Clinical problems with developmental anomalies of the biliary tract

Cholestatic jaundice defined as conjugated hyperbilirubinemia is a typical feature of neonatal liver disease. Biliary atresia is the most common disorder producing cholestasis during the first 2 months of life. Syndromic and non-syndromic paucity of the intralobular bile ducts and choledochal cysts can also present with cholestasis during early life. Liver dysfunction from obstruction of the biliary tree must be differentiated from numerous disorders affecting hepatocytes such as congenital infection and inborn errors of metabolism. Early recognition and a stepwise diagnostic evaluation of the cholestatic infant are essential in successfully treating many metabolic and infectious liver diseases of the infant as well as surgically relieving obstruction in patients with biliary atresia.     

CD56 as a useful marker in the regenerative process of the histological progression of primary biliary cirrhosis

OBJECTIVE: Owing to recent contradicting results in the study of the regenerative process after hepatic injury in primary biliary cirrhosis, we investigated the use of CD56 in tissue repair during the histological progression of primary biliary cirrhosis. METHODS: Fifty-three specimens were classified into Ludwig's stages (1-4) as follows: 14 specimens as stage 1, 23 as stage 2, 14 as stage 3, and two as stage 4. Immunohistochemical stain was performed for CD56. The cell types expressing the marker were morphologically analyzed to determine their origin. RESULTS: In normal liver biliary epithelial cells (including the epithelium of terminal bile ducts and bile ductules), hepatocytes, and intermediate cells (features between hepatocytes and biliary cells, distributed in interface between hepatic parenchyma and portal tract) were CD56. In primary biliary cirrhosis specimens, biliary epithelial cells, hepatocytes, and intermediate cells were CD56 distributed as 10 out of 14 cases as stage 1 (71.43%), 18 out of 23 as stage 2 (78.26%), nine out of 14 as stage 3 (64.28%), and two out of two as stage 4 (100%). The total positive cases were 39 of 53 (73.58%). CD56 was expressed equally in all three types of cells. CONCLUSION: These findings indicate that the consistent and uniform expression of CD56 in biliary epithelial cells, hepatocytes, and intermediate cells during hepatic injury in primary biliary cirrhosis is probably related to cellular damage and may be important in tissue regeneration. Furthermore, we cannot distinguish a specific cell type from the three above mentioned ones (biliary epithelial cells, hepatocytes, intermediate cells) as a putative stem cell in primary biliary cirrhosis.     

Human hepatic stem-like cells isolated using c-kit or CD34 can differentiate into biliary epithelium

BACKGROUND & AIMS: Recent reports suggest that after bone marrow transplantation into rodents and humans, hematopoietic stem cells migrate into the liver and give rise to oval cells, hepatocytes, and biliary epithelial cells. We investigated this hypothesis further in the human liver using the hematopoietic markers c-kit and CD34. METHODS: Immunofluorescence confocal microscopy was performed using cytokeratin 19 (CK-19; biliary cell marker) with either c-kit or CD34. Immunomagnetic separation was then used to select c-kit- or CD34-positive cells. After attachment, cells were cultured for up to 7 days, and their growth and phenotypic characteristics were examined. RESULTS: In cirrhotic tissue, c-kit- or CD34-positive cells were located in the portal tracts surrounding bile ducts. Occasionally c-kit- (but not CD34-) positive cells that coexpressed CK-19 were observed integrated into bile ducts. In vitro, immunoisolated c-kit or CD34 cells gave rise to colonies of at least 2 morphologies expressing CK-19 or CD31 (endothelial cell marker). CD34- or c-kit-positive cells with similar properties were also isolated from normal liver. CONCLUSIONS: These findings indicate that cells present in human liver that express the markers c-kit or CD34 have the capacity to differentiate into biliary epithelial cell lineage and may therefore represent human biliary epithelial progenitor cells.     

