Detection of Endonuclease III-and 8-Oxoguanine Glycosylase-sensitive Base Modifications in γ-Irradiated DNA and Cells by the Aldehyde Reactive Probe (ARP) Assay

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Ionizing radiation generates diverse DNA lesions that differentially induce cell death and mutations. In the present study, calf thymus DNA (400 μg/ml) and HeLa cells were irradiated by ^<60>Co γ-rays, and abasic (AP) sites and endonuclease (Endo)III-and 8-oxoguanine glycosylase (hOGG1)-sensitive base modifications in DNA were quantitated by the aldehyde reactive probe (ARP) assay. The irradiation of calf thymus DNA in phosphate buffer generated 91 Endo III-and 100 hOGG1-sensitive base modifications and 110 AP sites per 10^6 base pairs (bp) per Gy. The yield of the lesions in Tris buffer was 41-to 91-fold lower than that in phosphate, demonstrating a radioprotective effect of Tris. The HeLa cell chromosomal DNA contained 12 Endo III-and 3.8 hOGG1-sensitive base modifications and less than 1 AP sites per 10^6 bp as endogenous damage, and their level was increased by irradiation. The yields of the damage at 1 Gy (roughly equivalent to the lethal dose of HeLa cells [1.6-1.8 Gy]) were 0.13 Endo III, 0.091 hOGG1, and 0.065 AP sites per 10^6 bp, showing that irradiation with a lethal dose brought about only a marginal increase in base damage relative to an endogenous one. A comparison of the present data with those reported for DNA strand breaks supports the primary importance of double-strand breaks and clustered lesions as lethal damages formed by ionizing radiation.
日本放射線影響学会の論文
2004-06-15