| Budget Amount *help |
¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 1993: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 1992: ¥1,400,000 (Direct Cost: ¥1,400,000)
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| Research Abstract |
Reactive oxygen species (ROS) is one of the most effective species to cause disease. Among ROS induced damages, DNA damage is important in relation to cancer and aging. We established systems to determine typical mutagenic oxidative DNA damages, 8-hydroxydeoxyguanosine (8OHDG) and DNA strand breaks (SB). We could determine 8OHDG in cellular DNA quantitatively by HPLC with electrochemical detection. We could detect SB by pulsed-field gel electrophoresis, however, the method was not suitable for quantitative assay. Lymphoblasts from Fanconi's anemia patients (whose cells are hypersensitive to ROS) induced 8OHDG more than those from controls with H202 at 20 mM.The defference was indicated to be derived from decreased catalase activity in patients' cells. To study the effect of asbestos, we used human promyelocytic leukemia cell line, HL60, which defferentiates to neutrophils and macrophages. Opsonized asbestos induced O2- release from neutrophil-like HL60, however, it did not induce 80HdG-increase in the cellular DNA.On the other hand, asbestos did not increase ROS release from macrophage-like HL60, it was phagocytozed by the cell and increased 80HdG.The 80HdG-increase was not inhibited by SOD or catalase, however, it was inhibited by cytochalasin B, a inhibitor for phagocytosis. For the evaluation of oxidative DNA damage in populations, we determined 8OHdG in white blood cells (WBC) from factory workers. Polymorphonuclear cells which release ROS upon various stimuli, contained similar 8OHdG levels to mononuclear cells. Contaminated red blood cells in the cell preparation increased 8OHdG artificially. We could not find correlations between 8OHdG levels and age or lifestyle factors, probably because the distribution range of 8OHdG levels in WBC was too small against the experimental error of 8OHdG determination.
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