Evaluating and interpreting bile duct changes in liver allograft biopsies

Despite great advances in treatment of liver disease, liver transplant remains the only cure for end stage liver disease and hepatocellular carcinoma. Successful management of immediate and intermediate post-transplant events determines the overall graft outcome, and pathologists play a key role in achieving long-term graft survival. In evaluating liver allograft biopsies, bile duct changes frequently pose diagnostic dilemma as they could occur in association with or the result of various causes of graft dysfunction, but in which accurate classification and interpretation is key to subsequent management. The approach to interpreting these changes share some similarity to the non-transplant liver biopsy interpretation but several additional factors come into play in transplant setting sometimes making interpretation more complex. This review describes bile duct changes occurring in association with various clinical entities in liver allografts, such as acute and chronic rejection, obstruction, small-for-size syndrome, recurrent biliary diseases, and recurrent fibrosing cholestatic hepatitis C, and others, and discusses an approach to interpreting and reporting them.     

大鼠原位肝移植术后胆道并发症的影响因素

目的 研究大鼠原位肝移植术后胆道并发症的各种影响因素,为建立更稳定的大鼠原位肝移植模型提供实验依据.方法 雄性SD大鼠87只,完全随机法分组,其中移植组48只、非移植组36只、假手术组3只,实验共分胆管-胆管重建移植组、胆管-十二指肠重建移植组、胆管-胆管重建+肝动脉结扎非移植组、胆管-胆管重建非移植组、胆管-十二指肠重建+肝动脉结扎非移植组、胆管-十二指肠重建非移植组和假手术组7组.各手术处理组以出现明显胆道并发症或移植后14 d作为观察时间终点.通过肝脏大体标本肉眼观察、病理学检测和血清学分析等判断胆道并发症的发生率及其严重程度并进行各组间比较.结果 移植组总胆道并发症发生率为69.6%,明显高于非移植组(25.0%)和假手术组(0%),差异有统计学意义(均P<0.05).在移植组和非移植组中,胆管-十二指肠重建组的胆道并发症发生率明显高于胆管-胆管重建组(90.9%比50.0%,38.9%比11.1%,均P<0.05).非移植组中,结扎肝动脉组的胆道并发症高于不结扎肝动脉组(38.9%比11.1%,P<0.05).结论 经典"二袖套法"大鼠原位肝移植模型尚不完善,仍需进一步改进.就大鼠原位肝移植模型而言,胆管-十二指肠重建方式并非理想的选择.     

Regeneration of extrahepatic bile ducts by tissue engineering with a bioabsorbable polymer

With the widespread adoption of laparoscopic cholecystectomy and living-donor liver transplantation in recent years, complications involving the biliary system, stenosis in particular, are increasing. Various invasive and non-invasive techniques are now available for the treatment of biliary stenosis, but all are compromised by a high risk of recurrence and other problems. As a potential solution, our group has developed a bioabsorbable polymer (BAP) tube for implantation as a bypass graft. In the study reported here, we implanted this BAP tube and confirmed bile duct regeneration at the graft site after the tube had been degraded and absorbed into the body. We briefly describe our findings on extrahepatic biliary tissue regeneration, focusing on the possibility of its clinical application. This artificial bile duct may promote the development of novel treatments for biliary disease.     

