乙型肝炎相关慢加急性肝衰竭合并脓毒症的病理特征

目的观察乙型肝炎相关慢加急性肝衰竭(HBV-ACLF)合并脓毒症肝脏病理改变。方法选择50例HBV-ACLF肝移植患者的移植前肝脏标本,制成石蜡切片,经HE染色、Masson三色染色和抗细胞角蛋白-7(CK-7)免疫组织化学染色,镜下观察肝脏病理变化。结果 HBV-ACLF合并脓毒症的病理改变为在肝硬化基础上发生大块/亚大块坏死,部分肝硬化结节残留,结节边缘细胆管扩张,腔内有浓缩胆汁,细胆管上皮细胞萎缩甚至消失。结论 HBV-ACLF合并脓毒症时肝脏表现为残留结节边缘细胆管胆汁淤积,但需要注意与其他类型的胆汁淤积鉴别。     

HBV所致终末期肝病患者肝组织胆管增生的研究

目的在乙型肝炎病毒（HBV）所致的终末期肝病患者肝组织内，常可见大量的胆管增生，但其发生机制及临床意义尚不清楚。为阐明该类患者胆管增生的发生机制及其与卵圆细胞增生及肝细胞再生的关系，我们对8例HBV相关的终末期肝病患者及2例正常人肝组织进行了免疫组化染色及图像分析。方法肝组织连续切片后进行免疫组化染色，观察指标为细胞角蛋白CK7、CK8、CK18、CK19、OV6、增殖细胞核抗原（PCNA）、谷胱甘肽S转移酶（GSTπ）、白蛋白（ALB）及甲胎蛋白（AFP）。结果在所有8例患者肝组织汇管区内均可见典型增生的胆管及非典型增生的胆管，且对CK7、CK8、CK9、CK19、OV6及PCNA染色呈阳性反应，但两种类型的胆管在染色强度上存在明显差异。一些非典型增生的胆管细胞表现出肝卵圆细胞的形态学及免疫组化特征。某些小型肝细胞样细胞在形态及免疫组化特征方面介于肝卵圆细胞及成熟肝细胞之间。结论在HBV相关的终末期肝病患者肝组织内，胆管增生可能存在不同起源。某些非典型增生的胆管细胞实际上就是活化的肝卵圆细胞。非典型增生的胆管细胞与肝细胞再生密切相关。小型肝细胞样细胞可能是肝卵圆细胞与成熟肝细胞之问的中间过渡细胞。     

慢性重型肝炎患者肝组织内胆管增生的免疫组织化学研究

在慢性重型肝炎患者的肝组织切片中,除肝细胞大面积坏死、纤维组织增生、炎症细胞浸润及不同程度的肝细胞再生等病理特征外,还常有明显的胆管增生现象,但这种胆管增生的本质及意义尚不清楚。新近提出肝组织内胆管增生与肝干细胞     

大鼠胆汁淤积性肝硬化的动态病理变化及其意义

目的：观察胆汁淤积性大鼠肝硬化形成过程中模型动态变化特点。方法：结扎大鼠胆总管制备大鼠胆汁淤积性肝硬化模型；结扎后3天、1周、2周、3用、4周、5周后杀鼠取材，观测大鼠一般状况，肝功能、肝组织羟脯氨酸（Hyp）含量测定，肝组织病理学及CK-19免疫组织化学观察。结果：与假手术组大鼠比较。模型组大鼠肝重、脾重、肝／体比、脾／体比均呈先增加后降低的动态变化；血清Alb含量持续下降，而TP含量呈先高后低的变化；血清ALT活性在结扎3天后最高，2周时最低；GGT活性及TBil含量自造模后呈持续增高；而ALP活性在2周时最高；肝组织Hyp含量从结扎后呈持续增加；结扎3天后汇管区已见胆管增生，至3周时增生逐渐加重。CK-19免疫组织化学染色观察，胆管上皮细胞第3天起开始增生，1周、2周时增生明显，3周后增生渐减。4周、5周时肝组织呈花环样改变，肝实质细胞显著减少。可见肝细胞去分化现象。结论：①胆汁淤积性大鼠肝硬化模型主要病理改变有胆汁淤积、胆管上皮细胞增生、肝细胞减少、胶原增生和沉积与新生的胆管上皮细胞关系密切。②血清ALT水平由高到低，5周末仅比假手术组稍高。AST活性一直维持在较高水平。③肝细胞的减少方式除了凋亡、坏死外，可见去分化现象。     

