Detection of xenobiotic-induced DNA damage by the comet assay applied to human and rat precision-cut liver slices

Plazar, Janja; Hreljac, Irena; Pirih, Primož; Filipič, Metka; Groothuis, Geny M. M.

doi:10.1016/j.tiv.2007.02.012

Cited by 23 publications

(15 citation statements)

References 37 publications

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“…Moreover, this assay was found to be more sensitive than the comet assay when we compare our results regarding PAHs genotoxicity with published data on the same cell line (Plazar et al, 2007;Winter et al, 2008;Tarantini et al, 2009). Concerning the comparison of the γ-H2AX assay with the micronucleus test, it is now well established that the micronucleus formation correlates well with H2AX phosphorylation (Medvedeva et al, 2007;Terradas et al, 2009;Yoshikawa et al, 2009).…”

Section: Genotoxicity Of Pahs In Human Cell Linessupporting

confidence: 58%

Comparative potency approach based on H2AX assay for estimating the genotoxicity of polycyclic aromatic hydrocarbons

Audebert

Zeman

Beaudoin

et al. 2012

Toxicology and Applied Pharmacology

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a b s t r a c t a r t i c l e i n f oPolycyclic Aromatic Hydrocarbons (PAHs) constitute a family of over one hundred compounds and can generally be found in complex mixtures. PAHs metabolites cause DNA damage which can lead to the development of carcinogenesis. Toxicity assessment of PAH complex mixtures is currently expressed in terms of toxic equivalents, based on Toxicity Equivalent Factors (TEFs). However, the definition of new TEFs for a large number of PAH could overcome some limitations of the current method and improve cancer risk assessment. The current investigation aimed at deriving the relative potency factors of PAHs, based on their genotoxic effect measured in vitro and analyzed with mathematical models. For this purpose, we used a new genotoxic assay (γH2AX) with two human cell lines (HepG2 and LS-174T) to analyze the genotoxic properties of 13 selected PAHs at low doses after 24 h treatment. The dose-response for genotoxic effects was modeled with a Hill model; equivalency between PAHs at low dose was assessed by applying constraints to the model parameters. In the two cell lines tested, we observed a clear dose-response for genotoxic effects for 11 tested compounds. LS-174T was on average ten times more sensitive than HepG2 towards PAHs regarding genotoxicity. We developed new TEFs, which we named Genotoxic Equivalent Factor (GEF). Calculated GEF for the tested PAHs were generally higher than the TEF usually used. Our study proposed a new in vitro based method for the establishment of relevant TEFs for PAHs to improve cancer risk assessment.

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Section: Genotoxicity Of Pahs In Human Cell Linessupporting

confidence: 58%

Comparative potency approach based on H2AX assay for estimating the genotoxicity of polycyclic aromatic hydrocarbons

Audebert

Zeman

Beaudoin

et al. 2012

Toxicology and Applied Pharmacology

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“…1, Table 1). The extent of DNA damage is comparable to that observed in metabolically active rat and human liver slices and HepG2 cells (Plazar et al, 2007;Straser et al, 2011). In zebrafish embryos Kosmehl et al…”

Section: Resultssupporting

confidence: 60%

“…In the comet assay, the measured % DNA damage is not normally distributed (Plazar et al 2007), therefore we analyzed and presented the data in two separate ways. 1) The values of % DNA in the comet tail from all embryos in the treatment group were pooled (n = 330), and the differences in % of DNA in the tail between the treatments were analyzed with nonparametric one-way ANOVA (Kruskal-Wallis test) with Dunn's post test.…”

Section: Discussionmentioning

confidence: 99%

“…The embryos were exposed to MMS (2.5 and 25 mgL -1 = 22.7 and 227 µM) and BaP (0.63 and 6.3 mgL -1 = 2.5 and 25 µM) as model genotoxic chemicals, and NaCl (54.4 mgL -1 = 1mM) as a nongenotoxic chemical. The concentrations of MMS and BaP were selected according to their genotoxic potencies in different in vivo and in vitro test systems (Rank and Jensen, 2003;Kosmehl et al, 2006;Plazar et al, 2007). No embryo mortality was observed at the selected concentrations of the tested chemicals.…”

Section: Experimental Designmentioning

confidence: 99%

“…For preparation of the slides with separated single cells, we used the technique of mechanical squashing on the slides, previously described for the preparation of single cells for the comet assay from tissue slices (Plazar et al, 2007). The slides were precoated with 0,5% NMP agarose, and then covered with a bottom layer of 1 % NMP agarose (80 µL).…”

Section: Optimization Of the Zebrafish Embryo Mounting And Comet Assaymentioning

confidence: 99%

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A method for the assessment of DNA damage in individual, one day old, zebrafish embryo (Danio rerio), without prior cell isolation

Eleršek

Plazar

Filipič

2013

Toxicology in Vitro

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Genotoxic effects of p,p′‐DDT (1,1,1‐trichloro‐2,2‐bis‐(chlorophenyl)ethane) and its metabolites in Zebra mussel (D. polymorpha) by SCGE assay and micronucleus test

Binelli

Riva

Cogni

et al. 2008

Environ and Mol Mutagen

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This is the first study to evaluate the potential genotoxicity of p,p'-DDT (1,1,1-trichloro-2,2-bis-(chlorophenyl)ethane) and its metabolites (p,p'-DDD and p,p'-DDE) in the sentinel mollusc Zebra mussel (Dreissena polymorpha). DNA damage was measured using the single cell gel electrophoresis (SCGE) assay and the micronucleus test (MN test), which represent two of the more sensitive biomarkers for genotoxicity evaluation. Three different concentrations (0.1, 2, and 10 mug/L) of each compound were administered in water for 168 hr, maintaining mussels at constant laboratory conditions and collecting several specimens every 48 hr for biochemical analyses. At the same time, the bioaccumulation process and the concentration/effect relationship were checked by GC-MS/MS analyses of mussel soft tissues. The SCGE assay results showed a clear and significant (P < 0.05) relationship between DNA injuries and tested doses for all the homologues throughout the 7-day exposure period. The final DNA damage due to p,p'-DDE was almost double that of the other two homologues that showed the same toxicity pattern. The micronucleus frequency analysis confirmed the genotoxicity potential of the three homologues and p,p'-DDE showed the highest irreversible DNA damage. The capability of Zebra mussels to biotransform the administered compound in the other homologues was demonstrated by multiple regression analyses carried out between the MN frequencies and the concentrations of the different homologues in the mussel soft tissues. A greater genotoxic potential of the p,p'-DDE with respect to the other two chemicals was revealed.

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Detection of xenobiotic-induced DNA damage by the comet assay applied to human and rat precision-cut liver slices

Cited by 23 publications

References 37 publications

Comparative potency approach based on H2AX assay for estimating the genotoxicity of polycyclic aromatic hydrocarbons

Comparative potency approach based on H2AX assay for estimating the genotoxicity of polycyclic aromatic hydrocarbons

A method for the assessment of DNA damage in individual, one day old, zebrafish embryo (Danio rerio), without prior cell isolation

Genotoxic effects of p,p′‐DDT (1,1,1‐trichloro‐2,2‐bis‐(chlorophenyl)ethane) and its metabolites in Zebra mussel (D. polymorpha) by SCGE assay and micronucleus test

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