MRCP与ERCP鉴别肝外胆管良恶性狭窄的对比研究

目的：探讨肝外胆管良、恶性狭窄患者的磁共振胰胆管成像（magnetic resonance cholangiopan-creatography，MRCP）与内镜逆行胆胰管造戥术（endoscopic retrograde cholangiopancreatography，ERCP）的征象，并比较两种检查方法诊断的准确性。方法：对46例经手术或病理检查明确病因的肝外胆管狭窄患者的术前MRCP和ERCP资料进行回顾性分析，根据图像肝外胆管狭窄的形态描述为边缘不规则或平滑、狭窄不对称或对称、狭窄中断或呈鼠尾状及双管征等，计算MRCP与ERCP诊断的灵敏度、特异度及准确性，用标尺准确测量肝外胆管狭窄的长度，并用t检验来比较。结果：肝外胆管癌表现为不规则边缘和不对称狭窄（92％）较良性狭窄（分别为22％，35％）普遍。MRCP诊断肝外胆管癌与良性狭窄的灵敏度、特异度及准确性分别是84％、71％及78％，ERCP分别是72％、71％及72％。肝外胆管癌引起狭窄长度平均是（30．0±8．5）mm，良性狭窄平均长度是（13．6±9.1）mm（P〈0.001）。结论：MRCP在鉴别诊断肝外胆管良恶性的病变引起胆管狭窄与ERCP相比是较准确的。     

Regeneration of extrahepatic bile duct--possibility to clinical application by recognition of the regenerative process.

The biliary epithelium is continuously exposed to highly cytotoxic bile acids and pathogens and thus is at persistent risk for injury. The monolayer mucosal epithelium protects the body from these dangers and once injured. The bile duct repair process essentially involves reconstruction of the bile duct with migrating cells, but there are many questions about the process. It is reported that implantation of a bioabsorbable polymer tube as a bypass graft into the extrahepatic bile duct resulted in bile duct regeneration in the graft site after the artificial duct had been degraded and absorbed. We briefly describe our findings on extrahepatic biliary tissue regeneration with the possibility for clinical applications in mind. The creation of this artificial bile duct may be able to promote the development of the treatment for biliary diseases.     

Primary sclerosing cholangitis--diagnosis and therapy

Primary sclerosing cholangitis (PSC) is a chronic cholestatic liver disease of unknown origin, characterized by inflammation, fibrosis, and obliteration of bile ducts, which ultimately results in biliary cirrhosis. The condition most commonly affects intrahepatic and extrahepatic bile ducts together, but sometimes only intrahepatic or extrahepatic ducts are involved. PSC is often associated with inflammatory bowel disease, especially ulcerative colitis. The majority of patients are initially asymptomatic, and identified on the basis of elevated serum levels of alkaline phosphatase or gamma-glutamyl transpeptidase, especially while screening patients with ulcerative colitis. Diagnosis is based on characteristic cholangiographic appearance with focal bile duct dilatations proximal to areas of stricturing that produce a beaded appearance. Ursodeoxycholic acid is most effective medical therapy, with other symptomatic measures, while liver transplantation is the treatment of choice for patients with advanced liver disease.     

彩色多普勒超声对肝移植后胆道并发症的诊断价值

背景：肝移植后胆道并发症是引起移植后移植肝失功的重要原因,彩色多普勒超声是目前公认的肝移植后患者随访首选简便的检查方法,可以为临床上对肝移植后并发症的诊治提供较可靠的证据。 目的：探讨彩色多普勒超声在肝移植后胆道并发症诊断中的应用价值。 方法：回顾性分析126例肝移植后常规超声检查时胆道并发症的发生情况及彩色多普勒超声检查结果。以磁共振胆胰管成像或超声造影或穿刺引流或临床诊断性治疗等诊断胆道并发症为标准,使用彩色多普勒超声观察肝移植后胆总管及肝内胆管是否有胆漏、狭窄、扩张、胆泥及结石形成。 结果与结论：126例中超声诊断胆道并发症的共33例,其中胆漏4例(3.2%),胆道狭窄11例(8.7%),胆道结石或(和)胆泥18例(14.3%)。提示彩色多普勒超声对肝移植后胆道并发症的诊断具有重要价值,可为临床的诊治提供依据。     

Two cases of common bile duct stone after liver transplantation

Biliary complications after orthotopic liver transplants are a continuing cause of morbidity and mortality. Biliary stones and sludge are less well known complications of hepatic transplantation, although they have long been recognized. Recently we experienced two cases of biliary stones developed after liver transplantation. One 32-year-old male, who frequently admitted due to recurrent cholangitis, was treated with percutaneous transhepatic biliary drainage and choledochojejunostomy with cholecystectomy. The other 58-year-old male, who had stones in commone bile duct, was treated by endoscopic manipulation. They are in good condition without recurrent bile duct stones or its accompanying complications. Although stones and sludge are relatively infrequent after liver transplantation, surgical or interventional radiologic treatments are usually performed for treatment.     