e抗原阴性慢性乙型肝炎合并非酒精性脂肪性肝病的病理特征

目的观察HBeAg阴性慢性乙型肝炎合并非酒精性脂肪性肝病患者肝组织病理特征。方法收集72例慢性乙型肝炎合并非酒精性脂肪性肝病患者的肝组织标本及临床相关资料,肝组织标本常规行HE、Masson三色及网状纤维染色,观察其病理改变特点,并进行分级、分期。结果HBeAg阴性慢性乙型肝炎合并非酒精性脂肪性肝病患者肝活检病理学改变以肝细胞脂肪变性为主,肝细胞脂肪变性主要位于腺泡Ⅲ带,呈弥漫分布,重度患者则可扩大至腺泡Ⅱ带甚至全腺泡,轻度患者汇管区炎及界面性炎均不明显,主要为小叶内点灶状坏死,大部分患者仅为血窦壁及中央静脉管壁及周围纤维组织增生,且多为纤细的网状纤维,汇管区纤维组织增生不明显,重度及肝硬化患者则汇管区纤维组织增生显著,彼此相连,破坏正常的肝小叶结构,形成假小叶。结论HBeAg阴性慢性乙型肝炎合并非酒精性脂肪性肝病患者肝组织病理改变具有一定病理学特征;其病理学特征应从炎症、纤维化程度与肝细胞脂肪变性范围、肝细胞损伤等方面综合分析,肝组织活检有助于早期明确诊断,从而指导临床合理治疗。     

磷酸化p38丝裂原活化蛋白激酶在急性肝衰竭小鼠模型及HBV相关慢加急性肝衰竭患者肝组织中的表达及意义

目的研究磷酸化后的p38丝裂原活化蛋白激酶(p-p38MAPK)在氨基半乳糖(D-Gal N)或脂多糖(LPS)诱导的急性肝衰竭小鼠模型以及HBV相关慢加急性肝衰竭(ACLF)患者肝脏中的表达及其意义。方法采用D-Gal N/LPS诱导C57BL/6小鼠构建急性肝衰竭模型,分别在给药0、0.5、1、2、4、6、8 h设立实验组,每组4只,处死小鼠取肝组织标本进行HE染色观察肝组织结构的病理变化,分别运用蛋白免疫印迹技术半定量检测及免疫组化染色定位检测肝组织中p-p38MAPK的表达;同时对照研究pp38MAPK在HBV-ACLF、乙型肝炎肝硬化、慢性乙型肝炎(CHB)患者肝组织中的表达情况。组间比较采用独立样本t检验。结果蛋白免疫印迹法检测p-p38MAPK在肝衰竭小鼠肝组织匀浆中的表达随时间变化持续增高,给药6 h组半定量分析表达量显著高于正常对照组,差异具有统计学意义(t=-2.727,P=0.034)。免疫组化染色结果显示随存活时间的延长,小鼠肝组织炎症程度逐渐加重,炎症早期主要为窦细胞表达p-p38MAPK,随着肝组织损害加重,肝细胞表达p-p38MAPK渐多,坏死肝组织附近分布大量p-p38MAPK阳性肝细胞;正常人肝组织中p-p38MAPK的表达量很低,CHB患者肝组织内可见浸润淋巴细胞及肝细胞表达pp38MAPK,并且随病程进展呈增多趋势,与临床观察病变程度逐渐加重相一致。结论 D-Gal N/LPS诱导的小鼠急性肝衰竭模型中,p-p38MAPK表达随肝组织损害加重,表明其在肝衰竭病变过程中占重要地位;在HBV-ACLF患者致病过程中,p-p38MAPK信号通路可能发挥重要作用。     

