Induction of DNA damage in mammalian cells by hydrogen peroxide generated by glucose oxidase immobilized in agarose slides

A new model system has been developed to study the influence of reactive oxygen species on isolated mammalian cells in conjunction with the comet assay. The glucose-glucose oxidase system was used as a hydrogen peroxide generating source. The level of DNA damage was assessed in the splenocytes and t...

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Bibliographic Details
Published inBiophysics (Oxford) Vol. 62; no. 4; pp. 553 - 556
Main Authors Sirota, N. P., Glukhov, S. I., Sirota, T. V., Mitroshina, I. Yu, Kuznetsova, E. A.
Format Journal Article
LanguageEnglish
Published Moscow Pleiades Publishing 01.07.2017
Springer Nature B.V
Subjects
Biological and Medical Physics
Biophysics
Bone marrow
Comet assay
Controlled conditions
Deoxyribonucleic acid
DNA
DNA damage
DNA repair
Enzymes
Glucose oxidase
Hydrogen peroxide
Leukocytes
Mammalian cells
Molecular Biophysics
Physics
Physics and Astronomy
Reactive oxygen species
Splenocytes
Toxicity
comet assay
DNA damage
hydrogen peroxide
glucose oxidase
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Summary:A new model system has been developed to study the influence of reactive oxygen species on isolated mammalian cells in conjunction with the comet assay. The glucose-glucose oxidase system was used as a hydrogen peroxide generating source. The level of DNA damage was assessed in the splenocytes and the cells of bone marrow of mouse and in human leukocytes both in untreated cells and in cells treated with hydrogen peroxide generated by glucose oxidase using the alkaline comet assay in vitro. Various options for the location of the enzyme in the slides have been studied: in the layer with the cells, in the layer above the cells, or in solution on the surface of the slides. The option where glucose oxidase was in the upper layer of 0.5% agarose over the layer of the cells was optimal. It provided separation of the enzyme from the cells and avoided obstruction to the hydrogen peroxide exposure. For the whole blood study, the content of endogenous glucose must be taken into account. This approach can be used to study the level of DNA damage induced in vitro and for the detection of DNA repair, thereby expanding the possibilities of the method, while the experiments are conducted under controlled conditions.
ISSN:0006-3509
1555-6654
DOI:10.1134/S0006350917040200