带血管蒂空肠修复胆管狭窄与缺损

目的 探讨用带血管蒂空肠组织修复胆管狭窄与缺损的手术方法。方法 采用带血管蒂空肠片修复胆管狭窄与缺损15例。系用带血管蒂空肠段行缺损胆管桥式吻合8例。结果 共23例患胆管狭窄与缺损病人,经手术本组除两例术后并发胆瘘行外引流在两周内治愈外,其余21例均获满意效果。结论 本术式具有防止胆汁返流和逆行感染,保持肝外胆管生理机能及保持胆汁的正常pH环境等优点。     

Congenital extrahepatic biliary atresia

Ten cases of congenital extrahepatic biliary atresia were studied ultrastructurally. Samples of liver were obtained from each and in six of the cases, fibrous tracts, which we hoped would contain extrahepatic bile ducts, were also secured. The observed extrahepatic biliary structures were real, but hypoplastic, bile ducts. In places, necrosed epithelial cells, without obvious inflammatory processes, could be observed. The ductular cell cytoplasmic changes and the inflammatory reaction are different according to whether extrahepatic or intrahepatic sites are considered. These differences, as well as the cytoplasmic modifications of liver parenchymal cells, seem to be the result of impaired bile flow. On the contrary, extrahepatic bile duct hypoplasia and necrosis seem to be directly related to the unknown origin of this disease. Whether the nuclear changes of hepatocytes are the expression of direct injury of the liver is another important question. If there is direct injury, it is possible that the disease might evolve as an independent liver disease despite a correctly performed and uncomplicated surgical intervention.     

原位肝移植后胆管铸型组肝功能分析及预警指标筛选

背景：当肝移植胆管铸型诊断明确时移植肝因为胆道梗阻和反复感染已经受到不同程度的损伤,因此早期发现胆管铸型显得尤为重要。 目的：检测原位肝移植后发生胆管铸型胆道并发症患者的肝功能,从中筛选出可供早期诊断胆管铸型的预警指标。 方法：回顾性分析经胆道内镜诊治或胆道造影已经确定发生肝移植后胆道并发症患者25例的肝功能指标,其中胆管铸型组7例,胆管吻合口狭窄5例,胆管铸型并吻合口狭窄13例;以胆道内镜检查肝移植后胆道正常的18例患者为对照组。 结果与结论：肝移植后1周,正常组与胆道并发症各分组受者血清肝功能指标差异均无显著性意义(P > 0.05)。肝移植后1个月胆管铸型组、胆管铸型并吻合口狭窄组血清γ-谷氨酰转肽酶水平显著高于胆道正常组(P  < 0.01),胆管吻合口狭窄组血清γ-谷氨酰转肽酶水平显著低于胆管铸型组(P  < 0.05)。提示胆管铸型是原位肝移植后潜在的并发症,γ-谷氨酰转肽酶可作为原位肝移植后胆管铸型组的预警指标,也是肝移植后鉴别黄疸原因的重要参考指标和胆道镜检查和外科治疗的指征。     

Limited impact of pre-existing donor specific HLA-antibodies (DSA) on long term allograft survival after first adult liver transplantation

