Jun Tojo

Interleukin-6 induces proliferation of rat hepatocytes in vivo

Abstract Background/Aims: The aim of this study was to assess the effect of interleukin-6 (IL-6) on the proliferation of hepatocytes and to study the interaction between IL-6 and hepatocyte growth factor (HGF) in vivo . Methods: IL-6 was injected at a dose of 200 μg/mg subcutaneously into rats every day for 14 days. Liver and blood samples were obtained at 1, 3, 7 and 14 days during IL-6 administration. Hepatocyte proliferative activity of sera was measured using 3 H-thymidine incorporation into cultured rat hepatocytes. To evaluate the proliferative activity of the hepatocytes in tissue sections, hepatic DNA content and immunostaining of the liver tissue sections for proliferating cell nuclear antigen (PCNA) were performed. Plasma HGF levels were measured using specific EIA. In addition, total RNA was extracted from the liver and expression of HGF mRNA was detected by RT-PCR. Results: The DNA contents of liver taken from IL-6-treated rats were increased during IL-6 administration compared with untreated rats. Sera taken from IL-6-treated rats at various intervals during administration also significantly increased 3 H-thymidine incorporation by cultured rat hepatocytes compared with sera from untreated rats, suppressing 3 H-thymidine incorporation at day 1 and 3 by anti-HGF antibody. IL-6 itself did not increase 3 H-thymidine incorporation. Increased expression of PCNA in these hepatocytes was noted from 1 day after IL-6 administration, and at 14 days, the number of PCNA-positive cells was sevenfold greater than in the livers of untreated rats. However, plasma HGF levels showed a peak at day 1, decreased gradually from day 3, and became undetectable by day 14. HGF mRNA expression in livers of IL-6-treated rats was suppressed from day 3 to day 14 of IL-6 administration. Conclusions: These data show that IL-6 induces an early phase of liver cell growth in vivo and suggest that an increase level of HGF mediates this effect.

Clinicolaboratory characteristics of patients with primary biliary cirrhosis associated with CREST symptoms.

Aims: Patients with primary biliary cirrhosis (PBC) occasionally suffer complications from other autoimmune diseases. When PBC was associated with calcinosis, Raynauds phenomenon, esophageal dysmotility, sclerodactyly and telangiectasias (CREST) symptoms, it has been proposed that it is a distinct clinical entity. This study aimed to investigate whether PBC associated with CREST symptoms is a distinct disease complex. Method: Clinicolaboratory data, HLA type of leukocytes and disease prognosis were compared between 31 patients with PBC associated with CREST symptoms and 68 patients with PBC alone. Results: The characteristic findings and significant differences observed in patients with PBC associated with CREST symptoms compared with PBC alone are as follows: all women with older age with milder clinical features of both PBC (asymptomatic PBC in 84%) and CREST syndrome (incomplete CREST in 81%), more frequent occurrence of esophageal varices (28.6 vs. 9.3%), better prognosis (87.5 vs. 45.5% in 10 years survival), lower serum levels of AST (39.8 vs. 63.6 IU/l) and IgM (460 vs. 676 mg/dl), higher prevalence of discrete speckled pattern of antinuclear antibodies (93.5 vs. 12.3%), higher median titers of anti-CENP-B antibodies (1.22 vs. 0.31), lower median titers of antimitochondrial antibody (1:80 vs. 1:160), and a higher prevalence of HLA-DR9 (54.5 vs. 24.3%). Conclusion: These findings support the presence of a subgroup in PBC as PBC associated with CREST symptoms.

A case of hepatic inflammatory pseudotumor with primary biliary cirrhosis.

Inflammatory pseudotumor (IPT) of the liver is an unusual non-neoplastic benign lesion. A 75-year-old man was hospitalized for esophageal varices and gastric cancer. Three years before admission, he had been diagnosed as having primary biliary cirrhosis (PBC) without Sjögrens syndrome. Computed tomography (CT) and magnetic resonance imaging (MRI) scans showed multiple masses (S3, S5, S6) less than 2 cm in diameter in the liver. Since these masses were difficult to distinguish from hepatocellular carcinoma, or metastatic liver carcinoma, one of the masses (S5) was removed during an operation for gastric cancer. Histological examination demonstrated marked infiltration of plasma cells and some histiocytes, findings consistent with the histological features of IPT. The coexistence of hepatic IPT and PBC in this case may have been an accidental event. However, the immunological and environmental factors associated with PBC are thought to be involved in the development of IPT; in addition, cholangitis associated with PBC could have contributed to the development of IPT.

Lipo prostaglandin E1 reduces the production of CXC chemokines in endotoxin-induced rat liver injury

