Electrochemical Detection of 8-Hydroxy-2-Deoxyguanosine Levels in Cellular DNA

  • Thomas M. Nicotera
  • Sofia Bardin
Part of the Methods in Molecular Biology™ book series (MIMB, volume 108)


Oxidative stress has been implicated in the etiology of many pathological states and known to result in DNA damage. Oxidative DNA damage can lead to mutagenesis (1, 2, 3) and has been associated with aging (4), diabetes mellitus (5), inflammatory disease (1) and carcinogenesis (1, 2, 3,6). The 8-hydroxy-2-deoxyguanosine lesion (8-OHdG) is often used in the assessment of oxidative DNA damage and has become the de facto marker for oxidative damage to DNA. 8-OHdG is one of the most prominent lesions observed following exposure to ionizing radiation (7) but also results from treatment with many xenobiotics (8) as well as by endogenous mechanisms. Thus, normal endogenous levels becomes a critical issue in the assessment of cellular 8-OHdG levels in pathological states. Furthermore, 8-OHdG is efficiently repaired by a DNA glycosylase specific for this lesion (9) and its contribution to mutagenesis is relatively weak (10, 11, 12). However, it is representative of approximately 20 additional oxidatively-denved lesions known to result from radiation damage (13) and whose mutagenic outcome are largely unknown in mammalian-cell systems.