The relevance of pre-existing HLA-antibodies (HLA-Ab) in liver transplantation (LTx) is controversial. While livers are allocated without HLA match or preoperative crossmatch testing, several studies point to an impact of donor specific antibodies (DSA) for acute rejection, bile duct complications or even graft loss.To investigate the role of DSA in long term liver allograft survival we analysed 177 pre transplant sera of first LTx patients with Luminex single antigen technology defining a MFI of >1500 as positive. The analyses were done retrospectively, and the DSA results had no impact on graft acceptance or patients’ therapy. Three year follow up was available for all patients.77% of our patients had any HLA-Ab pre transplantation, 55 patients were transplanted with DSA by chance.Acute rejections or ischemic type bile duct lesions (ITBL) were not higher in the DSA group, but ITBL was associated with a higher donor age. There was no difference in long term graft function or survival in patients without HLA-Ab, with non DSA or with DSA (p?=?0.712).We suggest that determination of pre-existing DSA in first LTx recipients is not appropriate and we conclude that pre-existing DSA in first LTx patients are not a contraindication for liver transplantation.     

Ultrasound of living donor liver transplantation

Liver transplantation is the most effective treatment for various end-stage liver diseases. Living donor liver transplantation (LDLT) was first developed in Asia due to the severe lack of cadaveric graft in this region. The Liver Transplant Service at Queen Mary Hospital (QMH), Hong Kong, has pioneered the application of LDLT to patients using both left lobe and right lobe grafts. The QMH liver transplant programme is the largest of its kind in China and Southeast Asia.Ultrasound (US) is often employed in the initial work-up of potential donor and recipient of LDLT. It is the imaging technique of choice to assess the early and late complications of LDLT, with colour Doppler ultrasound being the most useful in the evaluation of post-LDLT vascular complications. The use of ultrasound contrast agents improves the visualisation of the hepatic vasculature, possibly delaying or removing the need for more invasive investigations. Intra-operative ultrasound facilitates the determination of the resection plane during donor hepactectomy.Computed tomography (CT) or magnetic resonance imaging (MRI) can be used as the single imaging modality in the evaluation of LDLT candidates.Ultrasound is most useful as the initial screening test in detecting hepatic parenchymal abnormalities, while CT or MRI is the modality of choice in the demonstration of vascular and biliary anatomy of the potential liver donor.Biliary complications are more common in LDLT than in cadaveric liver transplantation. The ductal dilatation, resulting from biliary stricture, is clearly demonstrated by ultrasound. Bilomas can be aspirated under ultrasound guidance to confirm the diagnosis and to promote healing. Perihepatic fluid collections and abscesses are also common after LDLT. Intra-hepatic collections may represent seromas, haematomas or infarction. Ultrasound is a sensitive means of detecting these collections and can be employed to guide drainage in suitable patients.Transplant-related malignancies include recurrent neoplasia and post-transplant lymphoproliferative disease (PTLD). Ultrasound can be used to screen for recurrent disease and to detect PTLD in the transplanted liver.     

原位肝移植供肝血管及胆道修整

目的探讨原位肝移植供肝血管及胆道系统的修整处理方法。方法回顾性分析31例原位肝移植其供肝动脉变异及胆道内异常情况的处理资料。结果肝动脉变异5例,其中2例肝左动脉来自胃左动脉,2例肝右动脉来自肠系膜上动脉,1例肝总动脉来自肠系膜上动脉。来自肠系膜上动脉的2例肝右动脉,1例吻合到脾动脉的断端;另1例将腹腔干吻合到肠系膜上动脉的近端。供肝的肝内胆管行冲洗时发现有寄生虫2例。结论避免变异的供肝动脉损伤,选择适当的肝动脉吻合方式可以保证移植肝脏的动脉血供。正确的供肝胆道处理,可以减少胆道的并发症。     