慢性重型乙型肝炎的病理形态学表现及诊断

目的观察并总结慢性重型乙型肝炎（CSHB）的病理形态学特点。方法用常规HE染色、组织化学Masson三色、Sweet网织、D-PAS染色及免疫组织化学CK7、CD68、增殖细胞核抗原染色对42例CSHB病例进行回顾性观察和分析。结果CSHB的基本病变是在慢性肝病的背景上出现大块或亚大块坏死，坏死的特点是部分肝硬化结节坏死，部分肝硬化结节保留，保留的硬化结节仍可发生部分肝细胞坏死。坏死后的肝细胞常被清除，而间质成分保留。随着病程的进展，还表现出不同程度的肝细胞再生和细胆管增生。结论CSHB与急性、亚急性重型乙型肝炎的病变表现是一致的，有其特征性的病理形态学改变，对重型肝炎的诊断应结合病因加以分类。     

肝硬化患者高胆红素血症与肝脏病理组织学的关系

目的 为探讨肝硬化高胆红素血症的发病机理，我们对19例肝炎后肝硬化患者的尸检肝组织进行病理组织学观察。方法 据患者生前血清胆红素测定值将19例患者分成3组。血清胆红素低于85.5μmol/L为Ⅰ组，血清胆红素在85.5-171μmol/L之间为Ⅱ组，血清胆红素高于171μmol/L为Ⅲ组。3组例数分别为9，3，7。肝组织切片经HE染色，从癌旁与血清胆红素之间的关系，肝组织硬化程度，肝脏炎症活动程度，肝组织内毛细胆管淤胆程度观察肝组织形态与血清胆红素值之间的关系。结果 （1）3组病例中，轻度肝硬化例数分别为6，2，4例；（2）组织学活动指数（HAI）计分第Ⅰ组6分以下6例（66．7％），第Ⅱ，Ⅲ组均在7分以上，亚大块肝坏死例数分别为0，1，2例；（3）毛细胆管内重度淤胆例数分别为0，1，5例；（4）合并癌变例数分别为3，2，2全。结论 肝炎肝硬化患者合并高胆红素血症程度与肝细胞活动性炎症，坏死程度及肝细胞内毛细胆管淤胆有关，在临床治疗中应兼顾此两种因素，合理安排治疗。     

肝内胆汁淤积发生机制进展

胆汁淤积是指胆汁分泌的阻滞、抑制或胆汁流的障碍,胆汁到达十二指肠减少,导致胆汁成分在血中蓄积。有关肝内胆汁淤积的发生机制,近几年来有了一些新的认识,为胆汁淤积的诊断、治疗及预后估计提供了理论依据。一、肝内毛细胆管及肝内胆管病损原发性胆汁性肝硬化为肝内胆管病损的典型代表。可见叶间胆管出现增生、小叶间胆管壁上皮细胞呈变性、坏死,上皮细胞间有炎症细胞浸润或出现淋巴细胞增殖,在胆管的周围还可见浆细胞浸润。进而毛细胆管出现结     

肝内胆管改变与肝纤维化的关系

肝硬化是由多种原因引起的以弥漫性纤维组织增生、肝小叶结构破坏、假小叶形成为特征的肝病。在多种肝硬化动物模型及不同病因肝硬化患者中均可见胆管增生。多种神经肽、神经递质及激素等调节因子参与的信号通路调控胆管增生。增生的胆管通过介导星状细胞增殖、活化,促进肝纤维化的形成。总结了肝硬化时肝内胆管系统的改变及其对纤维化进程的影响,胆管细胞增生与肝纤维化相关的信号通路,以及胆管结构的动态演变对肝纤维化程度的预测价值。提出胆管增生可能成为干预肝纤维化的潜在靶点,为早期治疗及逆转肝纤维化提供新的思路和方法。     

Ductular proliferation in liver tissues with severe chronic hepatitis B： An immunohistochemical study

AIM: To clarify the pathogenesis of ductular proliferation and its possible association with oval cell activation and hepatocyte regeneration. METHODS: Immunohistochemical staining and image analysis of the ductular structures in the liver tissues from 11 patients with severe chronic hepatitis B and 2 healthy individuals were performed. The liver specimens were sectioned serially, and then cytokeratin 8 (CK8), CK19, OV6, proliferating cell nuclear antigens (PCNA), glutathione-S-transferase (GST),α-fetal protein (AFP) and albumin were stained immunohistochemically. RESULTS: Typical and atypical types of ductular proliferation were observed in the portal tracts of the liver tissues in all 11 patients. The proliferating ductular cells were positive for CK8, CK19, OV6 and PCNA staining. Some atypical ductular cells displayed the morphological and immunohistochemical characteristics of hepatic oval cells. Some small hepatocyte-like cells were between hepatic oval cells and mature hepatocytes morphometri-cally and immunohistochemically. CONCLUSION: The proliferating ductules in the liver of patients with severe chronic liver disease may have different origins. Some atypical ductular cells are actually activated hepatic oval cells. Atypical ductular proliferation is related to hepatocyte regeneration and small hepatocyte-like cells may be intermediate transient cells between hepatic oval cells and mature hepatocytes.     

