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An outbreak of cholangiocarcinoma in a printing company was reported in Japan, and these cases were regarded as an occupational disease (occupational cholangiocarcinoma). This study examined the expression status of programmed death-1 (PD-1) and programmed death-ligand 1 (PD-L1) in occupational cholangiocarcinoma. Immunostaining of PD-1, PD-L1, CD3, CD8, and CD163 was performed using tissue sections of occupational cholangiocarcinoma (n = 10), and the results were compared with those of control cases consisting of intrahepatic (n = 23) and extrahepatic (n = 45) cholangiocarcinoma. Carcinoma cells expressed PD-L1 in all cases of occupational cholangiocarcinoma, whereas the detection of PD-L1 expression in cholangiocarcinoma cells was limited to a low number of cases (less than 10%) in the control subjects. In cases of occupational cholangiocarcinoma, occasional PD-L1 expression was also noted in precancerous/preinvasive lesions such as biliary intraepithelial neoplasia and intraductal papillary neoplasm of the bile duct. Additionally, tumor-associated macrophages and tumor-infiltrating T cells expressed PD-L1 and PD-1, respectively. The number of PD-L1-positive mononuclear cells, PD-1-positive lymphocytes, and CD8-positive lymphocytes infiltrating within the tumor was significantly higher in occupational cholangiocarcinoma compared with that in control cases. These results indicate that immune escape via the PD-1/PD-L1 axis may be occurring in occupational cholangiocarcinoma., (© 2017 Japanese Society of Pathology and John Wiley & Sons Australia, Ltd.)

Intraductal papillary neoplasms of the bile duct are generally thought to arise from neoplastic papillary proliferation of epithelial cells lining the bile duct. We herein report a case with findings that strongly suggested that the biliary cystic tumor might have derived from a peribiliary gland. A 69-year-old female was found to have a cystic lesion with intracystic protrusions at the anterior segment of the right hepatic lobe and underwent hepatic anterior segment resection. Fluoroscopy of the resected specimen injected with contrast medium into the cyst revealed a connection between the cystic lesion and the bile ducts. The cyst was multilocular in appearance. On microscopic examination, the cyst was located within the portal tract of the inferior branch of the anterior segment and connected with the inferior branch of the bile duct. The wall of the hepatic cyst lacked an ovarian-like stroma. The tumor was composed of papillary and glandular components, and the tumor cells were similar to gastric foveolar and pyloric gland epithelia and regarded as adenoma. These tumor cells were positive for MUC 5AC, MUC6, and HIK1083. The tumor was finally diagnosed as an intraductal papillary neoplasm of the bile duct (adenoma, gastric type) arising from a peribiliary gland., (© 2011 The Authors. Pathology International © 2011 Japanese Society of Pathology and Blackwell Publishing Asia Pty Ltd.)

Intrahepatic cholangiocarcinoma (ICC) is reported to develop in non-biliary chronic advanced liver diseases (CALD). Herein, we characterize the pathological features of ICC arising in CALD in comparison with those in non-CALD livers. Of 471 surgically resected cases of ICC in Kanazawa, Japan and Seoul, Korea, 53 were associated with CALD (group A), while the remaining 418 arose in otherwise normal livers (group B). When ICC were classified into bile duct type, bile ductular type, variants, and intraductal papillary neoplasm of the bile duct (IPNB), the whole spectrum of subtypes were found in group A; the majority of ICC belonged to the bile duct type in both groups. In group A, bile ductular type was rather frequent (22.6%) compared with group B (8.4%). IPNB was more frequent in group B (22.5%) than group A (3.8%), and in group B, frequent in Seoul cases (24.8%), but rare in Kanazawa cases (2.3%). Variants of ICC were rare in both groups. These results imply that cholangiocarcinogenesis itself is upregulated in group A in comparison with group B and that the bile ductular type is specifically related to group A. Some unique environmental factors in Seoul may be responsible for the frequent development of IPNB., (© 2011 The Authors. Pathology International © 2011 Japanese Society of Pathology and Blackwell Publishing Asia Pty Ltd.)

Recently, biliary neoplasms resembling intraductal neoplasms of the pancreas have been documented. In this report, a rare case of intraductal tubular neoplasm (ITN) arising in the common bile duct is presented. A polypoid mass, 10 mm in diameter, was found in a 67-year-old woman in the intrapancreatic part of the common bile duct during the follow up to cholecystolithiasis and choledocholithiasis. Endoscopic polypectomy was performed for the lesion. Histology of the lesion revealed tubular neoplasm, composed of an admixture of tubular glands resembling pyloric gland adenoma with minimal atypia (low-grade tubular adenoma), and those resembling intestinal type tubular adenoma (high-grade tubular adenoma). There was no significant formation of papillae or oncocytic cytoplasm. Small foci of carcinoma in situ of the intestinal type were also observed. On immunostaining low-grade tubular adenoma was positive for MUC5AC and MUC6, and negative for MUC2 and cytokeratin (CK) 20, while high-grade tubular adenoma and carcinoma in situ were positive for MUC2 and CK20, and negative for MUC5AC. Although more case studies of ITN in the biliary tracts are required to clarify the tumorigenesis and pathological features, the lesion may be the biliary counterpart to pancreatic ITN.

There are peribiliary glands around the biliary tract, and these glands drain into the bile duct lumen. Interestingly, small amounts of pancreatic exocrine acini are intermingled with these glands. Experimental studies using animals suggest that the biliary tract shows some potential for pancreatic differentiation. It is noteworth that the biliary tract and pancreas have similar pathological features. IgG4-related sclerosing cholangitis and autoimmune pancreatitis are representative inflammatory diseases with similar features. Intraductal papillary neoplasms are found in the biliary tract and also in the pancreas: intraductal papillary neoplasm of the bile duct (IPNB) and intraductal papillary mucinous neoplasm of the pancreas (IPMN). IPNB and IPMN share common histologic and phenotypic features and biological behaviors. Interestingly, mucinous cystic neoplasm (MCN) arises in both the pancreas and the hepatobiliary system. Intraductal tubular neoplasia is found in both the biliary tract and pancreas as well. Intraepithelial neoplasm is found in the biliary tract and pancreas: biliary intraepithelial neoplasm (BilIN) and pancreatic intraepithelial neoplasm (PanIN). BilIN and PanIN are followed by conventional invasive adenocarcinoma, while IPNB and IPMN are followed by tubular adenocarcinoma and mucinous carcinoma in both organs. Further study of the biliary tract's pathophysiology based on its similarity to pancreatic counterparts is warranted.

