The Induction and Repair of Double-Strand DNA Breaks in Mammalian Cells as Detected by Neutral Elution

  • Govert P. van der Schans


One of the important types of damage induced by ionizing radiation in intracellular DNA is the double-strand break (dsb). This damage, involving disruption of both strands of the DNA-helix at the same or neighbouring sites, is produced at a much lower frequency than other lesions (single-strand breaks, nucleotide damage). One would expect, however, that double-strand breaks would be much more deleterious for the cell than lesions of the latter type as these probably are repaired more easily.

The neutral filter elution technique has proven to be a sensitive means of detecting DNA dsb after exposure to low doses of ionizing radiation (Bradley and Kohn, 1979). However, the results obtained with it reported in literature are in some cases controversial as well as in disagreement with results of other methods. For example, according to data of Radford (1985) and Radford and Hodgson (1985), the induction of dsb by 250 kV X-rays does not show a simple linear dose-effect relationship, whereas others (Ross and Bradley, 1981, Van der Schans et al. 1982a, and Woods et al. 1982) found data consistent with a linear relationship. Also Blöcher (1982) observed a linear relationship in his sedimentation studies.

Another discrepancy arose when repair of dsb was studied with the elution technique; half-lifes of about 10 min were found, in contrast to the earlier values of 1–2 h obtained in sedimentation experiments. It has been suggested that the fast-repair component seen in filter elution studies might represent repair of DNA single-strand breaks (ssb). This is unlikely since Bradley and Kohn (1979) have found that the different dsb/ssb ratios determined with filter elution for various agents are comparable with those found with other methods. The fact that most of the filter elutions are carried out at rather high pH, could be ruled out as a possible reason for this discrepancy, since we found that elution at neutral pH leads to the same results.

In this paper the reliability of the different methods for the detection of dsb induced by ionizing radiation and other agents will be discussed.


