Nobuaki Okano

Cytokine profile in the liver of primary biliary cirrhosis.

We characterized the cytokine profile in the liver of patients with primary biliary cirrhosis (PBC). Total RNA was extracted from the biopsy specimens of 9 patients with early-stage PBC, 10 with chronic hepatitis C (CHC), and 4 normal controls. cDNA was prepared and amplified with a polymerase chain reaction using primers for interferon (IFN)-γ and interleukin (IL)-2, -4, -5, -6, -10, -12 (p40), and -15. Cytokines such as IFN-γ and IL-5, -6, -10, -12, and -15 were expressed in most cases of PBC. Expression rates of IL-5 and IL-6 were higher than in CHC and controls. The higher expression rate of IL-5 in PBC was associated with eosinophil infiltration. IL-2 and IL-4 were rarely detected. Semiquantitative analysis revealed that the expression of IFN-γ and IL-10 was reversed in PBC and CHC: high IFN-γ and low IL-10 in PBC and high IL-10 and low IFN-γ in CHC. These results suggest that cytokine expression is skewed in PBC and both Th1 and Th2 cytokines may play a role in the pathogenesis.

Stromal Cell–Derived Factor-1 from Biliary Epithelial Cells Recruits CXCR4-Positive Cells: Implications for Inflammatory Liver Diseases

Although stromal cell–derived factor-1 (SDF-1) plays an important role in hematopoiesis in the fetal liver, the role after birth remains to be clarified. We investigated the role of SDF-1 and its receptor, CXCR4, in 75 patients; this included controls and patients with viral hepatitis, liver cirrhosis, primary biliary cirrhosis, primary sclerosing cholangitis, and autoimmune hepatitis. Interestingly, SDF-1 appeared up-regulated in biliary epithelial cells (BEC) of inflammatory liver disease. Furthermore, in inflammatory liver diseases, SDF-1 was expressed by BEC of interlobular and septal bile ducts and by proliferated bile ductules. The message expression of SDF-1 in BEC was confirmed at a single-cell level by RT-PCR and laser capture microdissection. The plasma levels of SDF-1 were significantly higher in patients with liver diseases than in normal controls. Flow cytometric analysis of the surface expression of CXCR4 showed that most liver-infiltrating lymphocytes express CXCR4 and the intensity was up-regulated more significantly in liver-infiltrating lymphocytes than in peripheral blood lymphocytes. These results suggest that increased SDF-1 production by BEC may play an important role in the recruitment of CXCR4-positive inflammatory cells into the diseased livers. These data are significant because modulation of the SDF-1/CXCR4 interaction has therapeutic implications for inflammatory liver diseases.

Clinicopathological features of acute-onset autoimmune hepatitis

The clinicopathological features of nine acute-onset autoimmune hepatitis (AIH) patients were compared with those of 29 classical AIH patients. The clinical features of acute-onset AIH showed significantly higher serum ALT levels, lower serum IgG levels and AIH score than those of classical AIH, although the type of auto-antibodies, age and gender were not different between the two groups. Pathological features showed that the stages of acute-onset AIH varied from stage 1 to stage 4 and were less advanced compared with those of classical AIH. One patient showed submassive hepatic necrosis. Both centrilobular necrosis and interface hepatitis were observed in 7 and 8 of 9, respectively. Three stage 1 patients with centrilobular necrosis and one patient with submassive hepatic necrosis were suggestive of acute presentation, while patients with stages 2 and 4 fibrosis were suggestive of acute exacerbation of chronic disease. An immunohistochemical study demonstrated that CD8 T cells were predominant at both interface hepatitis and centrilobular necrosis, while CD79alpha-positive B lineage cells were predominant at interface hepatitis. These results suggest that acute-onset AIH includes both acute presentation and acute exacerbation of chronic disease and that centrilobular necrosis might be a prevailing pathological feature.