Polyneuropathy, organomegaly, endocrinopathy, M protein, and skin changes (POEMS) syndrome is a rare multi-system disease. Reported herein is an autopsy case of POEMS syndrome in a subject who developed idiopathic portal hypertension (IPH). The patient was a 38-year-old woman who was initially admitted to the Saiseikai Central Hospital because of polyneuropathy and edema. Diagnosis of POEMS syndrome was established on additional symptoms (splenomegaly and papilloedema) and serum M-protein. Corticosteroid was given for 10 years. The patient was admitted again at the age of 48 years because of gastrointestinal bleeding due to portal hypertension. The patient died of hepatoencephalopathy at 58 years of age. The liver at autopsy demonstrated dense portal fibrosis and obliteration of small portal vein branches, which are characteristic histological findings of IPH. Portal hypertension is a rare symptom in POEMS syndrome. Only three cases of IPH associated with POEMS syndrome (including the present one) have been reported so far. In the previous two reports, liver biopsy failed to determine the cause of portal hypertension. This is the first report on the occurrence of histological findings compatible with IPH in the liver. Although it is not confirmed whether IPH is related to POEMS syndrome, elevated serum cytokines such as vascular endothelial growth factor and coagulation abnormality could have contributed to the development of IPH in the present case.

Recently the authors proposed a new staging and grading system for primary biliary cirrhosis (PBC) that takes into account necroinflammatory activity and histological heterogeneity. Herein is proposed a convenient version of this system. Scores for fibrosis, bile duct loss, and chronic cholestasis were combined for staging: stage 1, total score of 0; stage 2, score 1-3; stage 3, score 4-6; and stage 4, score 7-9. Cholangitis activity (CA) and hepatitis activity (HA) were graded as CA0-3, and HA0-3, respectively. Analysis of interobserver agreement was then conducted. Digital images of 62 needle liver biopsy specimens of PBC were recorded as virtual slides on DVDs that were sent to 28 pathologists, including five located overseas. All participants were able to apply this version in all 62 cases. For staging, kappa was 0.385 (fair agreement) and the concordance rate was 63.9%. For necroinflammatory activity, the kappa and concordance rate were 0.110 (slight agreement) and 36.9% for CA, and 0.197 (slight agreement) and 47% for HA, respectively. In conclusion, this new staging and grading system for PBC seems to be more convenient and practical than those used at present, but more instruction and guidance are recommended for the grading of necroinflammatory activity in practice.

Liesegang rings (LR) are acellular, laminated structures that may be deposited within and around cysts and inflamed or necrotic tissue. Previous reports have shown that the kidney and breast are commonly affected organs of LR. Herein is reported a rare case of biliary deposition of LR presenting as a tumor-like polypoid mass in the liver. A 70-year-old man was found to have a cystic lesion, measuring 3.0 cm in diameter, in the lateral segment of the liver. The lesion was accompanied by a solid mass, 1.8 cm in diameter, within the cystic cavity. Lateral segmentectomy of the liver was performed because clinical examinations could not exclude the possibility of hepatobiliary cystadenoma or cystadenocarcinoma. Pathology of the resected specimen indicated a soft polypoid mass, connecting to the cystically dilated bile duct with elongated stalk-like structures. Histologically the surface of the mass was covered by non-neoplastic biliary-type epithelium. Beneath the epithelium, extensive deposition of numerous ring-like laminated structures, which exhibited an identical appearance to LR, was observed. This is a unique and previously unrecognized lesion involving the occurrence of LR deposition in the hepatobiliary tracts, which further formed a grossly visible mass resembling a hepatic tumor.

Cellular senescence is defined as irreversible cell arrest and could work as a safeguard against tumorigenesis. This mechanism was examined in chronic viral hepatitis-related hepatocarcinogenesis. By using surgical resected or wedge biopsied liver specimens from 87 chronic viral hepatitis patients in whom 35 neoplastic nodules (dysplastic nodules and hepatocellular carcinoma) were complicated, P21 expression and senescence-associated beta galactosidase activity, a marker of senescence, were examined. All of these neoplastic nodules harbored portal tracts within the tumors. Hepatocytes expressing senescence markers and cytokeratin (CK)7-positive bile ductules including hepatic progenitor-like cells were increased in periseptal areas in cirrhosis. Interestingly, these cells appeared to form an anatomical complex that was completely lost in the periportal areas within the neoplastic nodules. In one-third of the neoplastic nodules, CK7-positive small neoplastic hepatocytes resembling hepatic progenitor cells proliferated zonally around the portal tracts. In conclusion, loss of a complex of senescent hepatocytes and ductular cell including hepatic progenitor-like cells in the periportal or periseptal areas may be associated with emergence of neoplastic hepatocytes and their proliferation followed by neoplastic nodules arising in liver cirrhosis. Zonal proliferation of CK7-positive small neoplastic hepatocytes resembling hepatic progenitor cells may develop during early hepatocarcinogenesis.

A case of intraductal papillary-mucinous neoplasm (IPMN) of the pancreas associated with polycystic liver and kidney disease is reported. A 63-year-old man was admitted to hospital with heartburn and upper abdominal pain. CT indicated multiple cysts in the liver and bilateral kidneys, and also showed multiple cystic lesions in the pancreatic head. Pancreatoduodenectomy was performed, and a pathological diagnosis of intraductal papillary-mucinous adenoma of branch duct type of the pancreas was made. He died of renal failure and hepatic failure due to recurrent cholangitis 12 years after the operation. An autopsy confirmed multiple cysts in the liver and kidneys. Multiple hepatic cysts were lined by biliary-type epithelium with no connection to the bile ducts. Peribiliary cysts were observed, and histology of the liver indicated von Meyenburg complexes. The kidneys showed that the cysts originated from all segments of the renal tubule. The autopsy findings suggested that the patient had suffered from autosomal-dominant polycystic kidney disease (ADPKD), but he did not have a family history of ADPKD. This case demonstrates that IPMN of the pancreas can occur as an extrarenal complication in patients with polycystic liver and kidney disease.

