| Project/Area Number |
10557248
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 展開研究 |
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| Research Field |
Laboratory medicine
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| Research Institution | Tokyo Medical and Dental University |
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Principal Investigator |
TERAOKA Hirobumi Tokyo Medical and Dental University Medical Research Institute Professor, 難治疾患研究所, 教授 (30019137)
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| Co-Investigator(Kenkyū-buntansha) |
SAITO Keiko Tokyo Medical and Dental University Medical Research Institute Research Assistant, 難治疾患研究所, 教務職員 (50178969)
YANOKURA Mieko Tokyo Medical and Dental University Medical Research Institute Research Assistant, 難治疾患研究所, 教務職員 (50143615)
TAKASE Kozo Tokyo Medical and Dental University Faculty of Medicine Professor, 医学部, 教授 (90211333)
YAMAMOTO Kohtaro Tokyo Medical and Dental University Medical Research Institute Professor, 難治疾患研究所, 教授 (40000971)
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| Project Period (FY) |
1998 – 1999
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| Project Status |
Completed (Fiscal Year 1999)
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| Budget Amount *help |
¥13,000,000 (Direct Cost: ¥13,000,000)
Fiscal Year 1999: ¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1998: ¥11,000,000 (Direct Cost: ¥11,000,000)
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| Keywords | flow cytometry / DNA strand breaks / SCID / cell cycle / β-ray / DNA double-strand breaks / V(D)J recombination / apoptosis |
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| Research Abstract |
The purpose of this research is to estimate quantitatively chromosomal DNA strand breaks by flow cytometry. At first, we analyzed relationship between cell cycle progression and DNA strand breaks induced by incorporation of [ィイD13ィエD1H]thymidine into human hemopoietic cell lines, such as HL-60, Molt-4 and Raji. The ends of chromosomal DNA strand breaks in a cell were labeled with the FITC-dUTP/TdT system. In 7.4 kBq [ィイD13ィエD1H]thymidine/ml, cell cycle of HL-60 progressed with a slight increase in the population of S-phase cells, and the relative value of FITC, which reflects the number of DNA strand breaks per cell, were significantly high at 8 and 18 h with decrease to the basal level at 24 h. Analysis of HィイD22ィエD2OィイD22ィエD2-resistant HL-60 variants revealed involvement of reactive oxygen species in the effects of [ィイD13ィエD1H]thymidine incorporation on cell cycle and cell fate. A human glioma cell line, M059J, lacking DNA-PK involved in DNA double-strand break repair showed higher sensitivity to neocarzinostatin (NCS) compared with M059K containing wild-type DNA-PK. Addition of NCS at 50 ng/ml resulted in increase in S-phase cell population at 30 min in M059J with gradual increase in FITC signals. In contrast, there were no significant changes in DNA histogram patterns in M059K in the presence of 50 ng NCS/ml, and transient increase in the relative value of FITC was observed maximally 2 h after the addition. These results demonstrated that the number of chromosomal DNA strand breaks per cell could be estimated semi-quantitatively by flow cytometry. Now it is possible to predict efficacy and side effects of individual therapy with radioactive materials and radiomimetic drugs.
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