Akihiro Deguchi

Combination therapy of bezafibrate and ursodeoxycholic acid in primary biliary cirrhosis: a preliminary study

Combination therapy of bezafibrate and ursodeoxycholic acid in primary biliary cirrhosis: a preliminary study

Detection of Hepatocellular Carcinoma Using 11C-Choline PET: Comparison with 18F-FDG PET

The purpose of this study was to retrospectively investigate the feasibility of 11C-choline PET, compared with 18F-FDG PET, for the detection of hepatocellular carcinoma (HCC). Methods: A total of 16 HCC lesions in 12 patients were examined with both 11C-choline PET and 18F-FDG PET. Tumor lesions were identified as areas of focally increased uptake, exceeding that of surrounding noncancerous liver tissue. For semiquantitative analysis, the tumor-to-liver (T/L) ratio was calculated by dividing the maximal standardized uptake value (SUV) in HCC lesions by the mean SUV in noncancerous liver tissue. Results: 11C-choline PET showed a slightly higher detection rate than did 18F-FDG PET for detection of HCC (63% vs. 50%, respectively), although this difference was not statistically significant. 11C-choline PET had a better detection rate for moderately differentiated HCC lesions but not for those poorly differentiated (75% vs. 25%, respectively). In contrast, 18F-FDG PET exhibited the opposite behavior, with corresponding detection rates of 42% and 75%, respectively. The mean 11C-choline SUV and T/L ratio in moderately differentiated HCC lesions were higher than those in poorly differentiated HCC lesions. In contrast, the mean 18F-FDG SUV and T/L ratio in poorly differentiated HCC were higher than those in moderately differentiated HCC. These differences, however, were also not statistically significant. Conclusion: 11C-choline PET had a better detection rate for moderately differentiated HCC lesions but not for poorly differentiated HCC lesions, whereas 18F-FDG PET produced the opposite result. 11C-choline is a potential tracer to complement 18F-FDG in detection of HCC lesions.

Effect of the anti-diabetic drug metformin in hepatocellular carcinoma in vitro and in vivo.

Metformin is a commonly used oral anti-hyperglycemic agent of the biguanide family. Recent studies suggest that metformin may reduce cancer risk and improve prognosis. However, the antitumor mechanism of metformin in several types of cancers, including hepatocellular carcinoma (HCC), has not been elucidated. The goal of the present study was to evaluate the effects of metformin on HCC cell proliferation in vitro and in vivo, and to study microRNAs (miRNAs) associated with the antitumor effect of metformin in vitro. We used the cell lines Alex, HLE and Huh7, and normal hepatocytes to study the effects of metformin on human HCC cells. In an in vivo study, athymic nude mice bearing xenograft tumors were treated with metformin or left untreated. Tumor growth was recorded after 4 weeks, and the expression of cell cycle-related proteins was determined. Metformin inhibited the proliferation of Alex, HLE and Huh7 cells in vitro and in vivo. Metformin blocked the cell cycle in G0/G1 in vitro and in vivo. This blockade was accompanied by a strong decrease of G1 cyclins, especially cyclin D1, cyclin E and cyclin-dependent kinase 4 (Cdk4). In addition, microRNA (miRNA) expression was markedly altered by the treatment with metformin in vitro and in vivo. In addition, various miRNAs induced by metformin also may contribute to the suppression of tumor growth. Our results demonstrate that metformin inhibits the growth of HCC, possibly by inducing G1 cell cycle arrest through the alteration of microRNAs.

Efficacy of zinc administration in patients with hepatitis C virus-related chronic liver disease.