Endoscopic retrograde cholangiography in the diagnosis of biliary malformations in infants

The differentiation of infantile biliary malformations from primary parenchymal diseases is difficult. The recent development of a pediatric side-viewing endoscope (PJF Endoscope; Olympus Corporation of America) provided an opportunity to investigate the usefulness of endoscopic retrograde cholangiography (ERC) for precise visualization of the extrahepatic biliary passages in infants with persistent cryptogenic cholestasis. ERC was performed in 12 patients, with visualization of the existing extrahepatic bile ducts in 4. The entire biliary system was visualized in one, excluding extrahepatic biliary atresia and choledochal cyst. The reduced caliber of the intrahepatic bile ducts and histological observations in a percutaneous liver biopsy supported the diagnosis of intrahepatic biliary hypoplasia in this case. An intact hepatic portochole cystostomy was documented in one, although the intrahepatic biliary system was not delineated. Atresia of the hepatic bile ducts proximal to the gallbladder was documented in two. Of the eight patients in whom extrahepatic bile ducts were not visualized by ERC, six had extrahepatic biliary atresia confirmed at exploratory laparotomy. The papilla of Vater could not be located in four of these six infants. The remaining two had neonatal hepatitis. ERC may offer a useful alternative to operative cholangiography in selected infants with persistent cholestasis and acholic stools.     

Effector role of neonatal hepatic CD8+ lymphocytes in epithelial injury and autoimmunity in experimental biliary atresia

BACKGROUND & AIMS: Lymphocytes populate the livers of infants with biliary atresia, but it is unknown whether neonatal lymphocytes regulate pathogenesis of disease. Here, we investigate this question by examining the role of T lymphocytes in the destruction of extrahepatic bile ducts of neonatal mice using an experimental model of biliary atresia. METHODS: Inoculation of neonatal mice with rhesus rotavirus followed by multistaining flow cytometry to quantify expression of interferon-gamma by hepatic lymphocytes, and real-time polymerase chain reaction for mRNA expression of pro-inflammatory cytokines. This was followed by determining the consequences of antibody-mediated depletion of lymphocyte subtypes on the development of biliary obstruction, and coculture and cell transfer experiments to investigate the effector role of lymphocyte subtypes on neonatal biliary disease. RESULTS: Rotavirus infection results in overexpression of interferon-gamma by neonatal hepatic T cells. Among these cells, depletion of CD4(+) cells did not change the course of inflammatory injury and obstruction of neonatal bile ducts. In contrast, loss of CD8(+) cells remarkably suppressed duct injury, prevented luminal obstruction, and restored bile flow. Coculture experiments showed that rotavirus-primed, but not na?ve, CD8(+) cells were cytotoxic to cholangiocytes. In adoptive transfer experiments, we found that primed CD8(+) cells preferentially homed to extrahepatic bile ducts of neonatal mice and invaded their epithelial lining. CONCLUSIONS: Primed neonatal CD8(+) cells can activate a pro-inflammatory program, target diseased and healthy duct epithelium, and drive the phenotypic expression of biliary atresia, thus constituting a potential therapeutic target to halt disease progression.     

正常胰胆管汇合患者胆汁淀粉酶升高与胆道疾病关系的研究

目的探讨正常胰胆管汇合（NPBJ）者胆汁淀粉酶升高与胆道疾病的关系。方法连续202例患者（胆管胆汁组）在内镜治疗胆道疾病时抽取胆管胆汁检测淀粉酶,其中68例同时检测胆汁脂肪酶,149例做胆汁细菌培养,27例测Oddi括约肌压力（SOM）,38例测胆管压力。另外73例（胆囊胆汁组）经皮经肝胆囊镜治疗胆囊结石,取胆囊中胆汁检测淀粉酶,31例进行胆囊黏膜活检。两组病例均除外先天性胰胆管汇合异常、胆肠吻合术和既往内镜乳头切开治疗者。结果胆管胆汁组95例（47．0％）淀粉酶升高,其中肿瘤（56．9％,29／51）与非肿瘤疾病（43．7％,66／151）差异无统计学意义（P〈0．05）,但肝门部胆管癌大多数（7／9）淀粉酶值升高;胆汁脂肪酶水平与淀粉酶有明显的相关性（r=0．561）;淀粉酶水平与Oddi括约肌和胆管压力无明显关联;胆汁细菌培养阳性率在淀粉酶值正常和升高者之问无明显差异。胆囊胆汁组34．3％淀粉酶升高,其中87．5％见胆囊上皮细胞异型增生,与淀粉酶正常者差异有统计学意义（P〈0．01）。结论NPBJ胆道疾病患者胰液向胆管逆流发生频度较高,淀粉酶升高者中肿瘤和非肿瘤疾病无显著差异,但淀粉酶升高者胆囊上皮细胞异型增生和肝门部胆管癌发生频度高。     

