Unusual hepatocellular lesions in primary biliary cirrhosis resembling but unrelated to hepatocellular neoplasms

Summary Structural, cellular and nuclear abnormalities of hepatocytes are a histological hallmark of well-differentiated, small hepatocellular carcinoma (HCC) or its borderline lesion. This study revealed that several hepatocellular abnormalities found in these hepatocellular neoplasms were also found in non-cirrhotic stages of primary biliary cirrhosis (PBC) in which HCC is unlikely to develop. These changes are small cell changes, consisting of the appearance of small hepatocytes arranged in thin trabecular or compact patterns with increased cellularity and basophilic cytoplasm. This was found in 36%, 71% and 100% in specimens of stages 1, 2 and 3, respectively. Large cell changes occurred and consisted of large hepatocytes with large nuclei and prominent nucleoli, found in 27%, 47% and 22% of the stages, respectively. Finally, liver cell rosettes were seen, showing variable acinar formation and present in 0%, 41% and 33% of the stages, respectively. These lesions were identified microscopically as irregularly shaped areas or vague nodules of hepatocytes without a fibrous rim, in the hepatic lobules. They showed an expansive growth or shaggy border against the surrounding hepatic parenchyma. Follow-up studies, including autopsies, failed to show development of HCC or its borderline lesion in PBC cases. Pathologists must make a diagnosis of HCC and its borderline lesion bearing in mind the occurrence of such unusual hepatocellular lesions probably of a reactive nature.     

Pathologic features of small hepatocellular carcinoma

Twenty-one nodules of small hepatocellular carcinoma (HCC) were examined. Histologically, the nodules often presented formation of plump trabeculae, marked nuclear atypism, or aggressive growth comprising capsular invasion, vascular invasion, and replacement of adjacent pseudolobules. Aside from these characteristic findings of HCC, it was important to reveal the following features for the diagnosis of well differentiated type of small HCC: variable thickening or distortion of trabecular structure in association with nuclear crowding, acinar formation, selective cytoplasmic accumulation of Mallory bodies, nuclear abnormalities consisting of thickening of nucleolus, hepatic cords in close contact with bile ducts or blood vessels, and hepatocytes growing in a fibrous environment. During the invasive growth, the tumor cells may well be subtly blended with benign hepatocytes, giving rise to a pattern of "mixed cellularity". It is also emphasized that connective tissue septa of pseudolobules could be a route of rapid tumor spreading.     

The way we teach general pathology in Oslo, Norway

Hepatocellular carcinoma was induced in rats by administering aflatoxin B₁ (AFB₁) for 6 weeks. Malignant tumours were preceded by foci and nodules of altered hepatocytes of three histological types, composed of basophilic, eosinophilic, and vacuolated cells. In addition, there were areas of altered hepatocytes that were considered as hyperplastic. Lectins were used as histochemical markers to compare the expression of membrane glycoproteins in hepatocellular carcinomas and hepatic nodules with non-nodular or control hepatocytes. There were marked changes in the lectin-binding patterns of the hepatocellular carcinoma cells and the eosinophilic nodules. The lectin-binding patterns of basophilic nodules, vacuolated nodules, and hyperplastic areas were similar to non-nodular or untreated hepatocytes. The similarity in the lectin-binding changes of the eosinophilic nodules and hepatocellular carcinomas suggests that the eosinophilic nodules may be an early stage in the development of carcinoma.     

PATHOLOGIC FEATURES OF SMALL HEPATOCELLULAR CARCINOMA

Twenty-one nodules of small hepatocellular carcinoma (HOC) were examined. Hlstologically, the nodules often presented formation of plump trabeculae, marked nuclear atypism, or aggressive growth comprising capsular invasion, vascular invasion, and replacement of adjacent pseudolobules. Aside from these characteristic findings of HCC, it was important to reveal the following features for the diagnosis of well differentiated type of small HCC: (1) variable thickening or distortion of trabecular structure in association with nuclear crowding, (2) acinar formation, (3) selective cytoplasmic accumulation of Mallory bodies, (4) nuclear abnormalities consisting of thickening of nuclear membrane, irregularities in shape, and enlargement of nucleus or nucleolus, (5) hepatic cords in close contact with bile ducts or blood vessels, and (6) hepatocytes growing in a fibrous environment. During the invasive growth, the tumor cells may well be subtly blended with benign hepatocytes, giving rise to a pattern of "mixed cellularity". It is also emphasized that connective tissue septa of pseudolobules could be a route of rapid tumor spreading.     