Hepatocellular necrosis may be complicated in the clinical course of T-cell lymphoma. Herein are reported two cases of peripheral T-cell lymphoma associated with a rapid increase of serum transaminases in their final course. The final diagnoses were enteropathy-type T-cell lymphoma of the jejunum (patient 1) and hepatosplenic T-cell lymphoma (patient 2). The livers of both patients had non-zonal, well-demarcated coagulation necrosis (approximately 70% in patient 1 and 40% in patient 2) that was infiltrated by lymphoma cells bearing a cytolytic granule content (granzyme B). Hepatocytes in the necrotic areas in patient 2 showed immunoreactivity for human simplex virus type 1. Although the mechanism of the hepatocyte necrosis is unclear, it is suggested that the hepatocyte submassive necrosis may be related to an infection complicated in the final course of lymphoma or by certain direct effects of the tumor cells.

Xanthogranulomatous cholecystitis (XGC) is characterized by the infiltration of numerous foamy macrophages. Bacterial infection is thought to be involved in the pathogenesis of XGC. Using XGC and cultured murine biliary epithelial cells (BEC), the participation of E. coli and the role of the scavenger receptor class A (SCARA), as well as chemokine(C-X-C motif) ligand 16 (CXCL16) and its receptor chemokine(C-X-C motif) receptor 6 (CXCR6), were examined in the pathogenesis of XGC. E. coli components and genes were detected in XGC on immunohistochemistry and polymerase chain reaction (PCR), respectively. SCARA-recognizing E. coli was found in foamy macrophages aggregated in xanthogranulomatous lesions. CXCL16, which functions as a membrane-bound molecule and soluble chemokine to induce adhesion and migration of CXCR6(+) cells, was detected on gallbladder epithelia, and CXCR6(+)/CD8(+) T cells and CXCR6(+)/CD68(+) macrophages were also accumulated. In cultured BEC, CXCL16 mRNA and secreted soluble CXCL16 were constantly detected and upregulated by treatment with E. coli and lipopolysaccharide through Toll-like receptor 4. These suggest that SCARA in macrophages is involved in the phagocytosis of E. coli followed by foamy changes and that bacterial infection causes the upregulation of CXCL16 in gallbladder epithelia, leading to the chemoattraction of macrophages via CXCL16-CXCR6 interaction and formation of the characteristic histology of XGC.

Scirrhous hepatocellular carcinoma (scirrhous HCC) is a rare histological subtype of HCC characterized by marked fibrosis along the sinusoid spaces. Carcinogenetic processes and pathological features at earlier stages of scirrhous HCC remain unclarified. In the present report two cases of minute scirrhous HCC suggesting different carcinogenesis, are described. The first case involved a 54-year-old man with liver cirrhosis related to HCV infection. This patient died of ruptured splenic aneurysm. At autopsy a hepatic tumor measuring 1.8 cm was found, and this tumor had a nodule-in-nodule appearance. Histologically, the inner part of the tumor was well-differentiated HCC with prominent collagen fiber deposition along the sinusoids (scirrhous HCC), whereas the outer part was a high-grade dysplastic nodule. The other patient was a 75-year-old woman who died of hepatic failure due to liver cirrhosis probably related to non-alcoholic steatohepatitis. At autopsy a hepatic tumor (1.2 cm in diameter) was incidentally found in the cirrhotic liver, and was histologically scirrhous HCC without dysplastic nodule elements. Carcinoma cells proliferated along the cirrhotic fibrous septa and replaced regenerative nodules. These two cases suggested that scirrhous HCC could occur in dysplastic nodules (the former case) and also develop de novo in cirrhotic liver (the latter case).

Tumor-stromal interactions are important for the progression of malignant tumors. The purpose of the present study was to examine interactions of cholangiocarcinoma (CC) cells and stromal fibroblasts with respect to stromal-derived factor-1 (SDF-1) and transforming growth factor (TGF)-beta1. Two cell lines of CC (HuCCT-1 and CCKS-1) and WI-38 fibroblast cell line were used for cell culture, and 12 CC tissue specimens for immunohistochemical studies. Invasion of CC cells was increased significantly by the supernatant from fibroblast cultures, but not by the supernatant from fibroblasts cocultured with CC cells. Expression of SDF-1 in cultured fibroblasts was downregulated by TGF-beta1 treatment, and coculture with CC cells and anti-TGF-beta1 neutralizing antibody restored the decreased SDF-1 expression, suggesting that TGF-beta1 secreted from CC cells might have reduced the expression of SDF-1 by fibroblasts and might have reduced the increased invasion of CC cells induced by the supernatant from fibroblasts. Immunohistochemical expression of TGF-beta1 in CC cells was focal or negative and that of SDF-1 was evident in stromal fibroblasts at the invasive front of CC. In conclusion, local mutual influence of TGF-beta1 secreted from carcinoma cells and SDF-1 expressed by stromal fibroblasts may be involved in invasion of CC cells.

The ciliated hepatic foregut cyst (CHFC) is a rare lesion that may arise from remnants of the embryonic foregut. Its wall is lined by pseudostratified ciliated columnar epithelium, and is characterized by the presence of bundles of smooth muscle. Herein is presented two cases of ciliated hepatic cyst without smooth muscle layer. One case was an incidental autopsy finding, and the other was a surgically resected cystic lesion of the liver. Both cysts were <2.0 cm in diameter, and were located subcapusularly in the medial segment (S4) of the liver. The histological appearance of the two cases was identical. They had an epithelial lining of ciliated pseudostratified cells with occasional goblet cells, and lacked a smooth muscle layer, as shown by the actin immunostaining. The lining epithelium contained cells positive for immunohistochemical staining of surfactant apoprotein A, suggesting the embryonic foregut origin of the cysts and differentiation toward bronchiolar structures, rather than ciliated metaplasia of the epithelium of the simple (cholangiogenic) cyst. It is considered that ciliated hepatic cysts of the present case are a rare histological variant of CHFC where the smooth muscle layer is inconspicuous or absent.