Phenotypic change in portal fibroblasts in biliary fibrosis

Portal fibroblasts have been considered responsible for biliary fibrosis. Since lipocytes show differentiation toward myofibroblast-like cells in hepatic fibrogenesis, we studied whether similar differentiation of portal fibroblasts could be observed in biliary fibrosis. We examined rat livers after bile duct ligation by double immunofluorescent staining of α-smooth-muscle actin (α-smA) and desmin and also by electronmicroscopy. In the portal tract of normal livers, α-smA-positive cells were noted only in the vessel wall, whereas desmin-positive cells were occasionally seen in the connective tissue as well. With the development of biliary fibrosis, α-smA was remarkably expressed in the portal connective tissue, while desmin was seen in a small portion of α-smA-positive cells around proliferating bile ducts. In normal livers, portal fibroblasts presented quiescent features, such as a small Golgi complex and a few cisternal profiles of endoplasmic reticulum under electron microscopy. After 7 days of bile duct ligation, portal fibroblasts proliferated, were arrayed in multilayers, and were associated with collagen bundles. Some of these fibroblasts had numerous cytoskeletal components, and developed rough endoplasmic reticulum and a dense body. These data suggest that portal fibroblasts appear to differentiate toward myofibroblasts in biliary fibrosis.     

Primary biliary cirrhosis: modalities of injury and death in biliary epithelium

BACKGROUND/AIM: Despite the number of studies on primary biliary cirrhosis, contrasting data remain concerning modalities of cholangiocyte death. Liver biopsies obtained from 40 patients with anti mitochondrial antibody-positive primary biliary cirrhosis, at various stages of the disease, were examined, and special attention was paid to the expression of subcellular damage and evidence of apoptosis. METHODS: Liver sections were stained with haematoxylin/eosin or Sirius red. Ductular mass was evaluated on sections after cytokeratin 7 staining. Apoptosis was evaluated on haematoxylin/eosin stained material or after processing for terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling assay. In 16 patients, part of the biopsy was processed for electron microscopy. Twenty histologically normal liver biopsies were used for control purposes. RESULTS: According to Scheuer's classification, 29 patients were classified as stage I-II, and 11 as stage III-IV. A strong staining of bile ducts was evident after immunohistochemistry for cytokeratin 7, often associated with ductular metaplasia in lobular zone 1. Cytokeratin 7-positive cells occupied 3.0+/-1.3% of liver mass as compared to 0.25+/-0.03% in controls. Ductular metaplasia accounted for 1.4+/-0.07% of all cytokeratin 7-positive cells. Regardless of staging, apoptotic bodies were seen only exceptionally in epithelial wall of bile ducts, whereas cholangiocyte damage leading to extensive lytic necrosis appeared responsible for most of the bile duct mass loss, as also confirmed by ultrastructural studies. A few terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling-positive nuclei were occasionally associated with the inflammatory infiltrate and evidence of apoptosis in hepatocytes was frequent, especially in zone 1. CONCLUSION: Regardless of staging, lytic necrosis and not apoptosis accounts for most of the bile duct loss in primary biliary cirrhosis. Furthermore, ductular metaplasia appears as a late event with highly variable pattern being observed between patients.     

肝移植术后胆道并发症的临床及病理分析

目的探讨原位肝移植术后胆道并发症的常见病理组织学及临床特点.方法回顾性分析1998年9月-2005年6月的肝移植术后胆道并发症患者173例（235例次）肝穿刺活组织检查的病理和临床资料.结果肝移植后胆道并发症发生于术后3～2 920 d,其中1～30 d、31～90 d、91～180 d、180 d以后的发生率分别为49.71%（86/173）、17.92%（31/173）、4.62%（8/173）、27.74%（48/173）.炎症类占72.25%（125/173）,行肝穿刺活组织检查171例次;梗阻类占27.74%（48/173）,行肝穿刺活组织检查64例次.病理表现以小叶间胆管上皮细胞变性及炎细胞浸润、汇管区炎症、小胆管增生、汇管区纤维化、小胆管及肝细胞胆汁淤积为主.上述病理表现在炎症和梗阻两类并发症患者中的检出率分别为100.00%（171/171）和100.00%（64/64）、100.00%（171/171）和96.87%（62/64）、9.36%（16/171）和73.44%（47/64）、3.51%（6/171）和79.69%（51/64）、50.29%（86/171）和87.50%（56/64）、63.16%（108/171）和93.75%（60/64）.结论原位肝移植术后胆道并发症以炎症类居多,多发生于术后30 d内,梗阻类多见于术后90 d,预后较差.肝穿刺活组织检查的病理组织学表现在胆道并发症的分类、程度评估及鉴别诊断中具有重要价值.     

