The Kinetics of Repair of Oxidative DNA-Damage (Strand Breaks and Oxidized Pyrimidines) in Human Cells

A R Collins, A G Ma, Susan Joyce Duthie

    Research output: Contribution to journalArticle

    574 Citations (Scopus)

    Abstract

    Single cell gel electrophoresis is a sensitive method for detecting DNA strand breaks. Cells embedded in agarose are converted to nucleoids by treating with detergent and high salt. DNA breaks render the nucleoid DNA susceptible to extension by electrophoresis, forming 'comets'. We find that when DNA breakage resulting from H2O2 treatment is examined, freshly isolated normal human lymphocytes are relatively resistant compared with transformed human cells. When incubated after treatment with H2O2, HeLa cells repair most strand breaks within 1 h, and a substantial fraction of the oxidised pyrimidines (detected by converting them to DNA breaks with endonuclease III) within 4 h. However, lymphocytes are less proficient at repair; during incubation for 4 h after treatment with H2O2, no detectable removal of endonuclease III-sensitive sites is seen. While the addition of deoxyribonucleosides promotes completion of repair of UV damage by lymphocytes, it has no significant effect on repair of oxidative damage.

    Original languageEnglish
    Pages (from-to)69-77
    Number of pages9
    JournalMutation Research. DNA Repair
    Volume336
    Issue number1
    DOIs
    Publication statusPublished - Jan 1995

    Keywords

    • OXIDIZED BASES
    • STRAND BREAKS
    • COMET ASSAY
    • HUMAN CELLS
    • DEOXYRIBONUCLEOTIDE POOL
    • HUMAN-LYMPHOCYTES
    • IONIZING-RADIATION
    • BASE DAMAGE
    • CONTAIN
    • Oxidised bases
    • Strand breaks
    • Comet assay
    • Human cells
    • Deoxyribonucleotide pool

    Cite this

    The Kinetics of Repair of Oxidative DNA-Damage (Strand Breaks and Oxidized Pyrimidines) in Human Cells. / Collins, A R ; Ma, A G ; Duthie, Susan Joyce.

    In: Mutation Research. DNA Repair, Vol. 336, No. 1, 01.1995, p. 69-77.

    Research output: Contribution to journalArticle

    Collins, A R ; Ma, A G ; Duthie, Susan Joyce. / The Kinetics of Repair of Oxidative DNA-Damage (Strand Breaks and Oxidized Pyrimidines) in Human Cells. In: Mutation Research. DNA Repair. 1995 ; Vol. 336, No. 1. pp. 69-77.
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    abstract = "Single cell gel electrophoresis is a sensitive method for detecting DNA strand breaks. Cells embedded in agarose are converted to nucleoids by treating with detergent and high salt. DNA breaks render the nucleoid DNA susceptible to extension by electrophoresis, forming 'comets'. We find that when DNA breakage resulting from H2O2 treatment is examined, freshly isolated normal human lymphocytes are relatively resistant compared with transformed human cells. When incubated after treatment with H2O2, HeLa cells repair most strand breaks within 1 h, and a substantial fraction of the oxidised pyrimidines (detected by converting them to DNA breaks with endonuclease III) within 4 h. However, lymphocytes are less proficient at repair; during incubation for 4 h after treatment with H2O2, no detectable removal of endonuclease III-sensitive sites is seen. While the addition of deoxyribonucleosides promotes completion of repair of UV damage by lymphocytes, it has no significant effect on repair of oxidative damage.",
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    N2 - Single cell gel electrophoresis is a sensitive method for detecting DNA strand breaks. Cells embedded in agarose are converted to nucleoids by treating with detergent and high salt. DNA breaks render the nucleoid DNA susceptible to extension by electrophoresis, forming 'comets'. We find that when DNA breakage resulting from H2O2 treatment is examined, freshly isolated normal human lymphocytes are relatively resistant compared with transformed human cells. When incubated after treatment with H2O2, HeLa cells repair most strand breaks within 1 h, and a substantial fraction of the oxidised pyrimidines (detected by converting them to DNA breaks with endonuclease III) within 4 h. However, lymphocytes are less proficient at repair; during incubation for 4 h after treatment with H2O2, no detectable removal of endonuclease III-sensitive sites is seen. While the addition of deoxyribonucleosides promotes completion of repair of UV damage by lymphocytes, it has no significant effect on repair of oxidative damage.

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    KW - BASE DAMAGE

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    KW - Strand breaks

    KW - Comet assay

    KW - Human cells

    KW - Deoxyribonucleotide pool

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