大鼠肝癌发生过程中胰岛素样生长因子-Ⅱ的表达及意义

目的:研究胰岛素样生长因子-Ⅱ(IGF-Ⅱ)在肝癌发生过程中的表达及意义.方法:采用二乙基亚硝胺(DEN)诱癌建立大鼠肝癌模型.应用放射免疫分析检测诱癌过程中血清IGF-Ⅱ浓度,应用免疫组织化学方法观察诱癌过程中大鼠肝组织IGF-Ⅱ的表达情况.结果:在大鼠肝癌的发生发展过程中,从肝细胞损伤期、增生-硬化期至癌变期3个阶段大鼠血清IGF-Ⅱ水平呈"高到低到高"的变化趋势;肝细胞损伤期嗜酸性变细胞中见IGF-Ⅱ阳性表达,癌变期癌灶中表达低于癌周增生灶、增生结节和非典型增生结节,而增生-硬化期阳性表达较少.结论:大鼠血清和肝中IGF-Ⅱ的高表达是肝癌发生发展过程中的早期及晚期事件,提示IGF-Ⅱ可能与肝细胞持续增殖及恶性转化有关.     

Analysis of hyperplastic foci in livers with hepatocellular carcinomas by flow cytometry and AgNOR staining

The phase S ratlo in cell cycles were analyzed in livers with hyperplastic foci (HPF) and in livers without HPF by nuclear DNA determinatlons using flow cytometry, and by stalning wlth argyrophilic protelns of the nucieolar organlzer reglon (AgNOR). Flow cytometric analysis was done on 50 fresh frozen speclmens of livers resected from 50 patients wlth hepatocellular carcinoma (HCC). Paraffin sections from the same patients were analyzed uslng AgNOR staining. There were 25 cases each wlth and without HPF. We examined the stage of fibrosis and the grade of inflammatory activity according to the modlfied Scheuer and Desmet scale. The incidence of HCC recurrence among these patlents was also studied. The average phase S ratio of the livers of the patients with HPF was 6.5+3.2%, and that of the livers of the patients without HPF was 4.0±2.5%. The ratio differed slgnificantly between the two groups (P<0.01). The average AgNOR score for HPF lesions of the HPF-positlve cases was 1.6020.34, that for non-HPF lesions In the HPF-positive cases was 1.2920.12, and that for the HPF-negative cases was 1.1920.14. Significant differences were found between the average AgNOR scores for HPF lesions of the HPF-posltive cases and the non-HPF lesions of the HPF-posltive cases (P<0.0l), as well as between the non-HPF lesions in the HPF-positive cases and the HPF-negatlve cases (P-cO.05). Severe fibrosis (stage 3) and cirrhosls (stage 4) were found In 76% of HPF-positive cases and 48% of HPF-negatlve cases. The llvers of HPF-posltlve patlents were slgnificantiy more cirrhotic than those of HPF-negative patients (R0.05). The association between HPF and the Inflammatory grade was not slgnlficant (Ao.05). The incldence of HCC recurrence among HPF-positive cases was significantly higher than that among the HPF-negative cases (P<0.05). The average phase S ratio of the recurrent HPF-positive patients was 7.48+3.48%, slgnlficantly higher than that of HPF negative cases (5.57*3.06%, Pc0.05). Hyperplastic foci of the llver was shown to be a hlghly prollferatlve lesion. The proliferative activity of the non-HPF lesions in the HPF-positive patients was also higher than that of the HPF-negative patients. Hyperplastic focl tended to be present in cirrhotic livers, but it was not associated wlth the grade of inflammatory activlty of the Iiver. Hyperplastic focl may represent an important predictor of recurrence after hepatic resection.     

