| Co-Investigator(Kenkyū-buntansha) |
ANDO Koichi Researcher, International Spece Radiation Laboratory, National Institute of Radiological Sciences, 研究員 (00159526)
HACHIYA Misao Researcher, Division of Radiation Health, National Institute of Radiological Sciences, 放射線障害医療部, 研究員 (00198756)
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| Budget Amount *help |
¥3,200,000 (Direct Cost: ¥3,200,000)
Fiscal Year 1999: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 1998: ¥2,000,000 (Direct Cost: ¥2,000,000)
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| Research Abstract |
Myeloid cells have various functions against external insults such as bacterial infection. In response to the insults, these cells generate hydrogen peroxide (H2O2) and hypochlorous acid (HOCl), a more potent microbicidal agent. Thus, hydrogen peroxide plays a key role in host defense against a variety of microorganisms. However, elevated levels of H2O2 may lead to overproduction of HOCl, which is, in turn, toxic to the cells. The HL6O is a cell line derived a patient with acute promyelocytic leukemia (M3), and using a variant clone of HL6O that is resistant to H2O2, we studied the role of H2O2 in these cells. When the growth curves of both cell tines were compared, the HP100 grew significantly faster than the HL60. Treatment of the HL60 cells with catalase (CAT) stimulated their cell growth. Further studies found that a sublime HP100 was relatively resistant to γ- irradiation and tumor necrosis factor α (TNFα) as compared to the parent cells. The increased production of H2O2 was seen
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in response to irradiation and TNFα in HL60 cells but not in HP100 cells. We also determined the levels of apoptosis-related proteins in both cell lines. The Bax was constitutively expressed in both cell lines. On the other hand, Bcl-2 was expressed in the HP100 cells but not in the HL60. However, no changes in the levels-of proteins were observed upon exposure to-either H2O2, γ-ray, or TNFα. These results suggest that apoptosis by H2O2 may occur through an independent of Bax/Bcl-2 pathway in the HL60 cells. To investigate further mechanisms for apoptosis in these cells, we compared the expression of H2O2-related genes. HP100 cells had much higher levels of CAT and c-myc mRNAs than HL60 cells. No difference in the levels of manganese superoxide dismutase (MnSOD), glutathione peroxidase (GSH-Px), and CuZnSOD mRNAs were observed between the two cell lines. Treatment with either γ-ray, TNFα, or H2O2 increased the levels of MnSOD and TNFα transcripts dose-dependently in the HL60 cells but did not affect those in the HP100 cells. Our findings suggest that endogenously produced H2O2 plays some role in the growth of the HL60 cells. We also conclude that irradiation and TNFa induce apoptosis through, at least in part, the generation of H2O2 ; increased levels in MnSOD or TNFα but not in CAT or GSH-Px may also lead to the accumulation of H2O2, which results in apoptosis. Less
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