Distribution of intra-epithelial lymphocytes along intrahepatic biliary tree (bIEL), and their density and phenotype were examined in normal and diseased livers, particularly in primary biliary cirrhosis (PBC). Immunohistochemically, bIEL were examined in 28 normal livers, 13 cases of chronic viral hepatitis (CVH), 13 cases of PBC, five cases of primary sclerosing cholangitis (PSC), seven cases of extrahepatic biliary obstruction (EBO), and 16 hepatolithiatic livers. In normal livers, bIEL were relatively dense at large and septal bile ducts compared to interlobular ducts. Most of them were positive for CD3 and CD8, while a few were positive for CD4, CD20 and CD57. In CVH, PSC and EBO, neither distribution, phenotype nor density of bIEL differed from normal liver. In hepatolithiasis, numbers of CD8(+)bIEL were increased in stone-containing ducts. In PBC, numbers of CD4(+)CD28(-)bIEL, which are reportedly responsible for target tissue destruction in autoimmune diseases, were markedly increased in damaged interlobular ducts. In conclusion, CD3(+)CD8(+)bIEL may be involved in immune homeostasis of intrahepatic bile ducts in normal livers and in CVH, PSC and EBO. Altered distribution and phenotypes of bIEL in PBC and hepatolithiasis may reflect their participation in biliary lesions. Increased CD4(+)CD28(-)bIEL in damaged bile ducts of PBC may be related to immune-mediated biliary damage.

Biliary lining epithelia of the bile ducts in biliary diseases are known to have intraepithelial atypical/proliferative lesions related to the development of cholangiocarcinoma. The purpose of the present study was to determine the histological criteria for these lesions based on interobserver agreement. Digital images of 30 intraepithelial atypical/proliferative lesions in the stone-containing intrahepatic bile ducts of hepatolithiasis (30 cases) were sent to 10 pathologists. At first, 10 pathologists made a diagnosis (either of reactive/regenerative change, low-grade or high-grade biliary intraepithelial neoplasia (BilIN-1 and BilIN-2), or in situ carcinoma (BilIN-3)) based on their own criteria. The histological criteria for these four lesions were then determined, and the digital images of the same lesions with proposed criteria were re-distributed. Interobserver agreement on these four lesions was slightly improved (kappa = 0.44, first diagnosis; 0.49, second diagnosis) and intraobserver agreement was 'almost perfect' (kappa = 0.82 at both first and second diagnosis). Interobserver agreement between BilIN-1 and BilIN-2 and that between BilIN-2 and BilIN-3 were 'moderate', although the agreement between regenerative/reactive change and BilIN-1 was 'fair'. In this report, we propose histological criteria for reactive/regenerative change, BilIN-1, BilIN-2 and BilIN-3. Improvement of interobserver agreement suggests their applicability in diagnostic and research fields.

A case is reported herein of intrahepatic cholangiocarcinoma (ICC) with multicystic, mucinous appearance and oncocytic change. Because of liver dysfunction, a 73-year-old woman was hospitalized in early October 2003. She was diagnosed as having ICC of the right hepatic lobe with occlusion of the hilar and perihilar bile ducts by imaging examination. Extended right lobectomy was performed but the patient died of liver failure on the next day. In surgically resected specimens, the tumor (3 x 3 cm) was mainly located in the right lobe, and tumors infiltrated along the biliary tree as well as invading into the adjacent hepatic parenchyma. The tumor mass had a sponge or honeycomb appearance. Microscopically, these tumors were composed of multiple microcysts filled by abundant mucin and lined by micropapillary adenocarcinoma cells. Their cytoplasm was acidophilic, appearing as an oncocyte, and carcinoma cells were positive for mitochondrial antigen in addition to biliary cytokeratins. There were no ovarian-like stromas around these cystic tumors, and communication of the biliary lumen with these carcinomatous cysts was not evident, thus different from biliary mucinous cystadenocarcinoma and intraductal papillary neoplasm of the liver. This is the third case of multicystic mucinous ICC and the present case might have been derived from intrahepatic peribiliary glands.

MAGE-A3 antigen is known to be neo-expressed in a large proportion of tumors but not detectable in normal tissues, and could be a target antigen recognized by autologous cytotoxic T lymphocytes. In the present study, the expression of MAGE-A3 at protein and mRNA levels was examined in intrahepatic cholangiocarcinoma (ICC) and its precursor lesions. Carcinomatous and dysplastic biliary cells expressed MAGE-A3 in their cytoplasm diffusely, although there was no MAGE-A3 expression in normal and hyperplastic biliary cells. MAGE-A3 was expressed in one of 10 cases (10%) of low-grade dysplasia, four of 13 (31%) cases of high-grade dysplasia/in situ carcinoma, and 32 of 68 invasive ICC cases (47%), respectively. The MAGE-A3 mRNA expression pattern was similar to that of MAGE-A3 protein. The incidence and intensity of MAGE-A3 expression increased along the progression of biliary neoplasia (P < 0.05). There was no correlation between MAGE-3 expression and histological differentiation or anatomical locations of invasive ICC. MAGE-A3 is a promising target molecule for the specific immunotherapy of ICC.

Primary retroperitoneal cystic tumor is extremely rare, and its histogenesis and biological behavior remain speculative. Two surgical cases of retroperitoneal mucinous cystadenoma (Case 1, an 18-year-old woman; and Case 2, an 85-year-old woman) are reported. The cystadenomas in these cases were mainly lined by a monolayer of columnar or thin flat cells. Case 1 was positive for mucin and epithelial membrane antigen, whereas Case 2 was positive for a mesothelial marker (calretinin). Ciliated epithelium was also interspersed in Case 2. Some parts showed papillary projections, resembling well-differentiated papillary mesothelioma. Within the cyst walls of both cases, ovarian-like stroma that was positive for both estrogen and progesterone receptors was found. Interestingly, focal nodular hyperplasia of the liver was also detected in Case 1. We believe the retroperitoneal mucinous cystadenoma might have arisen from the peritoneum via mucinous epithelial metaplasia with a phenotype of extragenital Mullerian system.