人肝癌与癌旁肝AFP阳性细胞形态的图象分析及增殖活性的研究

采用免疫组化、双标免疫组化及图象分析技术，研究了２７例人体肝细胞癌与癌旁肝甲胎蛋白（ＡＦＰ）阳性细胞的量化形态特征及以其增殖细胞核抗原（ＰＣＮＡ）标记率显示的增殖活性。主要结果显示：（１）ＡＦＰ阳性与阴性癌及癌旁近癌肝细胞间难以用量化形态标准区分（Ｐ＞０．０５）；核面积与核浆比提示癌旁ＡＦＰ阳性肝细胞可能是尚无表型转化的癌前细胞。（２）ＡＦＰ阳性癌细胞ＰＣＮＡ的平均标记率（１５．７０％±６．２５％）显著低于肝癌的平均标记率（Ｐ＜０．０１），表明大多数ＡＦＰ阳性癌细胞处于非增殖状态。（３）癌旁近癌肝可见ＡＦＰ与ＰＣＮＡ双标阳性肝细胞。     

Morphometric study of hepatocytes containing hepatitis B surface antigen

The development of hepatocellular carcinoma (HCC) is probably related to infection with hepatitis B virus (HBV). Hepatocytes in livers of patients with HCC have been reported to show putative preneoplastic changes such as hyperplasia, dysplasia, or adenomatous regeneration. To determine quantitatively whether these morphologic changes are associated with HBV-infected cells, the authors performed morphometry of hepatitis B surface antigen (HBsAg)-positive hepatocytes in the nontumorous portion of 10 livers with HCC and in 10 livers without HCC. The diameter of nuclei and cytoplasm of HBsAg-positive hepatocytes was measured after demonstration of HBsAg by the peroxidase-antiperoxidase method. As controls, HBsAg-negative hepatocytes in the same liver sections were measured as well as hepatocytes of 20 age-matched HBsAg-negative patients with normal liver or alcoholic cirrhosis. HBsAg-positive hepatocytes exhibited significantly larger nuclei and a higher nucleocytoplasmic ratio than control hepatocytes. In addition, HBsAg-positive cells were often arranged in foci that consisted of two cell populations: hypertrophic (enlarged nuclei and nucleocytoplasmic ratio) and hyperplastic (two-cell-thick plates of small cells with a high nucleocytoplasmic ratio). While precancerous cells have been difficult to identify, these morphologic changes are frequently associated with the development of malignant neoplasia.     

长春花碱和细胞松驰素D对肝癌细胞粘弹性的影响

用微管吸吮技术测定了正常肝细胞和肝实质细胞癌细胞的粘弹性，以三参数标准线性固体模型拟实验结果，进一步研究了两种细胞骨架干扰剂处理后肝细胞和肝癌细胞粘弹性系数的变化。     

Tumorigenic role of YAP in hepatocellular carcinogenesis is involved in SHP2 whose function is different in vitro and in vivo

Yes-associated protein (YAP) is a nuclear effector of the cell-density sensing Hippo pathway and interacts with Src homology phosphotyrosine phosphatase 2 (SHP2), which controls cell proliferation and survival. The tumor promoting/suppressing activities of YAP and SHP2 during liver tumorigenesis remain controversial. This study aimed to investigate the tumorigenic roles of YAP and SHP2 in hepatocellular carcinogenesis. Cell density associated subcellular distributions of YAP and SHP2 in normal human hepatocytes (THLE-2) and hepatocellular carcinoma (HCC) cells (SK-Hep1, SNU-182) were investigated by Western blotting and cell block immunohistochemistry. The effects of YAP knockdown on proliferation, migration and invasion were studied using YAP-specific siRNAs. The prognostic significance of YAP and SHP2 expressions was investigated immunohistochemically using a tissue microarray (TMA) from 50 HCC cases. High-cell density decreased the nuclear expression of YAP and SHP2 in normal hepatocytes as compared with low-cell density. However, in HCC cells, nuclear YAP and SHP2 were observed regardless of cell density. Nuclear YAP influenced SHP2 expression and cell proliferation. In particular, YAP knockdown impacted nuclear levels of SHP2 protein in SK-Hep1 cells. In HCC tissues, nuclear YAP expression was elevated and cytoplasmic SHP2 expression was diminished as compared with adjacent non-tumor tissues. Notably, these expressions were found to be significantly associated with poor recurrence-free and overall survival rate in patients with HCC. Consequently, the tumor promoting role of YAP is involved in SHP2 which functions as a tumor promoter in vitro but as a tumor suppressor in vivo. YAP and SHP2 can be unfavorable prognostic markers in HCC.     