Expression of collagens type I and IV, osteonectin and transforming growth factor beta-1 (TGFbeta1) in biliary atresia and paucity of intrahepatic bile ducts during infancy.

BACKGROUND/AIMS: Biliary atresia and paucity of intrahepatic bile ducts are the main causes of neonatal cholestasis leading to hepatic fibrosis. Fibrotic evolution is slow in paucity of bile ducts as compared to the rapid progression to biliary cirrhosis in biliary atresia when cholestasis persists despite hepatoportoenterostomy. Our aim was to compare the expression of collagens type I and IV, alpha-smooth muscle actin, osteonectin and transforming growth factor beta1 in biliary atresia and paucity of bile ducts. METHODS: Liver biopsies were obtained in 12 children with biliary atresia and in five with paucity of bile ducts. Collagens type I and IV, alpha-smooth muscle actin were detected with immunostaining. Collagens type I and IV, osteonectin and transforming growth factor beta1 mRNAs were detected by in situ hybridization. RESULTS: Expression of mRNA and proteins was roughly parallel. In ductular proliferation areas of biliary atresia: (1) the expression of collagens type I and IV and osteonectin was increased, and was localized to periductular myofibroblasts; (2) transforming growth factor beta1 was expressed around biliary ductules, probably in inflammatory cells, and also in biliary cells. Osteonectin expression was also increased in the lobules. In paucity of bile ducts, there was no overexpression of collagens type I and IV and transforming growth factor beta1, except in the only child with marked fibrosis. However, osteonectin expression was enhanced at the periphery of the lobules, even when fibrosis was mild or absent. CONCLUSIONS: These findings suggest that in biliary atresia ductular proliferation areas are the site of a marked production of extracellular matrix proteins in periductular myofibroblasts, probably secondary to transforming growth factor beta1 production by inflammatory cells and by biliary cells. The weak expression of transforming growth factor beta1 could explain the slow progression of fibrosis in paucity of bile ducts.     

Assessment of a postoperative anastomotic stricture following correction surgery of a type IVa choledochal cyst using Gd-EOB-DTPA-enhanced magnetic resonance cholangiography

Choledochal cyst is a relatively uncommon disease which is characterized by congenital dilatation of the intra and/or extrahepatic part of the biliary tree. Type IVa choledochal cysts are managed surgically through total excision of the entire extrahepatic part of the abnormal bile ducts and a simultaneous hepaticoenterostomy. Postoperative anastomotic stricture after excision of choledochal cysts and hepaticojejunostomy is a well-known late complication. We report a case of a 17-year-old female in whom gadoxetic acid-enhanced magnetic resonance cholangiography assisted in the evaluation of a biliary stricture following bile duct procedures after choledochal cyst correction surgery.     

经皮肝穿刺胆道内支架植入治疗恶性胆道梗阻64例

目的：总结经皮肝穿刺胆道内支架植入治疗恶性梗阻性黄疸的经验，探讨其临床疗效及价值。方法：64例患者均采用X线透视下经皮肝穿刺胆道内支架植入术治疗恶性梗阻性黄疽。根据梗阻部位的不同解剖决定放置支架的方式。结果：64例患者中，50例植入单支支架于肝总管或(和)胆总管，14例植入2支以上支架于总管和分支胆管，其中2例肝内胆管支架的桥接通过肝实质。58例患者2周内血清胆红素降低75％以上。结论：经皮肝穿刺刺内支架植入是治疗恶性胆道梗阻性的有效方法。     

Expression of anti-OV6 antibody and anti-N-CAM antibody along the biliary line of normal and diseased human livers