We report a surgical case of liver tumor, 40 x 35 mm in size, with squamous cell carcinoma (SCC) and hepatocellular carcinoma (HCC) components in a 60-year-old Japanese man with steatohepatitis. Most of the SCC component showed typical intercellular bridge and keratinization, while most of the HCC components showed a thick trabecular pattern with mild to moderate nuclear atypia. Both components transit each other without undifferentiated foci; however, a small foci showing glandular structure was intermediated. No cyst formation was found in the liver. The primary site of the squamous cell carcinoma was not detected in general clinical and radiological examination. Immunohistochemical analysis revealed that part of the HCC components neighboring the SCC showed patchy and weak expression of cytokeratin 7. There are several possibilities for the origin of squamous cell carcinoma in this case: marked squamous metaplastic change of cholangiocarcinoma and/or HCC, and carcinoma originating from pleuripotential stem cells. Irregular fatty changes, scattered giant mitochondria and acellular fibrosis with bridging were seen in the liver; however, this patient had no episode of hepatitis-associated viral infection. This is an interesting case of combined hepatocellular and cholangiocarcinoma with marked SCC components arising in a non-cirrhotic fibrotic liver.

Recently, it was shown that osteopontin (OPN) is involved as a chemoattractant cytokine in the recruitment of macrophages and T lymphocytes in the granulomas of diverse etiologies and also plays an important role in the production of autoantibodies and development of autoimmune diseases. Primary biliary cirrhosis (PBC) is characterized by immune-mediated bile duct damage with frequent epithelioid granulomas. In this study, the expression of OPN was immunohistochemically examined in 25 PBC and 52 control livers. Epithelioid cells within granuloma in PBC expressed OPN variably. These cells were also positive for CD68, suggesting their histiocyte/macrophage lineage. In addition, strong expression of OPN was seen in the cytoplasm of mononuclear cells infiltrating around granulomas and also damaged bile ducts in PBC. The number of such positive mononuclear cells and the ratio of OPN-positive cells/total infiltrating cells in portal tracts were higher in PBC than in controls. The majority of these OPN-positive cells were found to be IgG- or IgM-producing plasma cells. These suggest that in PBC, OPN is an important immune molecule in portal tracts, and contributes to the recruitment of mononuclear cells into epithelioid granuloma and also participates in bile duct injury via B-cell differentiation and plasma cell expansion.

Autoimmune hepatitis (AIH) and primary biliary cirrhosis (PBC) are representative autoimmune liver diseases in which hepatocytes and intrahepatic bile ducts, respectively, are selectively damaged by autoimmune mechanisms. Bile duct injury and loss is characteristic of PBC and chronic non-suppurative destructive cholangitis (CNSDC), in particular, is a histological hallmark of PBC. In this report, we present an unusual case of AIH accompanied by CNSDC-like bile duct injury in a 46-year-old woman. The patient's serum aminotransferase level was abnormally high. The serum levels of alkaline phosphatase, gamma-GTP and IgG were also elevated, but the IgM level was within normal limits. The titer of antismooth muscle antibody (SMA) was 1:80, while antinuclear autoantibody (ANA) and the M2 fraction of antimitochondrial antibody (AMA) were both negative. Liver biopsy disclosed CNSDC-like bile duct injuries and severe interface hepatitis and lobular hepatitis with perivenular zonal necrosis were observed. The aggregate score of the International Autoimmune Hepatitis Group corresponded to the category of probable AIH. Immunohistochemically, histocompatibility leukocyte antigen-DR, which is aberrantly expressed in the damaged bile ducts of PBC, was not found in the injured bile ducts of this case. Laboratory data were much improved by treatment with prednisone, but ursodeoxycholic acid was not effective. Although the possibility of an overlapping syndrome of AIH- and AMA-negative PBC could not be excluded, this case was diagnosed as AIH with CNSDC-like bile duct lesions.

MRL/MP mice bearing the lymphoproliferative gene lpr (known as MRL/MP-lpr/lpr or MRL/Ipr mice) are known to spontaneously develop severe autoimmune diseases such as glomerulonephritis, arteritis and arthritis at an early stage of their life. They have also been reported to develop severe sialadenitis, suggesting that this mouse could be a model for Sjögren's syndrome. Primary biliary cirrhosis, an autoimmune disease characterized by chronic non-suppurative destructive cholangitis and the occurrence of antimitochondrial antibodies, is frequently associated with Sjögren's syndrome. In this study, we examined whether cholangitis and/or antimitochondrial antibodies occur in this mouse model, using more than 100 young and old MRL/Ipr mice. We frequently found portal inflammation associated with cholangitis of small intrahepatic bile ducts, especially in older mice. There was also infiltration of inflammatory cells (monocytes) as well as CD4-positive T cells. Epithelioid granuloma and bile-duct loss were also occasionally found. These histological features resemble primary biliary cirrhosis. In addition, antimitochondrial antibodies were shown by immunocytochemistry to be present in the sera of MRL/Ipr mice. There is currently no established animal model for primary biliary cirrhosis. Therefore, further studies on MRL/Ipr mice, with respect to pathogenesis of primary biliary cirrhosis, are warranted.

To investigate the molecular mechanism for enhanced fibrous stroma formation in intrahepatic cholangiocarcinoma (ICC), we surveyed the expression pattern of basement-membrane-type heparan sulfate proteoglycan (HSPG; also known as perlecan) at the core protein and the mRNA level in ICC as well as in other liver neoplasms and reactive hepatic diseases. Immunohistochemistry of paraffin-embedded liver sections with hyaluronidase pretreatment showed that HSPG was present in small amounts in normal liver around the bile ducts and the blood vessels within the portal area. There was no evident expression within the hepatic lobules. Intense immunoexpression of HSPG was seen in the tumor-specific fibro-myxoid stroma of ICC and metastatic liver cancer originating from the colon. However, tumor-specific stroma of hepatocellular carcinomas showed little or no expression of HSPG. At the mRNA level, signals for HSPG were found in tumor cells of cholangiocarcinoma and metastatic colonic carcinomas, and in myofibroblasts in the tumor fibro-myxoid-specific stroma. From immunoprecipitation and reverse transcription-polymerase chain reaction (RT-PCR) analyses, a cultured human intrahepatic cholangiocarcinoma cell line (CCKS1), was found to express high levels of HSPG core protein and mRNA. These findings suggest that biliary and metastatic colon carcinoma cells as well as stromal myofibroblasts have a potential for HSPG production. In order to investigate the growth, invasion and metastatic ability of ICC, further study of the 'self-made' stromal component of ICC may provide a new approach.