肝硬化中肝脏祖细胞的激活与扩增

目的证实肝硬化组织中存在人肝脏祖细胞（HPCs），探讨HPCs分布及激活的强度与肝脏炎症程度的关系，提供HPCs向肝细胞分化的依据。方法对30例肝硬化及3份正常组织标本进行常规组织学观察，对门静脉炎症程度进行评分，并用胆管上皮标志物（细胞角蛋白7）和肝星状细胞激活标志物（a平滑肌肌动蛋白）进行免疫组织化学染色，对符合HPCs、中间型肝细胞以及小管样反应的细胞进行计数和半定量评分。结果在正常肝组织中无门静脉周围HPCs和小管样反应增殖。在肝硬化组织中，增殖的HPCs起源于肝门静脉区域，随着门静脉炎症程度加重，HPCs及小管样反应从肝硬化结节周围向肝实质扩散并出现中间型肝细胞增殖，其周围有显著的肝星状细胞激活。HPCs及小管样反应增殖程度随着门静脉炎症程度的加重而增加。HPCs数目与中间型肝细胞数目之间存在直线正相关。丙氨酸氨基转移酶和天冬氨酸氨基转移酶与HPCs及中间型肝细胞数目存在直线正相关。结论在人肝硬化中存在祖细胞的激活，炎症反应是HPCs激活的触发因素，HPCs向肝实质内迁移并向肝细胞方向分化是肝再生的重要途径。     

Cell-kinetic study of proliferating bile ductules in various hepatobiliary diseases

ABSTRACT— Aims and Methods: Proliferat bile ductules are classifiable histologically into typical and atypical types. To clarify their histogenesis and regulation, we examined their phenotype, proliferating and degrading characteristics, using liver sections from 58 patients with various hepatobiliary diseases. Results: Typical ductules were found in all cases. Atypical ductules were also frequently found in extrahepatic biliary obstruction (EBO), chronic hepatitis (CH), as well as in primary biliary cirrhosis (PBC) and primary sclerosing cholangitis (PSC). Typical ductules completely expressed biliary-type cytokeratins, while atypical ductules lacked complete biliary-type cytokeratins and often connected with periportal hepatocytes. Proliferative indices of typical ductules in diseased livers were higher than those in normal livers, while those of atypical ductules were low in PBC and PSC and high in EBO and CH. Apoptosis was detected in typical and atypical ductules. Perforin was preferably expressed on typical and atypical ductules, compared with CD95. Conclusions: These findings suggest that typical ductules reflect active proliferation of preexisting ductules. Atypical ductules might be classifiable into two categories: those in PBC and PSC primarily reflect ductular transformation (metaplasia) of periportal hepatocytes, while those in EBO and CH reflect active proliferation and transformation of hepatocytes. Apoptosis via perforin/granzyme B pathway may be involved in the maintenance of homeostasis in ductular proliferation as degrading fraction.     

