Yasuhiro Mizoguchi

Effects of an anti-tumor polysaccharide, schizophyllan, on interferon-γ and interleukin 2 production by peripheral blood mononuclear cells

We examined the effect of schizophyllan, a neutral glucan isolated from the culture filtrate of Schizophyllum commune Fries, on the production of interferon-gamma (IFN-gamma) and interleukin 2 (IL 2) from the mitogen-stimulated human peripheral blood mononuclear cells (PBMC). When the levels of IFN-gamma and IL 2 in the culture medium of phytohemagglutin (PHA)- or concanavalin A (Con A)-stimulated PBMC were measured by radioimmunoassay and enzyme-linked immunosorbent assay (ELISA) respectively, a significant increase in the production of both cytokines by schizophyllan was demonstrated. These results suggest that the increased production of IFN-gamma and IL 2 may be responsible for the anti-tumor activity of this glucan.

Release of peptide leukotrienes from rat kupffer cells

Kupffer cells isolated from the normal rat liver were incubated with calcium ionophore A23187, and the levels of peptide leukotrienes (LTC4, LTD4, and LTE4) contained in the culture supernatant were determined by the combined technique of reverse-phase high-performance liquid chromatography and radioimmunoassay. In response to A23187, Kupffer cells released LTC4, LTD4, and LTE4. After 10 min-preincubation of Kupffer cells with AA861, a 5-lipoxygenase inhibitor, the generation of LTC4, LTD4, and LTE4 from A23187-stimulated Kupffer cells was significantly suppressed. Platelet activating factor (PAF), a phospholipid mediator, significantly enhanced the release of LTC4, LTD4, and LTE4 from Kupffer cells stimulated with A23187. These results suggested that Kupffer cells may participate in inflammatory and immunologic events in the liver tissue by the release of peptide leukotrienes.

Inhibition of DNA synthesis by methylglyoxal bis(guanylhydrazone) during lymphocyte transformation

The phytohemagglutinin induced DNA synthesis in guinea pig lymph node cells was inhibited remarkably by methylglyoxal bis(guanylhydrazone). This inhibitory effect was dependent on the time of its addition to the lymph node cell culture after stimulation with phytohemagglutinin. If methylglyoxal bis(guanylhydrazone) was added 48 hr after the stimulation, no inhibition of DNA synthesis was observed. Exogenous spermidine added at an early time of cell culture reversed the inhibitory effect of methylglyoxal bis(guanylhydrazone). However, no reversion occurred when spermidine was added at a late time of the cell culture.

Identification and fine structure of proliferating hepatocytes in malignant and nonmalignant liver diseases by use of a monoclonal antibody against DNA polymerase alpha

To analyze the process of liver regeneration and the initiation of hepatocellular carcinoma (HCC), we studied histochemically the morphologic features of proliferating parenchymal cells stained for DNA polymerase alpha (DPA), in 31 patients with various diseases, by use of a monoclonal antibody against DPA. In specimens from patients with acute viral hepatitis with confluent necrosis, most stained hepatocytes were small, with basophilic cytoplasm, and were located next to the necrotic areas. Under electron microscopy, stained granules were seen in the nucleus. Most stained hepatocytes had immature organelles. In specimens from patients with cirrhosis of the liver, the number of stained hepatocytes greatly differed in different pseudolobules. In specimens from patients with adenomatous hyperplasia, stained hepatocytes, mostly small and basophilic, were found diffusely; electron microscopy showed slightly indented nuclei with a few organelles and less condensed chromatin than normal. In specimens from patients with HCC, most stained cancer cells were small and basophilic; electron microscopy showed indented nuclei with a few organelles and less than normal condensed chromatin. Staining showed that during regeneration, immature hepatocytes reentered the cell cycle and repaired a large necrotic area. It was conceivable that in the initiation of HCC, some small hepatocytes with indented nuclei and less condensed chromatin might become HCC cells.

Effects of arachidonic acid metabolites and interleukin-1 on platelet activating factor production by hepatic sinusoidal endothelial cells from mice

The effects of interleukin‐1 (IL‐1) and arachidonic acid metabolites, prostaglandin (PG) E1, leukotriene (LT) B4 and LTC4, on the amount of platelet activating factor (PAF) produced and released by mouse hepatic sinusoidal endothelial cells were investigated. When hepatic sinusoidal endothelial cells were incubated with 1 μg/mL of calcium ionophore (CaI) A23187, PAF production increased until 20 min after the start of incubation, but when the cells were incubated with PGE1 and CaI A23187, PAF production was significantly suppressed. On the other hand, when hepatic sinusoidal endothelial cells were incubated with IL‐1β, LTB4 or LTC4 alone, PAF production significantly increased compared with that of the cells incubated without these substances. However, when the cells were incubated with PGE1 alone or with IL‐1β, LTB4 or LTC4 and CaI A23187, these effects were not observed. These results indicated that PAF produced by hepatic sinusoidal endothelial cells is affected by IL‐1 and arachidonic acid metabolites, suggesting that immune and inflammatory reactions in these cells may be regulated by chemical mediators produced by the cells themselves.