The incidence and significance of hepatocellular carcinoma (HCC) in non-alcoholic steatohepatitis (NASH) has not been previously evaluated in detail. We recently experienced a case of NASH with multicentric HCC in a female patient. At the age of 58 years, the patient was diagnosed with non-insulin-dependent diabetes mellitus, treated by insulin therapy. The patient did not drink alcohol. She was negative for all serological markers of hepatitis B and C virus infection. Because of liver dysfunction, a needle biopsy was performed at the age of 62 years, and pathological findings, such as fatty change, Mallory's body, nuclear glycogen and pericellular fibrosis, suggested a diagnosis of NASH. Subsequently, four nodules were detected in the liver by imaging. Liver biopsies were performed from each nodule. One nodule was pathologically diagnosed as a pseudolymphoma, while three other nodules were moderately differentiated HCC (10 years after the diagnosis of non-alcoholic steatohepatitis), well-differentiated HCC (11 years later) and dysplastic nodule (11 years later), suggesting multicentric occurrence of HCC. This case suggests that HCC could be a late complication of NASH.

Primary gall-bladder carcinoma producing granulocyte-colony stimulating factor (GCSF) is extremely rare. Only four cases, histologically investigated, have been reported to date in the English literature. We report a case of a 48-year-old female with primary gall-bladder carcinoma, associating with leukocytosis (15 700/mm3) and a high level of serum GCSF (54.0 pg/mL). The tumor was, histologically, a poorly differentiated adenocarcinoma with marked interspersed neutrophils invading into the primary tumor itself and the right lobe of the liver. Tumor cells distinctly showed positive immunoreaction in the cytoplasm with anti-GCSF antibody, and in the nucleus for anti-p53 antibody. After surgery, the leukocytosis and serum level of GCSF began to decrease. These findings confirmed the present case of GCSF-producing gall-bladder carcinoma, exhibiting leukocytosis. A total of five cases, including our case, reported as a GCSF-producing gall-bladder carcinoma were clinicopathologically reviewed.

Sialyl-Tn, Tn and T antigens are caused by aberrant or incomplete glycosylation of apomucins and are related to the aggressiveness of malignant neoplasms. Using 41 liver samples from patients with cholangiocarcinoma (including four with cirrhosis), 21 with combined hepatocellular-cholangiocellular carcinoma and 17 with hepatocellular carcinoma, the expression of sialyl-Tn, Tn and T antigens were characterized immunohistochemically and the correlation with apomucin profiles was evaluated. The prevalence of sialyl-Tn, Tn and T antigens expression was 89, 95 and 51% in cholangiocarcinoma without cirrhosis; 25, 75, and 0% in cholangiocarcinoma with cirrhosis; 29, 90, and 48% in combined hepatocellular-cholangiocellular carcinoma; and 0, 12 and 6% in hepatocellular carcinoma, respectively. Sialyl-Tn antigen was frequently expressed in cholangiocarcinoma without cirrhosis compared with cholangiocarcinoma with cirrhosis and combined hepatocellular-cholangiocellular carcinoma (P < 0.01). Although sialyl-Tn expression was associated with MUC1, MUC6 and MUC7 expression, the expression sites among them were not identical in the individual cases. These data suggest that the different expressions of sialyl-Tn antigen among cholangiocarcinoma without cirrhosis, cholangiocarcinoma with cirrhosis and combined hepatocellular-cholangiocellular carcinoma may reflect the biological features inherent to these tumors, such as the ability of invasion.

To date, nine apomucins have been characterized and their expression in malignant and premalignant lesions is under evaluation. The purpose of this study was to characterize immunohistochemically the expression of MUC2 (colonic/ intestinal type), MUC5AC (gastric surface type), and MUC6 (pyloric gland type) apomucins in 55 patients with gallbladder carcinoma (10 with in situ carcinoma, 45 with invasive carcinoma), 20 patients with gallbladder dysplasia, and 15 patients with non-dysplastic gallbladder. MUC2 was expressed mainly in 'goblet type' cells. The frequency was increased in non-dysplastic gallbladder (47%), dysplasia (75%), and in situ carcinoma (100%), and decreased in invasive carcinoma (58%). Carcinoma cells expressing MUC2, which were usually distributed at superficial areas, and well-differentiated carcinoma expressed MUC2 more extensively than moderately and poorly differentiated ones. MUC5AC was frequently expressed in gallbladder irrespective of non-dysplastic epithelia, dysplasia and carcinoma. MUC5AC was expressed also in carcinoma cells at deeply invasive sites. MUC6 was expressed frequently in pseudopyloric gland metaplasia as well as dysplasia and carcinoma. In conclusion, non-dysplastic gallbladder has a similar phenotype to gastric pyloric mucosa. Gallbladder carcinoma exhibits both intestinal and gastric surface phenotypes in the early stage of carcinogenesis, with the gastric surface phenotype dominant in invasive carcinoma.

Lipomatous pseudohypertrophy of the pancreas was found at autopsy in a 52-year-old Japanese woman with cirrhosis due to chronic hepatitis B. Clinically, there were no clear symptoms of pancreatic insufficiency during the entire course. Marked atrophy and fat deposition of the pancreas had already been detected by computed tomography (CT) at least 6 years before her death. She died of hepatic failure due to decompensated cirrhosis. Autopsy revealed uniform enlargement of the pancreas due to massive fat replacement (lipomatous pseudohypertrophy): the exocrine glandular elements showed marked atrophy and loss, while the islets of Langerhans were preserved. Although the etiology and pathogenesis of lipomatous pseudohypertrophy is still unclear, this case suggests that this condition is causally related to chronic hepatitis B or other chronic advanced hepatic lesions.