Immunohistochemical evidence for hepatic progenitor cells in liver diseases

BACKGROUND/AIM: Proliferative bile ductular reactions occur in a variety of liver diseases in humans. It is a matter of debate whether such reactions result from progenitor cell proliferation with biliary and hepatocytic differentiation, versus biliary metaplasia of damaged hepatocytes. We investigated bile ductular reactions in liver diseases, paying particular attention to the presence of cells with intermediate (hepatocytic/biliary) features (oval-like cells). METHODS: Five specimens each were selected of submassive hepatic necrosis and cirrhosis due to hepatitis B, hepatitis C, autoimmune hepatitis, alcohol injury, primary biliary cirrhosis and primary sclerosing cholangitis. Immunohistochemical stains were performed for biliary markers (cytokeratins [CKs] 7 and 19), as well as hepatocytic markers (HepParl and alpha-fetoprotein[AFP]) in sequential sections. The degree of staining of each cell type (biliary, hepatocytic, intermediate) was graded semiquantitatively. RESULTS: Hepatocytes always stained diffusely for HepParl, occasionally for CK7, and rarely for CK19. Biliary cells were always diffusely positive for CK7 and CK19, and rarely for HepParl. Intermediate cells were identified in all cases and showed widespread staining for both HepParl and CK7, and less commonly for CK19. AFP was not expressed in any cell type. The morphologic and immunohistochemical features of bile ductular reactions were similar in the different diseases. CONCLUSIONS: Proliferating hepatic parenchymal cells with intermediate (hepatocytic/biliary) morphologic features and combined immunophenotype can be identified in a variety of acute and chronic liver diseases. The similarity of bile ductular reactions among chronic hepatitic, alcoholic and biliary diseases suggests that they result from proliferation of oval-like progenitor cells.     

HBcAg and HBsAg expression in ductular cells in chronic hepatitis B

The possible involvement of bile duct epithelium (BDE) in chronic hepatitis B was examined by immunohistochemical investigation of HBcAg and HBsAg expression in biliary cells in 47 liver biopsies with both viral antigens detectable in hepatocytes. HBsAg- and HBsAg-positive cells were identified in nine and five cases, respectively, in atypical and occasionally in typical ductules in cases of acute excacerbation, chronic active hepatitis and active cirrhosis. Atypical ductules were usually located in areas of periportal fibrosis and in cirrhotic septa. Liver cell plates expressing viral markers and undergoing ductular transformation (positive reaction of hepatocytes to BDE-specific, wide-spectrum keratin) were also observed in acinar zone 1, at the periphery and within parenchymal nodules in a number of cases. The presence of both viral antigens in atypical ductules in cases of advanced chronic liver disease most probably expresses the persistence of the virus in cells deriving from biliary metaplasia of infected hepatocytes. However, the detection of the virus in a few typical ductules is indicative of a direct viral infection. According to these findings, ductular cells seem to serve as a suitable host for HBV, their genotype permitting viral replication and antigen production.     

Immunohistochemical study on bile ductular proliferation in various hepatobiliary diseases

ABSTRACT- Proliferation of the two types of bile ductules, typical and atypical, in the portal and periportal areas was examined in various liver diseases other than cirrhosis to determine any difference in their immunohistochemical properties and presumed histogenesis. While the typical ductules with a well-formed lumen were frequently seen in a large spectrum of diseases, atypical ductules with a poorly defined lumen were encountered much more frequently in prolonged biliary diseases, including primary biliary cirrhosis and primary sclerosing cholangitis, than in nonbiliary hepatic diseases. Immunocytochemically, cytoplasmic keratin was intensively positive in typical ductules, and the degree of its intensity and extent was variable in atypical ductules. Simultaneously, some of the periportal hepatocytes revealed weak staining for keratin. Luminal borders of typical ductules usually revealed an expression of both carcinoembryonic antigen and epithelial membrane antigen, while atypical ductules and periportal hepatocytes lacked epithelial membrane antigen. The atypical ductules, together with the adjoining hepatocytes, appeared on occasion to form anastomosing cords in prolonged biliary diseases. Thus, atypical ductules seem likely to originate from ductular transformation of the periportal hepatocytes and the typical ductules might result from the proliferation of preexisting interlobular bile ducts and ductules.     

Application of keratin immunocytochemistry and sirius red staining in evaluating intrahepatic changes with acute extrahepatic cholestasis due to hepatic duct carcinoma

A series of 10 cases of biliary obstruction due to primary cholangiocarcinoma has been studied with histological and immunocytochemical means. The total duration of cholestasis (as manifested by jaundice) was between 2 and 11 weeks with variable period of preoperative drainage. Liver biopsy specimens taken during surgery for cholangiocarcinoma were investigated for the presence of ductular proliferation and the development of fibrosis, as demonstrated by Sirius Red F3BA collagen staining. The differentiation of epithelial components was evaluated by AEC-immunostaining with chain-specific monoclonal antibodies specifically directed against human keratins type 7, 18 and 19. Keratin 7, normally occurring only in the ductular system, was expressed in hepatocytes at the periphery of the hepatic lobule (zone I) following about 4 weeks' cholestasis, when an increase of ductular profiles in the enlarged portal areas had become manifest. Such keratin 7 positive cells, however, still retained all morphological aspects of hepatocytes. Keratin 19, normally also restricted to the ductular system in liver, is not expressed by zone I hepatocytes even after longer duration (up to 11 weeks) of cholestasis. It is concluded that the increase in ductular profiles during the first week is mainly due do proliferation of pre-existing ductules, while ductular metaplasia occurs in more chronic cholestasis. Development of fibrosis, not always strictly paralleling the multiplication of ductular profiles in sections through a portal tract, represents an early change, and is clearly apparent after 2 weeks of obstruction.     