Embryology of Extra‐ and Intrahepatic Bile Ducts,the Ductal plate

In the human embryo, the first anlage of the bile ducts and the liver is the hepatic diverticulum or liver bud. For up to 8 weeks of gestation, the extrahepatic biliary tree develops through lengthening of the caudal part of the hepatic diverticulum. This structure is patent from the beginning and remains patent and in continuity with the developing liver at all stages. The hepatic duct (ductus hepaticus) develops from the cranial part (pars hepatica) of the hepatic diverticulum. The distal portions of the right and left hepatic ducts develop from the extrahepatic ducts and are clearly defined tubular structures by 12 weeks of gestation. The proximal portions of the main hilar ducts derive from the first intrahepatic ductal plates. The extrahepatic bile ducts and the developing intrahepatic biliary tree maintain luminal continuity from the very start of organogenesis throughout further development, contradicting a previous study in the mouse suggesting that the extrahepatic bile duct system develops independently from the intrahepatic biliary tree and that the systems are initially discontinuous but join up later. The normal development of intrahepatic bile ducts requires finely timed and precisely tuned epithelial–mesenchymal interactions, which proceed from the hilum of the liver toward its periphery along the branches of the developing portal vein. Lack of remodeling of the ductal plate results in the persistence of an excess of embryonic bile duct structures remaining in their primitive ductal plate configuration. This abnormality has been termed the ductal plate malformation. Anat Rec, 291:628–635, 2008. © 2008 Wiley‐Liss, Inc.     

Expression of cytokeratin 7 and 20 in pathological conditions of the bile tract

Expression of cytokeratin 7 (CK7) and cytokeratin 20 (CK20) helps to establish the origin of biliary and metastatic carcinomas. We investigated the expression of CK7 and CK20 in inflammatory, metaplastic and neoplastic conditions of the bile ducts, and evaluated possible relationships between the CK expression pattern and extrahepatic bile duct/gallbladder carcinomas (EBDCs) or intrahepatic bile duct carcinomas (IBDCs). We used immunohistochemistry for the investigation of 48 formalin-fixed, paraffin-embedded specimens grouped as: A) lithiasic or inflamed surgically resected extrahepatic bile ducts/gallbladders: all were CK7+/CK20+; B) percutaneous liver biopsies from patients with chronic hepatitis C primary biliary cirrhosis and primary sclerosing cholangitis: all were CK7+/CK20-; C) EBDCs: all were CK7+/CK20+, except for two cases which were CK7-/CK20-; D) IBDCs: all were CK7+/CK20-, except for one case showing CK20 positivity. Metaplastic changes were seen only among specimens in groups A and C: in these cases, CK20 was either focally or diffusely expressed. Our study suggests that the expression of cytokeratins under specific stimuli can be different from normal tissues, and that sometimes CK20 expression can be related to and precede the occurrence of metaplastic alterations.     

A cytokeratin immunohistochemical study of cholestatic liver disease: evidence that hepatocytes can express 'bile duct-type' cytokeratins

A cytokeratin immunohistochemical study was performed on 38 liver biopsies from cases of primary biliary cirrhosis, primary sclerosing cholangitis, extrahepatic biliary obstruction or drug-induced liver disease in order to analyse the cytoskeletal changes in detail. On paraffin sections of 27 cases, a variable number of hepatocytes were reactive with a polyclonal anti-cytokeratin antiserum that, in the normal liver, stains bile duct cells only. On cryostat sections of 23 cases, a variable number of hepatocytes were immunoreactive with a monoclonal antibody specifically directed against cytokeratin no. 7 and were most numerous in cases of long-standing cholestasis irrespective of the aetiology. In three cases of primary sclerosing cholangitis and two cases of primary biliary cirrhosis a few hepatocytes were also weakly positive with a monoclonal antibody specific for cytokeratin no. 19. Since cytokeratins no. 7 and no. 19 are, in the normal liver, restricted to bile duct cells, these results further support the concept of 'ductular metaplasia' of hepatocytes, the mechanism of which remains unclear.     