Prediction of the ablated area by the spread of microbubbles during radiofrequency ablation of hepatocellular carcinoma

Abstract: Background/Aim: Radiofrequency ablation (RFA) is effective for the treatment of hepatocellular carcinoma (HCC). To prevent the ablation of adjacent organs and vessels, the spread of microbubbles generated by heating during RFA was observed by ultrasonography (US) and used to predict the ablated area; however, several reports documented that discrepancies existed between the spread of microbubbles and the ablated area.

Pachydermoperiostosis associated with juvenile polyps of the stomach and gastric adenocarcinoma

Pachydermoperiostosis (PDP) is a rare syndrome, and the presence of digital clubbing, radiographic periostosis, and coarse facial features are the main diagnostic criteria. Here, we report patient with the primary form of PDP in whom juvenile polyps and gastric cancer developed within 9 years of follow-up. A 27-year-old Japanese man, diagnosed as having the primary form of PDP at 14 years of age, was referred to our department for assessment of chronic anemia. On upper gastrointestinal endoscopic examination, multiple polypoid lesions with a huge polyp were found in the stomach, and biopsy findings indicated juvenile polyps, although no polypoid lesion had been present at the age of 18 years. Bleeding from these polyps was suspected, and endoscopic mucosal resection of the polypoid lesions was performed. Histology of the huge polyp showed hamartoma, adenoma, and adenocarcinoma in part. This is the first case report of the primary form of PDP associated with gastric cancer. In this patient, juvenile polyps and gastric cancer developed within 9 years of follow-up, indicating that the primary form of PDP may be a high risk factor for gastric cancer, and that periodical follow-up with upper gastrointestinal endoscopy is important.

Expression of Perforin and Fas Ligand mRNA in the Liver of Viral Hepatitis

Cytotoxic T lymphocytes (CTLs) play an important role in the pathogenesis of viral hepatitis. We studied the expression of mRNAs of perforin and Fas ligand (Fas-L) in biopsy specimens from chronic hepatitis B (CHB) (15 cases) and hepatitis C (CHC) patients (13 cases). Both perforin and Fas-L mRNAs were detected in all cases of both CHB and CHC. No messages were detected in the control livers from two cases of fatty liver, a case of Gilberts syndrome, and a case of Dubin–Johnson syndrome. Semiquantitative analysis revealed a positive correlation between the intensity of perforin and Fas-L mRNAs in both CHB and CHC. In CHB, the intensity of both perforin and Fas-L mRNAs showed a positive correlation with the histological activity and serum alanine aminotransferase level, while the correlation was not apparent in CHC. These results suggest that both perforin and Fas/Fas-L systems are involved in the pathogenesis of liver cell injury of CHB and CHC.

High frequency of circulating HBcAg-specific CD8 T cells in hepatitis B infection: a flow cytometric analysis

Viral antigen‐specific T cells are important for virus elimination. We studied the hepatitis B virus (HBV)‐specific T cell response using flow cytometry. Three phases of HBV infection were studied: Group A, HBeAg (+) chronic hepatitis; Group B, HBeAb (+) HBV carrier after seroconversion; and Group C, HBsAb (+) phase. Peripheral T cells were incubated with recombinant HB core antigen (HBcAg), and intracytoplasmic cytokines were analysed by flow cytometry. HBcAg‐specific CD4 and CD8 T cells were identified in all three groups and the number of IFN‐γ‐positive T cells was greater than TNF‐α‐positive T cells. The frequency of IFN‐γ‐positive CD4 and CD8 T cells was highest in Group C, compared with Groups A and B. No significant difference in the HBcAg‐specific T cell response was observed between Group A and Group B. The HBcAg‐specific CD8 T cell response was diminished by CD4 depletion, addition of antibody against human leucocyte antigen (HLA) class I, class II or CD40L. Cytokine‐positive CD8 T cells without HBcAg stimulation were present at a high frequency (7 of 13 cases) in Group B, but were rare in other groups. HBcAg‐specific T cells can be detected at high frequency by a sensitive flow cytometric analysis, and these cells are important for controlling HBV replication.

