In Vitro Comparison of the Potential DNA Damage Effects of H2O2 and Etoposide for 30 and 60 Minutes Exposures With The Comet Assay


İpek Tekneci S., Üstündağ A., Duydu Y.

11th INTERNATIONAL CONGRESS OF THE TURKISH SOCIETY OF TOXICOLOGY, Antalya, Turkey, 02 November 2022, pp.121-122

  • Publication Type: Conference Paper / Summary Text
  • City: Antalya
  • Country: Turkey
  • Page Numbers: pp.121-122
  • Ankara University Affiliated: Yes

Abstract

The comet assay providing the estimation of both DNA damage and repair is a fast, sensitive, and

affordable technique. For long years, DNA damage and repair in individual cells and tissues have

been evaluated by using the comet assay. Also, the comet assay is the standard method used in

ecogenotoxicology, molecular epidemiology, human and environmental biomonitoring. Single/

double DNA strand breaks, alkali labile sites, base-pair damages, and apoptotic nuclei can be

determined by using this method.

Positive controls are chemicals known to induce DNA damage by the way of direct or indirect, and

also they cause single/double DNA strand breaks identified by the comet assay. The examples of

positive controls used in the comet assay are hydrogen peroxide (H2O2), methyl methane sulfonate

(MMS), ethyl methane sulfonate (EMS), etoposide, and ethyl nitroso urea (ENU). The choice of a

suitable positive control is an essential part of genotoxicity tests. However, it has been stated that

the results obtained from different positive controls at discrete concentrations and time intervals

varied. Accordingly, we aimed to compare H2O2, and etoposide, at different concentrations for 30

minutes, and 1h, respectively.

In our study, the alkali comet assay (pH>13) was performed to compare the results obtained from

two different positive controls by using 3T3 cell lines. H2O2 was tested at 1, 5, 10, 25, 50, 75, 100

and 200 μM for 1h. For 30 minutes, up to 75 μM H2O2 concentrations were tested. As for etoposide,

six different concentrations (0,05, 0,1, 0,5, 1, 5, 10 μM) were used for 30 minutes and 1h.

According to the comet assay results, the exposure of 25 and 50 μM H2O2 for 30 minutes

significantly induced DNA damage more than for 60 minutes. DNA strand breaks resulted from

the exposure of 1, 5, and 10 μM etoposide concentrations for 30 minutes were higher than for 60

minutes (p<0,05, one-way ANOVA).