Experimental graft-versus-host disease (GVHD) across minor histocompatibility antigens was developed by injecting spleen and bone marrow cells (9:1) of congenic B10.D2 mice into sublethally irradiated BALB/c mice, and the histologic features of the liver were studied for up to 14 months after transplantation. Both intrahepatic and extrahepatic bile ducts were severely involved in the GVHD process and showed features of non-suppurative cholangitis. Inflammatory cells, mainly lymphocytes, abutted the bile ducts and infiltrated into the duct epithelial layer together with a variety of degenerative changes in the epithelial cells. The peak inflammatory activity occurred about 2-3 weeks after transplantation. Thereafter, the inflammatory cell infiltration around the bile ducts and in the bile duct epithelial layer gradually became reduced in severity, although the infiltration persisted during the entire 14 month observation period. Periductal and duct wall fibroplasia began about 1 week after transplantation and gradually progressed. After 2-3 months post-transplantation, distinct ductal and periductal fibrosis of both intrahepatic and extrahepatic bile ducts was observed. This histologic feature resembled that of primary sclerosing cholangitis (PSC). These results suggest that PSC lesions might develop as a result of chronic cellular immunologic mechanisms in GVHD across minor histocompatibility barriers.

Interferon (IFN)-alpha is regarded as an efficient therapy for chronic hepatitis C, despite the fact that less than 50% of patients receiving IFN-alpha are known to show an initial biochemical response, and several adverse reactions related to this therapy are becoming a serious clinical problem. For a more efficient and safer treatment of IFN-alpha, several pretreatment factors to predict a favorable or unfavorable response to IFN-alpha therapy are now being evaluated, such as hepatitis C virus (HCV)-RNA levels in serum and the genotypes of HCV. Recently, the hepatic iron concentration has been reported to influence the outcome of IFN-alpha therapy for chronic viral hepatitis. In the present study, whether hemosiderin deposition in liver is a histologic predictor of response to IFN-alpha therapy was evaluated, as well as which anatomical location showing the hemosiderin deposition was more closely related to the response to this therapy. Two factors, high titer of HCV-RNA in serum and hemosiderin deposition in portal endothelial cells, were found to be predictable factors of poor response to IFN-alpha therapy, and these two factors were found to be related to each other. Results showed that the hemosiderin deposition in portal endothelial vessels is an easily evaluable histologic finding, and clinicians and histopathologists are encouraged to use this finding when selecting patients with chronic hepatitis C suitable for IFN-alpha therapy.

In situ expression of apoptosis and its related antigens has rarely been evaluated in human liver tumors. Therefore, investigation using in situ nick end-labeling and immunohistochemical methods of the in situ expression of apoptosis, proliferating cells, and apoptosis-related antigens in 7 normal livers, 20 cholangiocarcinomas (CC) and 17 hepatocellular carcinomas (HCC) was done. Apoptotic cells as determined by the nick end-labeling method and proliferating cell nuclear antigen-positive cells were present in all specimens, and the percentage of them was significantly higher in CC than in HCC. Bcl-2 protein was present only in one CC and one HCC, but was occasionally noted in bile ducts in non-cancerous livers. C-myc and Fas antigens were not found in any of the cases. Lewisy antigen was expressed in 8 CC, but was absent in the other cases although bile ducts in non-cancerous livers frequently expressed Lewisy. p53 protein was present in 8 CC, but was absent in the other cases. Serial section observations showed that apoptotic cancer cells were consistently negative for proliferating cell nuclear antigen; bcl-2-positive cells did not show apoptosis; p53-positive cancer cells showed apoptosis. Some Lewisy-positive cancer cells showed apoptosis, while others did not. These data suggest that apoptosis and cell proliferation are involved in CC and HCC, and their degree is more severe in CC than in HCC. p53 protein (stimulative) may regulate apoptosis in some cases, whereas c-myc, Fas and Lewisy are not related to apoptosis in CC and HCC in vivo. Many other factors may regulate apoptosis in CC and HCC in vivo.

A case of chronic hepatitis C at the pre-cirrhotic stage complicated with hepatocellular carcinoma is reported. The patient, a 64 year old female, showed elevated levels of serum alkaline phosphatase and immunoglobulin M. Antimitochondrial antibodies were negative by indirect immunofluorescence. Western blotting using beef heart mitochondria and recombinant polypeptides coding for mitochondrial antigens revealed that the patient's serum was positive only for the E2-subunit of the branched chain ketoacid dehydrogenase complex. In the non-neoplastic liver, chronic non-suppurative cholangitis surrounded by epithelioid granuloma, resembling the granulomatous destructive cholangitis of primary biliary cirrhosis, was found. The damaged bile ducts were immunohistochemically minimally positive or ambiguous for HLA-DR, and their expression of the E2-subunit of the pyruvate dehydrogenase complex E2 (PDC-E2) was diffuse or granular, and not typical of primary biliary cirrhosis. There was no bile duct loss, and orcein-positive copper binding granules reflecting chronic cholestasis were negative in periportal hepatocytes. The overall features in this case were consistent with primary biliary cirrhosis presenting an infrequent profile of antimitochondrial antibodies and atypical expression of HLA-DR and PDC-E2 on biliary epithelial cells, with late superimposition on chronic hepatitis C. However, it is also possible that this is a case of chronic hepatitis C with hepatitis-associated bile duct damage accompanied by granulomatous reaction. Either way, this case raises new diagnostic issues in the differential diagnosis of chronic liver diseases presented with granulomatous cholangitis.