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  1. 1.
    Ames, B. N (1983) Dietary carcinogens and anticarcinigens. Science 221, 1256–1264.PubMedCrossRefGoogle Scholar
  2. 2.
    Cerutti, P. A. (1985) Prooxidant states and tumor promotion Science 225, 375–381.CrossRefGoogle Scholar
  3. 3.
    Breimer, L. H. (1990) Molecular mechanisms of oxygen radical carcinogenesis and mutagenesis: the role of DNA base damage. Mol Carcinogenesis 3, 188–197CrossRefGoogle Scholar
  4. 4.
    Fraga, C. G., Shigenaga, M. K., Park, J-W, Deagn, P., and Ames, B. N. (1990) Oxidative damage to DNA during aging: 8-hydroxy-2′-deoxyguanosine in rat organ DNA and urine. Proc Natl. Acad Sci. USA 87, 4533–4537PubMedCrossRefGoogle Scholar
  5. 5.
    Dandona, P., Thusu, K., Cook, S., Snyder, B., and Nicotera, T (1996) Oxidative DNA damage diabetes mellitus. Lancet 347, 444–445.PubMedCrossRefGoogle Scholar
  6. 6.
    Floyd, R A. (1990) The role of 8-hydroxyguanosine in carcinogenesis Carcinogenesis 11, 1447–1450.PubMedCrossRefGoogle Scholar
  7. 7.
    Kasai, H., Crain, P. F., Kuchino, Y., Nishimura, S., Ootsuyama, A., and Tanooka, H (1986) Formation of 8-hydrodroxyguanine in cellular DNA by agents producing oxygen radicals and evidence for its repair. Carcinogenesis 8, 1849–1851.CrossRefGoogle Scholar
  8. 8.
    Fiala, E. S., Conaway, C. C., and Mathis, J. E. (1989) Oxidative DNA and RNA damage in the livers of Sprague-Dawley rats treated with the hepatocarcinigen 2-nitropropane. Cancer Res. 49, 5518–5522.PubMedGoogle Scholar
  9. 9.
    Chung, M. H., Kasai, H., Jones, D. S., Inoue, H., Ishikawa, H., Ohtsuka E., and Nishimura, S. (1991) An endonuclease activity of Escherichia Coll that specifically removes 8-hydroxyguamne residues from DNA. Mutat. Res. 254, 1–12.PubMedGoogle Scholar
  10. 10.
    Kuchino, T., Mori, F., Kasai, H., Inoue, H., Iwai, S., Miura, K., Ohtsuka E, and Nishimura, S. (1987) Misreading of DNA templates containing 8-hydroxydeoxuguanosine at the modified base and at adjacent residues Nature (Lond.) 327, 77–79CrossRefGoogle Scholar
  11. 11.
    Shibutani, S, Takeshita, M, and Grollman, A. P (1991) Insertion of specific bases during DNA sysnthesis past the oxidation-damaged base 8-oxodG. Nature (Lond.) 349, 431–434CrossRefGoogle Scholar
  12. 12.
    Wood, M. L., Dizdaroglu, M., Gajewski, E., and Essigmann, J. M. (1990) Mechanistic studies of ionizing radiation and oxidative mutagenesis, genetic effects of a single 8-hydroxyguanine (7-hydro-8-oxoguanine) residue inserted at a unique site in a viral genome. Biochem. 29, 7024–7032.CrossRefGoogle Scholar
  13. 13.
    Cadet, J. (1994) DNA repair caused by oxidation, deamination, ultraviolet radiation and photoexcited psoralens in DNA adducts, in Identification and Biological Significance, (Hemminki, K., Dipple, A., Shuker, D. E. P., Kadlubar, F. F., Segerback, D., and Bartsch, H., eds.), IARC Scientific Publication No. 125. International Agency for Research on Cancer, Lyon, pp. 245–276.Google Scholar
  14. 14.
    Roy, D., Floyd, R. A., and Liehr, J. (1991) Elevated 8-hydroxyguanosine levels in DNA of diethylstilbesterol-treated Syrian hamsters: covalent DNA damage by free radicals generated by redox cycling of ditheylstilbesterol. Cancer Res. 51, 3882–3885.PubMedGoogle Scholar
  15. 15.
    Wei, H. and Frenkel, K. (1991) In vivo formation of oxidized DNA bases in tumor promoter-treated mouse skin. Cancer Res. 51, 4113–4449.Google Scholar
  16. 16.
    Malins, D. and Haimanot, R. (1991) Major alterations in the nucleotide structure of DNA in cancer of the female breast. Cancer Res. 51, 5430–5432.PubMedGoogle Scholar
  17. 17.
    Olinski, R., Zastawny, T., Budzbon, J., Skowkowski, J., Zegarski, W., and Dizdaroglu, M. (1992) DNA base modifications in chromatin of human cancerous tissues. FEBS 309, 193–198.CrossRefGoogle Scholar
  18. 18.
    Maccubbin, A. E., Przybyszewski, J., Evans, M., Budzinski, E. E., Patryzc, H. B., Kulesz-Martin, M., and Box, H. B. (1995) DNA damage in UVB-irradiated keratinicytes. Carcinogenesis 16, 1659–1660.PubMedCrossRefGoogle Scholar
  19. 19.
    Frenkel, K., Zhong, Z., Wei, H., Karkoszka, J., Patel, U., Rashid, K., Georgescu, M., and Solomon, J. J. (1991) Quantitative high-performance chromatography analysis of DNA oxidized in vitro and in vivo. Anal. Biochem. 196, 126–136.PubMedCrossRefGoogle Scholar
  20. 20.
    Degan, P., Shinegawa, M. K., Park, E-M., Alperin, P. R, and Ames, B. N. (1991) Immunoaffinity isolation of urinary 8-hydroxy-2′-deoxyguanosine and 8-hydroxyguanine and quantitation of 8-hydroxy-2′-deoxyguanosine in DNA by polyclonal antibodies. Carcinogenesis 12, 865–871.PubMedCrossRefGoogle Scholar
  21. 21.
    Park, J-U., Cundy, K. C., and Ames, B. N. (1989) Detection of DNA adducts by high performance liquid chromatography with electrochemical detection. Carcinogenesis 10, 827–832.PubMedGoogle Scholar
  22. 22.
    Paul, C. R., Wallace, J. C., Alderfer, J. A., and Box, H. C. (1988) Radiation chemistry of d(TpApCpG) in oxygenated solution. Int. J. Radiat. Biol 54, 403–115.PubMedCrossRefGoogle Scholar
  23. 23.
    Marmur, J. (1961) A procedure for the isolation of deoxyribonucleic acid from micro-organisms. J. Mol. Biol. 3, 208–218.CrossRefGoogle Scholar

Copyright information

© Humana Press Inc., Totowa, NJ 1998

Authors and Affiliations

  • Thomas M. Nicotera
    • 1
    • 2
  • Sofia Bardin
    • 1
  1. 1.Department of Biophysics, Roswell Park Cancer InstituteNew York State Department of HealthBuffalo
  2. 2.New York State Department of HealthUniversity at Buffalo School of Medicine and Biomedical SciencesBuffalo

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