文章基本信息

标题：Analysis of 8-Hydroxyguanine (8-OH-Gua) Released from DNA by the Formamidopyrimidine DNA Glycosylase (Fpg) Protein: A Reliable Method to Estimate Cellular Oxidative Stress
其他标题：Analysis of 8-Hydroxyguanine (8-OH-Gua) Released from DNA by the Formamidopyrimidine DNA Glycosylase (Fpg) Protein: A Reliable Method to Estimate Cellular Oxidative Stress
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作者：Hiroshi ORIMO ; Nan MEI ; Serge BOITEUX 等
期刊名称：Journal of Radiation Research
印刷版ISSN：0449-3060
电子版ISSN：1349-9157
出版年度：2004
卷号：45
期号：3
页码：455-460
DOI：10.1269/jrr.45.455
摘要：To improve the analyses of a form of oxidative DNA damage, 8-hydroxyguanine (8-OH-Gua), we treated isolated DNA with formamidopyrimidine DNA glycosylase (Fpg) and analyzed the released 8-OH-Gua by using a high-performance liquid chromatography system equipped with an electrochemical detector (HPLC-ECD). The human lung carcinoma cells (A549) and human keratinocyte (HaCaT) were irradiated with γ-rays. After the isolated DNA was treated with the Fpg protein, we analyzed the released 8-OH-Gua by using an HPLC-ECD. With this method, the background level of 8-OH-Gua in DNA from human lung carcinoma cells was determined to be 3.4 residues per 10 7 guanine (Gua). A similar background level of 8-OH-Gua (3.1 residues per 10 7 Gua) was also detected in human keratinocyte DNA with this method. These background 8-OH-Gua levels in cellular DNA are comparable to that obtained previously by an analysis of 8-OH-dGMP after nuclease P1 digestion of cellular DNA (4.3 residues per 10 7 dCMP). A dose-dependent increase of 8-OH-Gua (0.17/10 7 Gua/Gy) was observed after cells were irradiated with γ-rays. Twenty hours after γ-irradiation with 60 Gy, 75% of the 8-OH-Gua produced in keratinocyte DNA was repaired. With our new analysis method, it is possible to detect the small changes in the 8-OH-Gua levels in cellular DNA induced by various environmental factors.
关键词：8-OH-Gua background level; Fpg protein; γ-irradiation