Following hepatic injury, proliferation of anastomosing ductules can be observed. The origin of this ductular reaction is not completely clear, although there is considerable evidence for proliferation of a putative hepatic progenitor cell, reported to be located in the canals of Hering (CoH) and showing morphologic similarities with rat oval cells. In this study, we analyzed the immunophenotype of solitary oval cell-like cells (SOC), intralobular groups of cuboidal cells that might represent lining cells of CoH, bile ductular cells (BDC), bile duct epithelial cells (BEC), and hepatocytes. We used the antibodies OV6, CK19, and CD56 (NCAM) in a double-staining method in a series of 111 liver specimens. The series consisted of a variety of liver diseases, primary liver tumors, and normal livers. In normal livers, SOC, CoH, BDC, and BEC were uniformly and predominantly CK19+, OV6+, and CD56-. In diseased livers SOC and BDC were CK19+, OV6+, and also CD56+. Occasionally, BEC was CD56+ in damaged bile ducts in diseased liver, e.g., PSC. CoH lining cells were not present in cirrhotic nodules and were indistinguishable from BDC in the fibrous septa. The consistent and uniform staining patterns of SOC, CoH, and BDC support the concept that these cells share the same biliary lineage and might represent one biliary structure. The expression of CD56 on these cells in diseased livers indicates that CD56 is a useful marker for a reparative or regenerative state of the biliary liver-cell constituents but not to discriminate a putative hepatic stem cell.     

Endoscopic biliary manometry in patients with suspected sphincter of Oddi dysfunction and in patients with cystic dilatation of the bile ducts

We studied the motility of the sphincter of Oddi in 12 patients with suspected sphincter of Oddi dysfunction, in four patients with cystic dilatation of the bile ducts (two Caroli's cases and two fusiform choledochal cyst cases), and in 33 patients with retained common duct stones. In these last 33 patients, the motor activity of the sphincter of Oddi was similar to that recorded in nine control subjects without pancreatic or biliary diseases. In the suspected Oddi dysfunction cases, both the basal sphincteric pressure and the frequency of the phasic contractions were significantly elevated (P<0.001). Patients with biliary cystic dilatation showed an increased basal pressure, but the frequency of the contractions was elevated in only those with choledochal cysts and the amplitude in only one of the two patients with Caroli's disease. Motor disorders of the sphincter of Oddi provide a basis for an alternative etiopathogenesis of cystic disease of the biliary system and a possible explanation for pain and dilatation of the bile duct in patients with suspected sphincter of Oddi dysfunction.     

Increased secondary bile acids in a choledochal cyst. Possible role in biliary metaplasia and carcinoma

Choledochal cysts are uncommon congenital or acquired lesions of the biliary tree. The incidence of biliary tract carcinoma in patients with choledochal cysts is 5-35 times greater than that of the general population. Factors responsible for the increased risk of carcinoma are unknown. The case of a young woman who underwent excision of a choledochal cyst 16 years after initial diagnosis and treatment by choledochocystduodenostomy is reported. Metaplasia of the epithelial lining of the cyst was found in the resected specimen. The relative composition of bile acids in cyst contents was as follows: lithocholate, 2%; deoxycholate, 88%; chenodeoxycholate, 5%; and cholate, 5%. Virtually all bile acids were recovered in unconjugated form. In contrast, the bile acid composition of hepatic bile was as follows: lithocholate, 0%; deoxycholate, 34%; chenodeoxycholate, 43%; and cholate, 23%. Bile acids were fully conjugated. These data suggest that stasis of bile within choledochal cysts contributes to bacterial overgrowth and generation of unconjugated secondary bile acids.     

胆道闭锁患儿肝脏组织c-kit+细胞的表达及意义

目的探讨胆道闭锁患儿肝脏组织c-kit+细胞表达及其与肝脏纤维化程度与预后的关系.方法20例胆道闭锁患儿肝脏组织标本,应用免疫组化PV6000二步法染色检测c-kit+细胞.结合常规病理检查进行纤维化程度分级.根据临床资料将患儿分为预后良好组与预后不良组,并取非肝胆疾病新生儿肝脏组织作为正常对照.结果胆道闭锁患儿c-kit+细胞数显著多于正常对照组(P＜0.01);预后不良组c-kit+细胞数显著多于预后良好组(P＜0.05).纤维化程度较重组c-kit+细胞数显著多于纤维化程度较轻组(P＜0.01).结论c-kit+细胞参与胆道闭锁患儿肝脏损害、纤维化等病理过程,c-kit+细胞过度表达与胆道闭锁患儿肝脏的纤维化程度及预后密切相关.     

三维磁共振胰胆管成像诊断小儿胆道闭锁价值分析

目的 研究磁共振成像(MRI)三维磁共振胰胆管成像(3D-MRCP)诊断小儿胆道闭锁(BA)的临床价值.方法 2016年4月～2020年4月我院收治的疑似BA患儿24例,接受常规MRI和3D-MRCP检查.对患儿采取Kasai术,以手术后组织病理学检查结果为金标准,判断3D-MRCP诊断BA的效能.结果 24例疑似BA...