A Crucial Role of Hepatocyte Nuclear Factor-4 Expression in the Differentiation of Human Ductular Hepatocytes

Ductular structures are suggested to be bipotential progenitor cells that may differentiate into hepatocytes or biliary epithelial cells (BEC). To better understand the differentiation process, we studied the expression of hepatocyte nuclear factor (HNF) in ductular structures. Matured hepatocytes in normal liver expressed HNF-1, HNF-4α, HNF-3β, and C/EBPα in the nucleus. Normal BEC expressed HNF-1 but did not express HNF-4α, suggesting an important role of HNF-4α in maintaining the phenotype of matured hepatocytes. Ductular structures were classified into ductular cells and ductular hepatocytes. Ductular cells showed glandular or bile duct-like appearance and strongly expressed cytokeratin-7. Ductular hepatocytes showed features between BEC and hepatocytes and heterogeneously expressed cytokeratin-7. Both ductular cells and ductular hepatocytes expressed HNF-4α, but the nuclear localization of HNF-4α was more prominent in ductular hepatocytes. The expression of HNF-4α mRNA in ductular hepatocytes was demonstrated at the single cell level by laser capture microdissection. Regenerative hepatocytes strongly expressed all HNFs in the nucleus, whereas residual hepatocytes in massive necrosis showed low or cytoplasmic expression. These results suggest that HNF-4α plays an important role in the differentiation and maintenance of the matured hepatocyte phenotype and that nuclear localization of HNFs is implicated in the accomplishment of their function.

CONCURRENT GASTRIC AND COLONIC LOW-GRADE MUCOSA-ASSOCIATED LYMPHOID TISSUE LYMPHOMATA IN A PATIENT WITHOUT HELICOBACTER PYLORI INFECTION

Mucosa‐associated lymphoid tissue (MALT) lymphomata observed simultaneously in the stomach and colon are rare. We report concurrent gastric and colonic low‐grade MALT lymphomata that originated from the same clone in a 58‐year‐old Japanese man without Helicobacter pylori infection. Endoscopy showed multiple erosive lesions in the gastric body and antrum, and a single flat elevation with an irregular margin in the sigmoid colon. Histopathological findings of both lesions suggested low‐grade MALT lymphoma. Lymphoepithelial lesions were evident in the gastric lesions, but not in the colonic lesion. Southern blot analysis of lymphoma cells revealed the same immunoglobulin heavy‐chain rearrangement pattern. The chromosomal translocation t(11;18)(q21;q21) was also observed. After six courses of cyclophosphamide, doxorubicin, vincristine and predonisolone, the gastric lesions disappeared endoscopically, while the colonic lesion persisted. A sigmoidectomy was consequently performed. The chromosomal translocation may be related to the pathogenesis of the present MALT lymphoma case without H. pylori infection. It is interesting that the gastric and colonic lesions differed in response to treatment and in their endoscopic and histologic features, despite having the same origin.

T cell repertoire in primary biliary cirrhosis : A common T cell clone and repertoire change after treatment

T cell repertoire was analyzed in three early-stage primary biliary cirrhosis (PBC) patients, using reverse transcription–polymerase chain reaction and single-strand conformation polymorphism. Multiple expanded clones were demonstrated in livers and peripheral blood lymphocytes (PBL) of all three patients. Comparison of the repertoire of different parts of the liver demonstrated the presence of common clones in various Vβ families. Comparison of the repertoire between the liver and PBL demonstrated that both CD4 and CD8 T cell clones were expanded. Sequence analysis of complementarity determining region 3 of the expanded clones revealed that relatively conserved amino acids were utilized in each patient and that an identical CD4 T cell clone having Vβ16 was present in all three patients. The number of expanded T cell clones in PBL decreased markedly after the treatment with prednisolone. These results suggest that common T cell clones may play a pathogenic role in PBC.