Objective. Zinc supplementation has been shown to contribute to inhibition of liver fibrosis and improvement in hepatic encephalopathy. However, little is known about the anti-inflammatory effect of zinc on hepatitis C virus (HCV)-related chronic liver disease (CLD). We therefore examined the effects of zinc administration on inflammatory activity and fibrosis in the liver of patients with HCV-related CLD. Material and methods. Polaprezinc, a complex of zinc and l-carnosine, was administrated at 225 mg/day for 6 months to 14 patients with HCV-related CLD, in addition to their ongoing prescriptions. Peripheral blood cell counts, liver-related biochemical parameters, serological markers for liver fibrosis, HCV-RNA loads, and serum levels of zinc and ferritin were evaluated before and after zinc administration. Results. Serum zinc concentrations were positively correlated with hepatic reserve before zinc supplementation. A significant increase in serum zinc level was observed after zinc supplementation (64±15 versus 78±26 mg/dl, p=0.0156). Treatment with polaprezinc significantly decreased serum aminotransferase levels (aspartate aminotransferase (AST): 92±33 versus 63±23 IU/l, p=0.0004; alanine aminotransferase (ALT): 106±43 versus 65±32 IU/l, p=0.0002), whereas alkaline phosphatase levels were significantly increased (305±117 versus 337±118 U/l, p=0.0020). Serum ferritin levels were significantly decreased by treatment with polaprezinc (158±141 versus 101±80 ng/ml, p=0.0117). The reduction rate of ALT levels by polaprezinc was positively correlated with that of ferritin (r2=0.536, p=0.0389). There was a tendency toward a decrease in serum type IV collagen 7S levels after treatment with polaprezinc. However, administration of polaprezinc did not affect peripheral blood cell counts, other liver function tests, or HCV-RNA loads. Conclusions. These findings suggest that polaprezinc exerts an anti-inflammatory effect on the liver in patients with HCV-related CLD by reducing iron overload.

Outer arm fucosylation of N-glycans increases in sera of hepatocellular carcinoma patients.

Serum glycans are promising markers for early-stage cancer detection, but the research remains challenging because low concentrations of serum glycoproteins are secreted from early-stage tumors. We have established an N-glycan profiling method using liquid chromatography electrospray ionization-mass spectrometry with high sensitive derivative, trimethyl(4-aminophenyl)ammonium chloride (TMAPA). The mass sensitivity of TMAPA-labeled oligosaccharides was enhanced more than 50 times compared with 2-aminopyridine (PA) labeled oligosaccharides, and the analytical period was significantly shortened compared with traditional HPLC 2D-mapping. Using this method, we found about 28 major N-linked oligosaccharides in human sera, and we investigated their alterations in patients who developed hepatocellular carcinoma (HCC). We found that outer arm fucosylation (attached GlcNAc via an alpha 1-3/4 linkage) in highly branched oligosaccharides increased significantly in sera of HCC patients. Normalizing the level of outer arm fucosylation by taking into account platelet concentration allowed us to distinguish more clearly between HCC and LC patients.

Repeated adenoviral administration into the biliary tract can induce repeated expression of the original gene construct in rat livers without immunosuppressive strategies

Background: Systemic adenoviral readministration appears to be limited by immunogenicity. Aims: We examined the feasibility of repeated adenovirus mediated gene transfer into the liver via the biliary tract. Methods: Recombinant adenoviruses carrying a reporter lacZ gene were infused retrogradely into the common bile duct of rats. Transduction efficiency of the lacZ gene was estimated histochemically and quantitatively. Results: Retrograde administration of recombinant adenoviruses into the common bile duct of rats resulted in efficient transgene expression in the liver, specifically in hepatocytes, but not in biliary epithelia. Transduction efficiency induced by intrabiliary adenoviral administration was not substantially different from that induced by intraportal adenoviral infusion. Transgene expression in the liver was however transient, and development of neutralising antibodies against adenovirus was observed in serum but not in bile. When adenoviruses were readministered into the common bile duct, successful re-expression of the transgene in the liver was achieved despite the existence of neutralising antibodies in serum. Interestingly, although proliferation of adenovirus specific T cells in response to adenoviral readministration was suppressed significantly by immunosuppressive FK506 treatment, levels of transgene expression in the liver achieved by intrabiliary adenoviral readministration were not significantly different between animals treated with and without FK506. Furthermore, third adenoviral administration into the common bile duct also induced successful transgene expression in the liver. Conclusions: These results suggest that adenovirus mediated gene transfer into the liver may be repeatable without immunosuppressive strategies in clinical settings by means of endoscopic retrograde cholangiography.

CD28-negative CD8-positive cytotoxic T lymphocytes mediate hepatocellular damage in hepatitis C virus infection.

