Journal Article

Glutathione and ascorbate are negatively correlated with oxidative DNA damage in human lymphocytes

Kevin J. Lenton, Hélène Therriault, Tamàs Fülöp, Hélène Payette and J. Richard Wagner

in Carcinogenesis

Volume 20, issue 4, pages 607-613
Published in print April 1999 | ISSN: 0143-3334
Published online April 1999 | e-ISSN: 1460-2180 | DOI: http://dx.doi.org/10.1093/carcin/20.4.607
Glutathione and ascorbate are negatively correlated with oxidative DNA damage in human lymphocytes

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Intracellular antioxidants, glutathione and ascorbate, and two molecular markers of oxidative DNA damage, 5-hydroxy-2′-deoxycytidine (5-OH-dCyd) and 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxo-dGuo), were measured in lymphocytes from 105 healthy volunteers. The analysis of 5-OH-dCyd and 8-oxo-dGuo was carried out by HPLC with electrochemical detection such that both compounds were detected on the same chromatography run. There was no significant difference in oxidative DNA damage when the extraction of DNA from cells using phenol was carried out under anaerobic conditions or in the presence of metal ion chelators. This indicates that auto-oxidation of DNA during sample preparation was minimal. Using the above methods, the average level of oxidative DNA damage in lymphocytes was 2.9 ± 1.4 for 5-OH-dCyd and 4.5 ± 1.8 for 8-oxo-dGuo lesions per 106 dGuo (n = 105). It is unlikely that artifactual oxidation contributed to the observed damage because the level of 5-OH-dCyd was comparable with that of 8-oxo-dGuo in lymphocyte DNA, whereas 8-oxo-dGuo outnumbers 5-OH-dCyd by a ratio of >5:1 when DNA is exposed to various oxidants, including ionizing radiation or Fenton reagents. Rather, the nearly equal levels of 5-OH-dCyd and 8-oxo-dGuo in cellular DNA implies that 8-oxo-dGuo may be more efficiently removed by DNA repair. Finally, and most importantly, the correlation of our endpoints revealed that the naturally occurring level of intracellular antioxidants was negatively correlated to the level of oxidative DNA damage with the strongest correlation observed for glutathione and 8-oxo-dGuo (r = –0.36; P < 0.001). These results strongly suggest that intracellular glutathione and ascorbate protect human lymphocytes against oxidative DNA damage.

Keywords: EC, electrochemical detection; FBS, fetal bovine serum; GC–MS, gas chromatography–mass spectrometry; ODS, octadecylsilyl; 5-OH-dCyd, 5-hydroxy-2′-deoxycytidine; 8-oxo-dGuo, 8-oxo-7,8-dihydro-2′-deoxyguanosine; PBS, phosphate-buffered saline.

Journal Article.  6648 words.  Illustrated.

Subjects: Clinical Cytogenetics and Molecular Genetics

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