Strand Break Chinese Hamster Ovary Cell Elution Procedure Alkaline Elution Fast Repair 
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  1. Blöcher, D., 1982, DNA double strand breaks in Ehrlich ascites tumour cells at low doses of X-rays. I. Determination of induced breaks by centrifugation at reduced speed. Int. J. Radiat. Biol., 42, 317–328.Google Scholar
  2. Blöcher, D., Nüsse, M. and Bryant, P.E., 1983, Kinetics of double strand break repair in the DNA of X-irradiated synchronized mammalian cells. Int. J. Radiat. Biol., 43, 579–584.Google Scholar
  3. Boye, E., 1980, Formation and repair of DNA double-strand breaks in superinfecting phage X after ionizing irradiation of Escherichia coli host cells. Radiation Research, 81, 427–440.PubMedCrossRefGoogle Scholar
  4. Bradley, M.O. and K.W. Kohn, 1979, X-ray induced DNA double strand break production and repair in mammalian cells as measured by neutral filter elution. Nucl. Acids Res. 7, 793–804.Google Scholar
  5. Bryant, P.E., and Blöcher, D, 1980, Measurement of the kinetics of DNA double strand break repair in Ascites tumour cells using the unwinding method. Int. J. Radiat. Biol., 38, 335–347.Google Scholar
  6. Burgi, E. and Hershey, A.D., 1963, Sedimentation rate as a measure of molecular weight of DNA. Biophysical J. 3, 309–321.CrossRefGoogle Scholar
  7. Cole, A., Shonka, F., Corry, P.,and Grant Cooper, W., 1975, CHO cell repair of single-strand and double-strand DNA breaks induced by K- and a-radiation. Molecular Mechanisms for the repair of DNA, edited by R.B. Setlow and P.C. Hanawalt (New York Plenum Press), pp. 665–676.Google Scholar
  8. Coquerelle, T., Bopp, A., Kessler, B., and Hagen, U., 1973, Strand breaks and 5’ end-groups in DNA of irradiated thymocytes. Int. J. Radiat. Biol., 24, 397–404.Google Scholar
  9. Dugle, D.L., Gillespie, C.J. and Chapman, J.D., 1976, DNA strand breaks, repair, and survival in X-irradiated mammalian cells. Proc. Natl. Acad. Sci. USA, 73, 809–812.Google Scholar
  10. Grdina, D.J., Guilford, W.H., Sidgestad, C.P. and Giometti, C.S., 1987, Role of radioprotectors in DNA damage and repair, damage to protein, and effects on cell progression. In: Book of “Program and Abstracts” of Symposium on “Perspectives in Radioprotection” at Bethesda, Maryland, USA, March 1987, p. 35–36.Google Scholar
  11. Jeggo, P.A., and Kemp, L.M., 1983, X-ray sensitive mutants of Chinese hamster ovary cell line: Isolation and cross-sensitivity to other DNA-damaging agents. Mutation Res., 122, 313–327.Google Scholar
  12. Kampf, G., Tolkendorf, E., Regel, K., and Abel, H., 1977, Cell inactivation and DNA strand break rates after irradiation with X-rays and fast neutrons. Studia biophysica, 62, 17–24.Google Scholar
  13. Kemp, L.M., Sedgwick, S.G. and Jeggo, P.A., 1984, X-ray sensitive mutants of Chinese hamster ovary cells defective in double-strand break rejoining. Mutation Res. 132, 189–196.PubMedCrossRefGoogle Scholar
  14. Lafleur, M.V.M., Van Heuvel, M., Van der Stroom, H.A. and Loman, H., 1976, Biological relevance of gamma-ray-induced alkali-labile sites in single-stranded DNA in aqueous solutions. Int. J. Radiat. Biol., 30, 223–228.Google Scholar
  15. Lehmann, A.R. and Ormerod, M.G., 1970, Double-strand breaks in the DNA of a mammalian cell after X-irradiation. Biochim. biophys. Acta, 217, 268–277.Google Scholar
  16. Lehmann, A.R. and Stevens, S., 1977, The production and repair of double strand breaks in cells from normal humans and from patients with ataxia telangiectasia. Biochim. biophys. Acta, 474, 49–60.Google Scholar
  17. Radford, I.R., 1985, The level of induced DNA double-strand breakage correlates with cell killing after X-irradiation. Int. J. Radiat. Biol., 48, 45–54.Google Scholar
  18. Radford, I.R. and Hodgson, G.S., 1985, 125I-induced DNA double strand breaks: use in calibration of the neutral filter elution technique and comparison with X-ray induced breaks. Int. J. Radiat. Biol., 48, 555–566.Google Scholar
  19. Ross, W.E., and M.O. Bradley, 1981, DNA double-strand breaks in mammalian cells after exposure to intercalating agents. Biochim. biophys. Acta, 654, 129–134.Google Scholar
  20. Shiloh, Y., Van der Schans, G.P. Lohman, P.H.M., and Becker, Y., 1983, Induction and repair of DNA damage in normal and ataxia-telangiectasia skin fibroblasts treated with neocarzinostatin. Carcinogenesis, 4, 917–921.PubMedCrossRefGoogle Scholar
  21. Van der Schans, G.P., 1978, Gamma-ray induced double-strand breaks in DNA resulting from randomly-inflicted single-strand breaks: Temporal local denaturation, a new radiation phenomenom? Int. J. Radiat. Biol., 33, 105–120.Google Scholar
  22. Van der Schans, G.P., Aten, J.B.T. and J. Blok, 1969, Determination of molecular weight distributions of DNA by means of sedimentation in a sucrose gradient. Anal. Biochem., 32, 14–30.Google Scholar
  23. Van der Schans, G.P. Centen, H.B. and Lohman, P.H.M., 1979, The induction of gamma-endonuclease-susceptible sites by (-rays in CHO cells and their cellular repair are not affected by the presence of thiol compounds during irradiation. Mutation Res., 59, 119–122.PubMedCrossRefGoogle Scholar
  24. Van der Schans, G.P., Centen, H.B., and Lohman, P.H.M., 1980, Studies on the repair defect(s) of ataxia-telangiectasia fibroblasts. Radiat. envir. Biophys., 17, 351.Google Scholar
  25. Van der Schans, G.P, Centen, H.B. and Lohman, P.H.M., 1982a, DNA lesions induced by ionizing radiation. Prog. Mutation Res., 4, 285–299.Google Scholar
  26. Van der Schans, G.P., Centen, H.B. and Lohman, P.H.M., 1982b, The induction and repair of double-strand DNA breaks in normal human and ataxia-telangiectasia cells exposed to 60Co- I -radiation, 4-nitroquinoline-1-oxide or bleomycin. In “A cellular and molecular link between Cancer, Neuropathology and Immune Deficiency. Eds.: Bridges and Harnden, Wiley and sons Ltd. Chichester, p 291–303.Google Scholar
  27. Van der Schans, G.P, Paterson, M.C. and Cross, W.G., 1983, DNA strand break and rejoining in cultured human fibroblasts exposed to fast neutrons or gamma rays. Int. J. Radiat. Biol., 44, 75–85.Google Scholar
  28. Van der Schans, G.P. Vos, O., Roos-Verheij, W.S.D. and Lohman, P.H.M., 1986, The influence of oxygen on the induction of radiation damage in DNA in mammalian cells after sensitization by intracellular glutathione depletion. Int. J. Radiat. Biol., 50, 453–465.Google Scholar
  29. Veatch, W. and Okada, S., 1969, Radiation-induced breaks of DNA in cultured mammalian cells. Biophysical J., 9, 330–346.CrossRefGoogle Scholar
  30. Ward, J.F., 1981, Some biochemical consequences of the spatial distribution of ionizing radiation-produced free radicals. Radiation Res., 86, 185–195.PubMedCrossRefGoogle Scholar
  31. Waters, R., Mirzay Ans, R., Meredith, J., Mallalah, G., Danford, N. and Parry, J.M., 1982, Correlations in mammalian cells between types of DNA damage, rates of DNA repair and the biological consequences. Prog. Mutation Res., 4, 247–259.Google Scholar
  32. Weibezahn, K.F., Lohrer, H. and Herrlich, P., 1985, Double-strand break repair and G2 block in Chinese hamster ovary cells and their radiosensitive mutants. Mutation Res. 145, 177–183.PubMedCrossRefGoogle Scholar
  33. Woods, W.G., Lopez, M. and Kalvonjian, S.L., 1982, Normal repair of gamma radiation-induced single-strand and double-strand DNA breaks in retinoblastoma fibroblasts. Biochim. biophys. Acta, 698, 40–48.Google Scholar

Copyright information

© Springer Science+Business Media New York 1989

Authors and Affiliations

  • Govert P. van der Schans
    • 1
  1. 1.TNO Medical Biological LaboratoryRijswijkThe Netherlands

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