Characterization of human extrahepatic biliary duct epithelial cells in culture

In order to study human bile duct cells in vitro, cystic ducts were obtained during cholecystectomy and treated with collagenase and mechanical abrasion to isolate biliary epithelial cells. The culture medium was supplemented with 50% of a bovine bile duct conditioned medium obtained by incubating minced bovine extrahepatic bile ducts for 24 hr in Dulbecco's modified Eagle's medium. Cells grew in monolayer and showed contact inhibition at confluency. The epithelial origin of primary cultures was verified by their growth pattern, ultrastructure, and indirect immunofluorescence for cytokeratin. The cultures showed specific immunofluorescence for lysozyme, collagen types I, III, and IV, fibronectin, and laminin, but were negative for collagen type V and factor VIII-associated antigen. Thus, these cultures provide an experimental model for the in vitro study of biliary atresia and other bile duct diseases.     

30%小体积肝移植大鼠模型的手术技巧及改良

背景：活体肝移植显著缓解了供肝短缺矛盾,如何使供者献出的肝量最小,同时受者得到最大的临床受益成为了目前的研究热点。 目的：探索一种简便、稳定的小体积肝移植大鼠模型的建立方法。 方法：以Kamada“二袖套法”非动脉化原位肝移植SD大鼠模型为基础并参考国内外文献,制作30%小体积肝移植模型。实验分为Ⅰ组为体内肝叶减除;Ⅱ组为体外肝叶减除组;Ⅲ组为改良后小体积肝移植模型。主要观察供体手术各阶段时间;移植后并发症和生存情况。 结果与结论：改良后模型较改良前的供肝获取时间和肝叶减除时间均显著缩短(P< 0.05)。Ⅱ组的供肝冷缺血时间显著长于Ⅰ组和Ⅲ组(P < 0.05)。改良后的小体积肝移植模型手术成功率较改良前显著提高(P< 0.05),改良后7及14 d存活率均高于改良前模型(P < 0.05)。改良模型较改良前生存时间延长了9 d。改良模型的手术总并发症例次较改良前组显著较少(P < 0.01)。提示采用此改良方法能建立稳定的30%小体积肝移植大鼠模型,而且在操作简便、减少并发症、提高手术成功率和术后存活率等方面有显著优势。     

Claudin-4 differentiates biliary tract cancers from hepatocellular carcinomas.

The recently identified claudins are dominant components of tight junctions, responsible for cell adhesion, polarity and paracellular permeability. Certain claudins have been shown to have relevance in tumor development, with some of them, especially claudin-4, even suggested as future therapeutic target. The aim of the present study was to analyze the expression of claudin-4 in the biliary tree, biliary tract cancers and hepatocellular carcinomas. A total of 107 cases were studied: 53 biliary tract cancers, 50 hepatocellular carcinomas, 10 normal liver and 10 normal extrahepatic biliary duct samples. Immunohistochemical analysis was performed on conventional specimens and on tissue microarrays as well. Claudin-4 was further investigated by Western blot analysis and real-time RT-PCR. Intense membranous immunolabeling was found for claudin-4 in all biliary tract cancers unrelated to the primary site of origin, namely intrahepatic, extrahepatic or gallbladder cancers. Normal biliary epithelium showed weak positivity for claudin-4. In contrast, normal hepatocytes and tumor cells of hepatocellular carcinomas did not express claudin-4. The results of Western immunoblot analysis and real-time RT-PCR were in correlation with the immunohistochemical findings. Cytokeratins, as CK7 (92%) and CK19 (83%) were mostly positive in biliary tract cancers, however, one-third of hepatocellular carcinomas also expressed CK7 (34%). HSA antibody (HepPar1) reacted with the majority of hepatocellular carcinomas (86%), while being positive in a low percentage of the biliary tract cancers (8%). In conclusion, this is the first report of a significantly increased claudin-4 expression in biliary tract cancers, which represents a novel feature of tumors of biliary tract origin. Claudin-4 expression seems to be a useful marker in differentiating biliary tract cancers from hepatocellular carcinomas and could well become a potential diagnostic tool.