The pathogenic mechanism for hepatocellular damage in hepatitis C virus (HCV) infection has not been clearly understood. Analysis of costimulatory molecules on lymphocytes may give us insight into the pathogenic mechanism of hepatocellular damage in HCV infection. Peripheral blood mononuclear cells (PBMCs) and liver infiltrating mononuclear cells (LIMCs) isolated from the HCV-infected patients were analyzed with antibodies directed against a variety of costimulatory molecules by flow cytometry. Blocking experiment against HLA-A24-restricted HCV-specific CTLs and immunohistochemical analysis were also performed. PBMCs expressing CD8, CD28, CD80, or CD154 were significantly reduced in HCV-infected patients compared with the healthy controls. CD28(+)CD8(+) PBMCs in the patients inversely correlated with ALT levels. Conversely, levels of CD28(−)CD8(+) LIMCs correlated with ALT levels. HCV-specific CTL activity was blocked by the treatment with anti-CD8 antibody, but not with anti-CD4 or anti-CD28 antibody. Immunohistochemical analysis revealed the accumulation of CD28(+) cells around the portal area in the liver of a patient with chronic active hepatitis C. These results suggest that CD28(+)CD8(+) T cells leave the circulation, move to the livers, and are activated in the portal area in proportion to the extent of liver diseases. CD28(−)CD8(+) T cells may finally function as effector T cells causing the hepatocellular damage in HCV infection.

Histopathologic comparison of anti-mitochondrial antibody-positive primary biliary cirrhosis and autoimmune cholangiopathy

The present study aimed to investigate whether antimitochondrial antibody (AMA)-positive primary biliary cirrhosis (PBC) and AMA-negative PBC with autoantibodies differ histologically, especially with respect to infiltrating cells in portal tracts involved by chronic non-suppurative destructive cholangitis. Liver specimens were stained from 15 primary biliary cirrhosis with AMA (group 1), nine patients consistently negative for AMA but positive for antinuclear antibodies (ANA) (group 2). Group 2 showed overlapping features of PBC and autoimmune hepatitis type 1, in a pattern recently termed autoimmune cholangiopathy (AIC). We analyzed the cell population, including lymphocytes, plasma cells, large histiocytes, eosinophils and neutrophils, which had infiltrated portal tracts involved by destructive lesions. Although serum immunoglobulin M levels were higher in group 1 compared to those in group 2 (P=0.0282), patients of both groups were broadly comparable with respect to clinical features and laboratory data. Histologically, the number of plasma cell and its percentage among inflammatory infiltrating cells in the portal tract were higher in group 2 than in group 1 (P=0.0015, P=0.0070, respectively). The percentage of lymphocyte infiltration among inflammatory infiltrating cells in the portal tract were higher in group 1 than in group 2 (P=0.0052). The percentage of plasma cell infiltration among inflammatory infiltrating cells in the portal tract was correlated to immunoglobulin G levels in group 2 (r=0.949, P=0.0016). In conclusion, AMA-positive PBC and AIC showed differences in inflammatory cell population in involved portal tracts in this preliminary study.

Profile of microRNAs associated with aging in rat liver.

Recent studies suggest that small non‑coding microRNAs (miRNAs or miRs) play an important role in the regulation of genes involved in various cellular and developmental processes. However, the expression of miRNAs during the aging process remains largely unknown. The aim of the present study was to analyze miRNA expression profiles in rat livers during the aging process. The livers of male Wistar rats at different stages of development (fetal, aged 3 days, and 1, 2, 4, 8 and 36 weeks of age) were used. Total RNA was extracted from the livers. We analyzed the expression levels of 679 rat miRNA probes. In addition, immunohistochemical staining for proliferating cell nuclear antigen (PCNA) was performed. Several up- and downregulated miRNAs were identified in the rat livers at 7 different fetal developmental stages and at 36 weeks of age. We observed the upregulation of miR‑29a, miR‑29c, miR‑195 and miR‑497, whereas miR‑301a, miR‑148b-3p, miR‑7a, miR‑93, miR‑106b, miR‑185, miR‑450a, miR‑539 and miR‑301b were downregulated in the aging rat livers. The number of PCNA-positive hepatocytes was decreased with age. In conclusion, our findings suggest that these up- and downregulated miRNAs play an important role in aging by regulating cell cycles that are involved in liver senescence. Further investigation is required to reveal additional target genes of the miRNAs expressed in the liver and the roles of miRNAs in the developmental process of aging in the liver.

Clinical characteristics of patients with autoimmune hepatitis seropositive for anticentromere antibody

Aim:  Recent studies have revealed that primary biliary cirrhosis patients with anticentromere antibody (ACA) commonly develop portal hypertension. However, the clinical characteristics of autoimmune hepatitis (AIH) remain uncertain. We investigated the clinical features of patients with AIH seropositive for ACA (ACA‐AIH), comparing them with those of patients with AIH seropositive for other immunofluorescent patterns of antinuclear antibodies (ANA) (other‐AIH).