Microvessel density and angiogenesis in primary hepatic malignancies: Differential expression of CD31 and VEGFR-2 in hepatocellular carcinoma and intrahepatic cholangiocarcinoma

Background

Microvessel density is an indicator of tumor-driven neoangiogenesis. Hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma (ICC) have distinct vascular patterns, which are also reflected in their imaging characteristics. Since a significant proportion of HCC are treated without biopsy confirmation, it is essential to discriminate HCC and ICC radiologically. The aim of our study was therefore to compare microvessel density and expression of VEGFR-2 in HCC and ICC, since these data may ultimately help us to better understand their imaging characteristics. Whereas CD31 documents vessel density, VEGFR-2 expression is an indicator of tumor-related neoangiogenesis.

磷脂酰肌醇蛋白聚糖-3的表达在肝细胞癌诊断中的意义

目的 探讨磷脂酰肌醇蛋白聚糖-3( GPC3)免疫组织化学染色在肝细胞癌病理诊断的应用价值。方法 制作14个含731例肝肿瘤和肿瘤旁肝组织芯片,其中肝细胞癌357例、胆管癌26例、肝细胞癌癌旁肝组织包括肝硬化171例、血管瘤旁肝组织93例、肝转移癌84例。全部病例采用免疫组织化学检测GPC3（1G12克隆）蛋白表达,实验设阳性对照。结果 72.0％的肝细胞癌(257/357)GPC3呈阳性反应,其余374例非肝细胞癌病例均为阴性,包括胆管癌、肝转移癌、肝细胞癌癌旁肝组织（包括肝硬化和血管瘤旁肝组织）。GPC3阳性率在不同的肝细胞癌组织学分级的差异有统计学意义(P＜0.01),阳性率高低排列为Ⅲ级(77.1％,64/83)、Ⅱ级(73.3％,187/255)、Ⅰ级6/12和Ⅳ级(0)为阴性。结论 GPC3免疫组织化学检测是肝细胞癌诊断的良好指标,敏感性达72.0％;也是区别瘤旁肝组织和肝转移癌的鉴别诊断指标,其特异性达100％。     

Greater age and hepatocellular aging are independent risk factors for hepatocellular carcinoma arising from non-B non-C non-alcoholic chronic liver disease

We previously reported that hepatocellular aging can be assessed by measuring the nuclear size of hepatocytes. We attempted to elucidate whether this method is useful to identify the high risk group of hepatocellular carcinoma (HCC) in the patients with non-B non-C non-alcoholic liver injury. Fourteen patients with HCC and 78 without HCC, both of whom presented with non-B non-C non-alcoholic chronic liver injury and underwent liver biopsy, were selected. Twelve histologically normal liver tissues were selected as controls. The relative nuclear size (RNS) was calculated as the average nuclear size of the hepatocytes divided by that of lymphocytes. Multiple clinicopathological parameters were studied. The RNS values of normal livers ranged from 1.32 to 2.10, showing a gradual increase in an age-dependent manner. The RNS values of the injured livers without HCC increased after middle age. Univariate analysis identified greater age, existence of diabetes and RNS, as significantly positive contributors and ALT value and the degree of steatosis as negative contributors for the occurrence of HCC. Only age and RNS retained significance in multivariate analysis. All of the HCC patients were older than 50 and showed RNS values higher than 2.00. Therefore, such patients are classified as a high risk group of HCC.     

Growth patterns and interstitial invasion of small hepatocellular carcinoma

Twenty-five caw of small hepatocellular carcinoma (HCC; dhmeter ≥30mm) were evaluated for overall morphologic features and growth patterns. The tumors often showed a uelldtfferentlated, normotrabecular histologic pattern and insidious interstitial invasion, which resembled benign hepetocytes scattered in connective tissues. As the tumor grew, B less-differentiated tumor area became predominant. Portal tracts Included in small HCC nodules were quantitatively assessed, revealing that they progressively reduced in number with tumor growth. The tumor margin was often reported to be unclear. The present results indicate that the histologk grade of tumor differentiation, capsular formation, existence of liver cirrhosis and patterns of interstitial invaslon are important factors for determining the nature of the margin. The score of argyrophilic nuclear organizer regions (AgNOR) was examined in 5 cases showing typical interstitial imaslon with the insidious type. In each case, the AgNOR score of the invading tumor cells was lower than that of turnor cells within the HCC nodules, but higher than benign hepatocytes in cirrhotic parenchyma. It clarified that the growth activity of well-differentiated HCC was rather suppressed upon their interstitial invasion.     