The present study attempted to assess the effect of prostaglandin E1 (PGE1) incorporated into lipid microspheres (Lipo PGE1) on chemokine production in endotoxin-induced rat liver injury. Male Wistar rats weighing 200-250 g were injected with 2 mg lipopolysaccharide (LPS) per kg intravenously. Lipo PGE1 was administered simultaneously at various concentrations (0.002, 0.02, 0.2, 2 µg/kg) in the tail vein. Blood samples and liver specimens were taken from the rats at 1, 3, 8, 12 and 24 h after injection with LPS alone or with LPS and Lipo PGE1. Serum macrophage inflammatory protein-2 (MIP-2) and cytokine-induced neutrophil chemoattractant (CINC) levels were measured by the enzyme-linked immunosorbant assay using the corresponding antibodies. Liver specimens were fixed, and the number of neutrophils that had infiltrated each liver section was determined under a microscope. Serum alanine aminotransferase (ALT) levels were significantly lower in the rats injected with LPS and Lipo PGE1 compared with those in the rats injected with LPS alone, and this difference was expressed in a PGE1 dose-dependent manner. Serum MIP-2 levels were significantly lower at 3 h (141.4+/-95.5 pg/ml) and 8 h (44.9+/-44.7 pg/ml) after injection with LPS and Lipo-PGE1 (2 µg/kg) than at the same times after injection with LPS alone (342.9+/-35.9 and 358.3+/-23.4 pg/ml, respectively). Similarly, serum CINC levels were significantly lower at 8 h (482.7+/-156.0 ng/ml) after injection with LPS and Lipo-PGE1 (2 µg/kg) than at the same time after injection LPS alone (723.3+/-29.0 ng/ml). No significant differences were observed at any time between serum tumor necrosis factor-alpha (TNF-alpha) levels in rats injected with LPS alone and in rats injected with LPS and Lipo-PGE1 (2 µg/kg). The number of neutrophils that had infiltrated the liver was significantly lower at 8 h after injection with LPS and Lipo PGE1 than at the same time after injection with LPS alone. This difference was expressed in a Lipo PGE1 dose-dependent manner. In conclusion, Lipo PGE1 reduces liver injury and serum levels of MIP-2 and CINC, but not TNF-alpha, in rats injected with LPS and also reduces the number of neutrophils that infiltrate in the liver.

High serum levels of vascular endothelial growth factor in patients with human hepatocellular carcinoma

Vascular endothelial growth factor (VEGF) is intimately involved in neovascularization. In addition, it is know that in human hepatocellular carcinoma (HCC), angiogenesis is indispensable for tumor growth. In this study, we measured the serum VEGF levels of patients with HCC and studies the relationship between the serum VEGF level and maximum tumor diameter as well as that between the serum VEGF level and the serum α-fetoprotein (AFP) level. Mean serum VEGF level were 5.33 ± 0.77, 3.97 ± 0.68, 2.64 ± 0.78, and 2.57 ± 0.97 ng/ml for patients with HCC, chronic hepatitis (CH), or liver cirrhosis (LC) and normal controls (NC), respectively, with that of the HCC patients being significantly (P < 0.05) higher than that of the LC patient or NC. In addition, the serum VEGF level was significantly (r = 0.53, P < 0.05) correlated with the maximum tumor diameter in the HCC patients, and the sera of the patients with hypervascular HCC showed a significantly (P < 0.01) higher VEGF titer than the sera of the patients with isovascular or hypovascular HCC. However, there was no significant correlation between serum VEGF level and serum AFP level. These findings suggest that VEGF may play an important role, apart from that in AFP production, in the development of HCC.

T-cell clonality of peripheral blood from patients with chronic hepatitis C before and after treatment with interferon α

Immune-mediated liver cell damage has been likely to occur in patients with chronic hepatitis C virus (HCV) infection. To clarify the T-cell clonality in patients with chronic hepatitis C, we analyzed T-cell receptor (TCR) Vβ chain messages expressed in peripheral blood mononuclear cells (PBMC) before and after treatment with interferon (IFN)-α. PBMC from 15 patients with chronic hepatitis C and from six healthy subjects were examined. The 15 patients were allocated to three groups based on their response to IFN-α treatment as follows: responders (group A, n=5), transient responders (group B, n=5), and non-responders (group C, n=5). Twenty-two TCR Vβ repertoires Vβ 1–Vβ 20 expressed in PBMC were analyzed by reverse-transcription polymerase chain reaction (RT-PCR) and single strand conformation polymorphism (SSCP). In responders, the mean total number of T-cell clones in 22 TCR Vβ repertoires was significantly decreased from 97.4±40.1 to 62.6±32.1 (P<0.01) after treatment. In transient responders, although one patient showed a decrease, four patients showed an increase, with the mean total number of T-cell clones increasing from 66.4±28.0 to 79.8±28.8 after treatment. In non-responders, the mean total number of T-cell clones was almost unchanged between before and after treatment. No clear tendency towards the use of any specific Vβ was observed in any group of patients. These results suggest that T-cell clones accumulated in the peripheral blood of patients with chronic hepatitis C and that the number of clones may change in response to IFN-α treatment.

Antithrombin III stimulates prostaglandin I2 production by cultured rat hepatic sinusoidal endothelial cells

Abstract We examined the production of prostanoids, specifically prostaglandin (PG) I2 and thromboxane (TX) A2, in cultured rat hepatic sinusoidal endothelial cells treated with antithrombin (AT) III. When cells were treated for 3 h with various concentration of AT III, production of 6-keto-PGF1α (a stable metabolite of PG I2) increased significantly and in a dose-dependent manner, compared with production by untreated cells. In terms of kinetics, significant increases were noted at 3 h (20.13 ± 1.38 pg/ml), 4 h (21.23 ± 0.63 pg/ml) and 24 h (36.58 ± 4.93 pg/ml) with AT III (300 μg/ml) stimulation, compared with production by the untreated cells (10.29 ± 1.21, 10.34 ± 1.66 and 22.64 ± 2.59 pg/ml, respectively). Moreover, this production was significantly reduced with increasing concentrations of heparin. On the other hand, TX B2 (a stable metabolite of TX A2) production was unaffected by AT III treatment. These data suggest that AT III may ameliorate the liver damage or disturbances in the sinusoidal microcirculation by stimulating the PG I2 production of hepatic sinusoidal endothelial cells.