cells and an increase in the BrdU-labeling index compared with the controls. These findings indicate that mucous secretion and cell turnover were enhanced in the mucosal epithelial cells of the extrahepatic bile duct in cholelithiasis, suggesting that the epithelial cells of the bile duct were protected and regenerating.

Development of the adult endocrine pancreas during metamorphosis in the sea lamprey, Petromyzon marinus L. II. Electron microscopy and immunocytochemistry.

The development of the adult endocrine pancreas was followed throughout metamorphosis in the sea lamprey using electron microscopy and immunocytochemistry. It was discovered that the caudal pancreas develops from the larval extrahepatic common bile duct through the process of transdifferentiation (dedifferentiation/redifferentiation). Early in metamorphosis the bile duct epithelial cells possess large vacuoles, resembling autophagic vacuoles, containing recognizable cell material. There is a loss of the large bundles of intermediate filaments characteristic of the larval bile duct epithelium. These same cells are then seen to contain granules immunoreactive for insulin. Pancreatic islets develop within the base of the bile duct epithelium from these transdifferentiated cells and migrate into the surrounding connective tissue to form the caudal pancreas. The cranial pancreas was found to develop from the epithelia lining the developing adult diverticulum and anterior intestine in a similar fashion as those in the larva. The second cell type to appear in either portion of the developing pancreas is similar to the third cell type of the adult: cells immunoreactive for somatostatin do not appear until late in metamorphosis in either region.

Sarcoma botryoides of the extrahepatic bile ducts. A light- and electron microscopic study of a case.

The clinical, light- and electron microscopic features of a sarcoma botryoides of the common bile duct of a 2 1/2-year-old boy are reported. Despite the absence of cross-striation under the light microscope, many tumour cells were found to contain abundant thin and thick filaments arranged in a sarcomere-like pattern at the electron-microscopic examination of originally paraffin-embedded material. Ultrastructural examination of embryonal rhabdomyosarcoma may be of value for a definite diagnosis also when only formalin-fixed and paraffin-embedded material is available.