Lack of evidence that bone marrow cells contribute to cholangiocyte repopulation during experimental cholestatic ductal hyperplasia.

BACKGROUND: Ductopenia is observed in end-stage human cholestatic diseases. The limited capability of cholangiocytes for proliferation is suggested to be the principal reason. Recently, bone marrow cells (BMCs) have been reported to behave as hepatic stem cells; however, their capability to differentiate into cholangiocytes in cholestasis remains unclear. METHODS: Normal mice were lethally irradiated to suppress the proliferation of self-BMCs; thereafter, the BMCs from enhanced green fluorescent protein (EGFP)-transgenic mice were transferred to recipients. Chronic cholestasis was induced by 0.1%alpha-naphtylisothiocyanate (ANIT) feeding. The proliferation of cholangiocytes and oval cells was assessed morphologically and immunohistchemically (cytokeratin-7 (CK-7), A6). Proliferative activity (proliferating cell nuclear antigen (PCNA) protein expression), hepatic growth factor (HGF) receptor (c-Met), stem cell factor receptor (c-kit), Notch2 and Hes1 expression were also evaluated. RESULTS: Marked cholangiocyte proliferation was observed in ANIT-fed mice. However, no EGFP/CK-7 double positive cells were identified in any of the liver specimens after BMCs transfer (Tx). In hepatic parenchyma, there were scattered EGFP-positive cells, although none of them were positive for CK-7. CONCLUSIONS: In spite of the significant ductular proliferations after ANIT feeding, no EGFP-positive cholangiocytes were confirmed by any other means in this chronic cholestasis model. Thus, different from hepatocytes, BMCs Tx seems not to contribute to the differentiation of cholangiocytes. Future studies are feasible to clarify the origin of proliferative cholangiocytes observed in this chronic cholestatic ductular hyperplasia model.     

Epithelial cell adhesion molecule (EpCAM) marks hepatocytes newly derived from stem/progenitor cells in humans

Epithelial cell adhesion molecule (EpCAM) is a surface marker on human hepatic stem/progenitor cells that is reported as absent on mature hepatocytes. However, it has also been noted that in cirrhotic livers of diverse causes, many hepatocytes have EpCAM surface expression; this may represent aberrant EpCAM expression in injured hepatocytes or, as we now hypothesize, persistence of EpCAM in hepatocytes that have recently derived from hepatobiliary progenitors. To evaluate this concept, we investigated patterns of EpCAM expression in hepatobiliary cell compartments of liver biopsy specimens from patients with all stages of chronic hepatitis B and C, studying proliferation, senescence and telomere lengths. We found that EpCAM(+) hepatocytes were rare in early stages of disease, became increasingly prominent in later stages in parallel with the emergence of ductular reactions, and were consistently arrayed around the periphery of cords of keratin 19(+) hepatobiliary cells of the ductular reaction, with which they shared EpCAM expression. Proliferating cell nuclear antigen (proliferation marker) and p21 (senescence marker) were both higher in hepatocytes in cirrhosis than in normal livers, but ductular reaction hepatobiliary cells had the highest proliferation rate, in keeping with being stem/progenitor cell-derived transit amplifying cells. Telomere lengths in EpCAM(+) hepatocytes in cirrhosis were higher than EpCAM(-) hepatocytes (P < 0.046), and relatively shorter than those in the corresponding ductular reaction hepatobiliary cells (P = 0.057). CONCLUSION: These morphologic, topographic, immunophenotypic, and molecular data support the concept that EpCAM(+) hepatocytes in chronic viral hepatitis are recent progeny of the hepatobiliary stem/progenitor cell compartment through intermediates of the transit amplifying, ductular reaction hepatobiliary cells.