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Archives of Toxicology

, Volume 90, Issue 8, pp 1983–1995 | Cite as

Complementarity of phosphorylated histones H2AX and H3 quantification in different cell lines for genotoxicity screening

  • Laure Khoury
  • Daniel Zalko
  • Marc AudebertEmail author
Genotoxicity and Carcinogenicity

Abstract

The in vitro micronucleus assay is broadly used, but is not per se able to discriminate aneugenic from clastogenic compounds, and cytotoxicity can be a confounding factor. In vitro genotoxicity assays generally rely on cell lines with limited metabolic capabilities. Recently, the use of histone H2AX and H3 phosphorylation markers (γH2AX and p-H3) was proposed to discriminate aneugenic from clastogenic chemicals. The aim of the present study was to develop a new genotoxic screening strategy based on the use of the γH2AX and p-H3 biomarkers in combination with cell lines with distinct biotransformation properties. First, we tested a training set of 20 model chemicals comprised of 10 aneugens, five clastogens and five cytotoxics on three human cell lines (HepG2, LS-174T and ACHN). Our data confirm the robustness of these two biomarkers to discriminate efficiently clastogens, aneugens and misleading cytotoxic chemicals in HepG2 cells. Aneugenic compounds induced either an increase or a decrease in p-H3 depending on their mode of action. Clastogens induced γH2AX, and cytotoxic compounds generated a marked decrease in these two biomarkers. Moreover, the use of different cell lines permits to discriminate direct from bioactivated genotoxins without the need of an exogenous metabolic activation system. Finally, we further evaluated this strategy using a test set of 13 chemicals with controversial genotoxic potential. The resulting data demonstrate that the combined analysis of γH2AX and p-H3 is an efficient strategy. Notably, we demonstrated that three compounds (fisetin, hydroquinone and okadaic acid) display both aneugenic and clastogenic properties.

Keywords

Genotoxicity H2AX H3 HepG2 Metabolism 

Notes

Acknowledgments

This work benefited from fruitful discussions with many genotoxicity experts who generously shared their experience, special thanks to Drs. David Kirkland, Richard Walmsley, Steven Bryce and Stephen Dertinger.

Funding

This work was supported in part by the French ANSES MYCO-TOX EST-13-214 and ANR “Contreperf” Grants.

Supplementary material

204_2015_1599_MOESM1_ESM.docx (88 kb)
Supplementary material 1 (DOCX 88 kb)
204_2015_1599_MOESM2_ESM.pptx (638 kb)
Supplementary material 2 (PPTX 637 kb)

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Copyright information

© Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  1. 1.INRA, UMR1331, Toxalim, Research Centre in Food ToxicologyToulouse Cedex 3France
  2. 2.Université de Toulouse, INPT, UPS, UMR1331ToulouseFrance

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