Hepatocellular carcinoma presenting well differentiated, normotrabecular patterns in peripheral or metastatic loci. Analysis of 103 resected cases

In a series of 103 surgically resected hepatocellular carcinomas (HCCs), the tumor boundaries were rather clearly defined in 85 cases (82.5%). Twelve tumors had an ill-defined margin because adjacent pseudolobules were replaced by HCC with a well differentiated growth pattern (group A1). This peripheral pattern was unrelated to the predominant histologic type of the major tumor nodules. Despite its close resemblance to normal hepatic parenchyma, we were able to determine the area of the normotrabecular HCC on the basis of, differences in stain-ability, variable nuclear crowding and frequent microacinar formation. Moreover, scattered occurrence of nuclear atypia and midtrabecular conversion were also observed within the normotrabecular HCC. In an additional 6 cases, a band-like zone of normotrabecular HCC was evident peripherally around the primary lesion (group A2). Fourteen other cases (13.6%) had intrahepatic metastatic lesions formed either within the portal vein or in pseudolobules. Of these, 5 cases showed intrapseudolobular metastases with a normotrabecular pattern (group B). For precise evaluation of surgical HCC specimens, it seems to be particularly important to realize that HCCs sometimes have a highly differentiated appearance in a locus of peripheral invasion as well as intrahepatic metastasis.     

alpha 1-Antitrypsin deficiency and hepatocellular carcinoma. Association with cirrhosis, copper storage, and Mallory bodies

A pathogenetic relationship between hepatic manifestations of alpha 1-antitrypsin deficiency (AATD) and hepatocellular carcinoma has previously been suggested but not established. We encountered three patients with hepatocellular carcinoma (HCC) and nonneoplastic hepatocytes containing globules positive for alpha 1-antitrypsin by immunoperoxidase staining or immunofluorescence microscopy. The AATD was severe in one patient, mild in the second, and not evaluated in the third; any pathogenetic relationship between the deficiency and HCC must therefore remain speculative. Nonneoplastic hepatocytes of all three patients demonstrated copper storage and, in two patients who were not alcoholic, Mallory bodies. Simultaneous presence of copper storage and Mallory bodies has not been reported in liver disease associated with AATD. The pathogenetic mechanisms of these findings are unknown.     

Hepatocellular Carcinoma Presenting Well Differentiated,Normotrabecular Patterns in Peripheral or Metastatic Loci

In a series of 103 surgically resected hepatocellular carcinomas (HCCs), the tumor boundaries were rather clearly defined in 85 cases (82.5%). Twelve tumors had an ill-defined margin because adjacent pseudolobules were replaced by HCC with a well differentiated growth pattern (group Al). This peripheral pattern was unrelated to the predominant histologic type of the major tumor nodules. Despite its close resemblance to normal hepatic parenchyma, we were able to determine the area of the normotrabecular HCC on the basis of, differences in stain-ability, variable nuclear crowding and frequent microacinar formation. Moreover, scattered occurrence of nuclear atypia and midtrabecular conversion were also observed within the normotrabecular HCC. In an additional 6 cases, a band like zone of normotrabecular HCC was evident peripherally around the primary lesion (group A2). Fourteen other cases (13.6%) had intrahepatic metastatic lesions formed either within the portal vein or in pseudolobules. Of these, 5 cases showed intrapseudo-lobular metastases with a normotrabecular pattern (group B). For precise evaluation of surgical HCC specimens, it seems to be particularly important to realize that HCCs sometimes have a highly differentiated appearance in a locus of peripheral invasion as well as intrahepatic metastasis. Acta Pathol Jpn 40: 887–893, 1990.     