To test the diagnostic utility of pancreatic digestive enzyme immunohistochemistry in liver cancers, the expression of three pancreatic digestive enzymes (trypsinogen, chymotrypsinogen and pancreatic lipase) was investigated in cholangiocarcinoma (CC) (n = 42), hepatocellular carcinoma (HCC) (n = 35), combined HCC-CC (n = 11) and metastatic adenocarcinoma (MA) of the liver (n = 34; 4 gastric cancer, 5 pancreatic cancer and 25 colon cancer). In CC, 15 (36%) expressed one or more of these enzymes, while the remaining 27 (64%) did not express any enzymes. In MA, 13 (38%) expressed one or more of these enzymes, while the remaining 21 (62%) did not express any enzymes. Expression of trypsinogen, chymotrypsinogen and lipase was noted in 15 CC (36%), 11 CC (25%) and 15 CC (36%), respectively, and in 9 MA (26%), 6 MA (18%) and 13 MA (38%), respectively. There was no significant difference in the positive ratio of each enzyme between CC and MA. In positive cases, the enzymes were expressed with a cytoplasmic granular pattern. In MA, there was no significant difference in the positive ratio of the enzymes among the primary sites. In contrast to CC and MA, these enzymes were not expressed in any cases of HCC and combined HCC-CC. These data suggest that pancreatic digestive enzyme immunohistochemistry may be useful for differential diagnosis between HCC and CC or MA as well as between combined HCC-CC and CC or MA, but it is not useful for differential diagnosis between CC and MA. A positive reaction for these enzymes is indicative of CC or MA and is against the diagnosis of HCC or combined HCC-CC, and a negative reaction is noncontributory to the differential diagnosis.

A case of mucinous cholangiocarcinoma is reported. The patient was a 49 year old woman with the complaint of lumbago. Imaging examination disclosed a tumor 3.5 cm in diameter in the right hepatic lobe, which showed low density on computerized tomography scans and low signal intensity on T1-weighted magnetic resonance imaging (MRI) and high intensity on T2-weighted MRI. The hepatic tumor expanded rapidly and multiple pulmonary metastases and peritoneal dissemination developed. The patient died due to respiratory failure 5 months after the initial symptom. An autopsy, a massive tumor (9.5 cm in diameter) containing abundant mucus with several surrounding daughter nodules was found in the right hepatic lobe. Metastases were widespread. Histologically, the tumor was composed mainly of numerous lobulated mucus lakes in which adenocarcinoma cells floated. Admixed signet ring cells were also noted. These gross and histologic features differ from those in other mucin-producing hepatic tumors such as biliary papillomatosis and mucinous cystoadenocarcinoma. Immunohistochemically, the adenocarcinoma cells were strongly positive for carcinoembryonic antigen, Lewis Y, Tn, and T antigens and moderately positive for carbohydrate antigen 19-9, Lewis X, sialyl-Lewis X and sialyl-Tn antigen. Mature MUC1 mucin and core protein of MUC1 mucin were also expressed to varying degrees. The rapidly expanding, widespread metastases and poor prognosis found in the present case may be the clinicopathological features of mucinous cholangiocarcinoma.

The classical nomenclature and categorization of neoplastic and non-neoplastic nodular lesions of the liver are being revised due to the tremendous volume of information recently published on this issue. The diagnostic histopathology of non-neoplastic nodular (tumor-like) lesions of the liver that are recognizable in biopsied, surgically resected and autopsied livers is reviewed using current terminology. Generally, such nodules are infrequent and even rare in routine liver specimens. Non-neoplastic nodules include focal nodular hyperplasia, nodular regenerative hyperplasia, compensatory hyperplasia of the liver, pseudonodule of the liver demonstrable by angiography, partial nodular transformation, focal fatty change, nodular hepatic area shown by modified angiography, cirrhotic large regenerative nodule with variable atypia, anoxic pseudolobular necrosis, intrahepatic bile duct adenoma, biliary and mesenchymal hamartoma, and mesenchymal nodular lesions such as inflammatory pseudotumor and pseudolymphoma, pseudolipoma, peliosis hepatis, solitary necrotic nodule, and so on. Some of these develop preferentially in non-cirrhotic or cirrhotic livers, while others occur with similar prevalence in cirrhotic and non-cirrhotic livers. Some occur multiply or diffusely and others singly. As to the pathogenesis of these nodules, it is speculated that hyperplasia due to disturbed intrahepatic circulation or hormonal imbalance, preneoplastic characteristics, abnormal metabolic disturbance, hamartoma or focal necrobiotic processes, and infection have a role. Knowledge and awareness of these non-neoplastic nodular lesions are necessary for precise diagnosis and differentiation of these nodular lesions from neoplastic hepatic nodules.

Pathologic changes of the spleen in primary biliary cirrhosis (PBC) were studied using autopsy cases. By analysis of the data complied in the Annual Registry of Autopsies of Japan (1977-92), it was found that the splenic weight of 184 cases of PBC (450 +/- 224 g) was significantly heavier than that of 41 control autopsy cases of non-biliary liver cirrhosis from the School of Medicine, Kanazawa University (341 +/- 189 g), and also of 210 control cases filed in the Annual Registry of Autopsies of Japan (1982; 334 +/- 174 g). Among the cases of PBC, the splenic weight was positively correlated with the liver weight. There was no significant difference in the splenic weight between the cases belonging to histologic stages 1-3 and those of stage 4. Fibrosis extending from the splenic trabecula, proliferation and dilatation of the splenic sinus, and congestion in the red pulp, all being compatible with chronic splenic congestion, were similarly observed in the spleen in PBC as well as in other non-biliary cirrhotic cases. The present study indicates that the PBC patients present splenomegaly in the non-cirrhotic histologic stage and splenomegaly persists thereafter. However, the histopathologic changes of the spleen peculiar to PBC were not specified in this autopsy study. Prolonged portal hypertension and other mechanisms, possibly related to immune disarrangement, may be responsible for the prominent splenomegaly in PBC.

A case of pseudolymphoma (reactive lymphoid hyperplasia) of the liver in a 66 year old female is presented. A tumor-like lesion was incidentally discovered in the liver during clinical follow up of diabetes mellitus. The hepatic lesion was resected because malignant lymphoma was suspected after a needle biopsy. Grossly, the lesion was well-defined and measured 1.0 x 1.5 x 1.0 cm. Microscopically, the lesion consisted of hyperplastic lymphoid follicles with distinctive germinal centers and interfollicular areas consisting of mature lymphocytes and plasma cells. An immunohistological study revealed that the lymphoid cells of the lesion were polyclonal in immunophenotypes. These histological and immunohistochemical findings strongly suggested a pseudolymphoma and not hepatic inflammatory pseudotumor. This case was diagnosed as pseudolymphoma of liver. Only a few cases of hepatic pseudolymphoma have so far been reported in the English literature.