Calcium-dependent prostaglandin biosynthesis by lipopolysaccharide-stimulated rat Kupffer cells

Isolated rat Kupffer cells produced and released prostaglandin (PG) E2, 6-keto-PGF1 alpha, and thromboxane B2 (TXB2) in response to lipopolysaccharide (LPS) stimulation. This elevation of PGE2, 6-keto-PGF1 alpha and TXB2 in the medium was not observed when cells were cultured in the absence of extracellular calcium or in the presence of an extracellular calcium chelator, EGTA. An intracellular calcium antagonist, TMB-8, also suppressed the production of PGE2, 6-keto-PGF1 alpha and TXB2 in a concentration-dependent manner. The intra-cellular calcium concentration of Kupffer cells elevated early after the addition of LPS determined by the use of fura-2 and a fluorescence microscopy. Moreover, calmodulin inhibitors, W-7 and W-13, apparently inhibited the production of PGF2, 6-keto-PGF1 alpha and TXB2. All these results suggest that LPS-induced PG production by stimulated rat Kupffer cells may be regulated by a calcium-calmodulin pathway.

There is No Correlation Between Function and Lymphokine Production of HBs‐Antigen‐Specific Human CD4+‐Cloned T Cells

The question whether antigen‐specific human CD4+ T cells can be classified on the basis of appropriate and fixed lymphokine production patterns and their corresponding functions still remains lo be elucidated. We generated ten CD4+ T‐cell clones specific for HBsAg from HBsAg‐positive but HBsAb‐negative individuals. Seven of these clones exhibited helper activity for HBsAb response, while the three other clones did not. Both helper‐ and non‐helper‐type T‐cell clones produced interleukin 4 (IL‐4) after antigenic stimulation. By stimulation with phytohaem‐agglutinin (PHA) plus phorbol myristate acetate (PMA). three of the seven helper‐type clones produced interleukin 2 (lL‐2) in addition toiL‐4. However, the other four helper‐type clones did not produce IL‐2 by such stimulation, although they continued the production of IL‐4. All non‐help)er‐type T‐cell clones produced a large amount of IL‐2. and some of them completely became an IL‐2 producer after certain stimulation. These results suggested that both helper‐ and non‐helper‐type CD4+ T‐cell clones specific for HBsAg might have no strict pattern of lymphokine production as m the TH1 /TH2 dichotomy of murine CD4+ T cells. The data also revealed that lymphokine‐producing capacity of individual cloned T cells is changeable depending upon the sort of activation.

Arachidonic acid metabolites in carbon tetrachloride-induced liver injury

SummaryThe changes in the levels of leukotrienes (LTs) and prostaglandins (PGs) in the liver tissue of rats with carbon tetrachloride (CCl4-induced liver injury were studied. As a result, after the administration of CCI4, the levels of LTs increased at an early stage while the levels of PGs increased at a later stage. This suggests that LTs may have an adverse effect on liver injury induced by CC14, and that PGs may not have a direct effect on liver injury. In addition, serum GOT and GPT levels improved with the administration of AA-861, a 5-lipoxygenase inhibitor, while these levels did not change with the administration of indomethacin, a cyclooxygenase inhibitor. These results suggest that arachidonic acid metabolites may play an important role in the induction of liver cell injury.

Control of ornithine decarboxylase activity by cyclic nucleotides in the phytohemagglutinin induced lymphocyte transformation

Summary N2,O2′-Dibutyryl guanosine 3′:5′-cyclic monophosphate enhanced the ornithine decarboxylase activity which was induced during lymphocyte transformation by phytohemagglutinin, while N6,O2′-dibutyryl adenosine 3′:5′-cyclic monophosphate tended to inhibit it. The increased ornithine decarboxylase activity was not observed when N2,O2′-dibutyryl guanosine 3′:5′-cyclic monophosphate alone was added to the non-stimulated lymphocytes. In contrast to the dibutyryl derivative, guanosine 3′:5′-cyclic monophosphate did not show any effect on enzyme induction following phytohemagglutinin stimulation.

Changes in leukotrienes and prostaglandins in the liver tissue of rats in the experimental massive hepatic cell necrosis model

When heat-killed Propionibacterium acnes is intravenously injected into rats followed by an intravenous injection of a small amount of Gram-negative lipopolysaccharide (LPS) 7 days later, massive hepatic cell necrosis is induced and most of the rats die within 24 hours of LPS injection. Using this experimental model, we studied the changes in the levels of leukotrienes (LTs) and prostaglandins (PGs) in the liver tissue and bile of rats with experimentally-induced massive hepatic cell necrosis. Both the levels of LTs and PGs in the liver tissue and LTs in the bile increased before the microscopic appearance of hepatic cell necrosis. These results suggest that arachidonic acid metabolites may play an important role in the induction of liver cell injury.