Nucleophosmin/B23在肝细胞癌组织中的高表达

目的探讨Nucleophosmin／B23（B23）在肝细胞癌（HCC）组织中的表达及其临床病理意义。方法采用重组蛋白表达和杂交瘤细胞技术制备B23重组蛋白和鼠抗B23单克隆抗体。收集103例HCC组织、12例肝局灶性结节性增生和17例肝血管瘤旁肝组织的临床病理档案资料,10例HCC及癌旁肝新鲜组织,采用免疫组织化学（ABC）方法、逆转录聚合酶链反应（RT-PCR）及免疫印迹等技术检测B23在这些组织中的表达,并与增殖细胞核抗原（PCNA）的表达相比较。应用统计学方法对结果进行分析。结果RT-PCR及免疫印迹结果显示：在mRNA和蛋白质水平,B23在HCC组织中的表达明显高于对应的癌旁肝组织。免疫组织化学结果显示：B23在4组（HCC组、癌旁肝组织组、肝局灶性结节性增生组、血管瘤旁肝组织组）中的表达差异有统计学意义（P〈0．001）,其中B23在HCC组织中的表达显著高于其他3组（P〈0．01）,PCNA在HCC组织中的表达也显著高于其他3组（P〈0．01）,相关性分析表明B23与PCNA在4组中表达强度及其差异具有相关性（r=0．4767,P〈0．01）。B23在HCC组织中的表达强度与患者血清AFP水平、肿瘤病理分级及是否伴有肝硬化之间的关系有统计学意义（P〈0．05）。结论B23在HCC中呈高表达,且显著高于非癌肝组织;B23可作为HCC细胞增生程度的潜在标记,并在临床病理上具有潜在的应用意义。     

Histopathological and morphometric analysis of atypical adenomatous hyperplasia of human cirrhotic livers

Atypical adenomatous hyperplasia (AAH) is a hyperplastic parenchymal nodular change in the cirrhotic liver, in which overt hepatocellular carcinoma (HCC) occasionally arises. AAH is defined as a sizable hepatocellular nodule with a variable degree of hepatocellular atypia not regarded as HCC, and is different from ordinary adenomatous hyperplasia in which hepatocellular atypia is absent. In the present study, we attempted to evaluate carcinogenetic processes and to find histological variables which indicate malignant transformation in AAH, using 49 surgically resected or autopsied nodules. AAH frequently showed morphological heterogeneity. Atypical lesions within AAHs were divisible into the following three categories from overall histopathological appearances: malignant (A), equivocal (B), or non-malignant (C) lesions. Analysis of combination of these three lesions, which were frequently intermixed in a given AAH, suggested that B lesions appear subsequent to C lesions, and A lesions finally appear in AAH nodules. Among the 14 histological variables, enlargement, hyperchromasia and irregular contour of nuclei were found to correlate well with A lesions. Increased nuclear density, iron resistance, reduction of reticulin fibres, clear cell change, sinusoidal dilatation and presence of abnormal arteries were suggestive of A or B lesions. Nuclear deviation toward the sinusoids, acinar and compact arrangements, fatty change and Mallory's hyaline alone were not useful indicators of A or B lesions. These results indicate that AAH is a preneoplastic or borderline lesion in which overt HCC is likely to evolve through several steps. Although a needle liver biopsy is a useful tool for diagnosis of benign, equivocal and malignant hepatocellular nodular lesions, the needle biopsy specimen should be carefully evaluated by considering the morphological heterogeneity of the AAH and a variable combination of 14 histological variables.     

肝癌细胞侵袭行为与其流变特性相关

以原代肝细胞、肝细胞癌（Ｈｅｐａｔｏｃｅｌｌｕｌａｒ ｃａｒｃｉｎｏｍａ,ＨＣＣ）细胞及对人工基底膜Ｍａｔｒｉｇｅｌ具有侵袭能力的ＨＣＣ细胞（ＨＣＣ－Ｉｎｖ细胞）为研究对象,研究了三种细胞的粘弹特性、与胶原蛋白Ⅰ的粘附力学特性及与肝血窦内皮细胞（Ｌｉｖｅｒ ｓｉｏｎｓｏｉｄａｌ ｅｎｄｏｔｈｅｌｉａｌ ｃｅｌｌ,ＬＥＣ）的粘附力学特性。结果发现,ＨＣＣ细胞的上述流变特性较之于肝细胞发生明显的变化,