Evidence of the in vitro genotoxicity of methyl-pyrazole pesticides in human cells

Abstract

Consumers are exposed daily to several pesticide residues in food, which can be of potential concern for human health. Based on a previous study dealing with exposure of the French population to pesticide residues via the food, we selected 14 pesticides frequently found in foodstuffs, on the basis of their persistence in the environment or their bioaccumulation in the food chain. In a first step, the objective of this study was to investigate if the 14 selected pesticides were potentially cytotoxic and genotoxic. For this purpose, we used a new and sensitive genotoxicity assay (the γH2AX test, involving phosphorylation of histone H2AX) with four human cell lines (ACHN, SH-SY5Y, LS-174T and HepG2), each originating from a potential target tissue of food contaminants (kidney, nervous system, colon, and liver, respectively). Tebufenpyrad was the only compound identified as genotoxic and the effect was only observed in the SH-SY5Y neuroblastoma cell-line. A time-course study showed that DNA damage appeared early after treatment (1h), suggesting that oxidative stress could be responsible for the induction of γH2AX. In a second step, three other pesticides were studied, i.e. bixafen, fenpyroximate and tolfenpyrad, which - like tebufenpad - also had a methyl-pyrazole structure. All these compounds demonstrated genotoxic activity in SH-SY5Y cells at low concentration (nanomolar range). Complementary experiments demonstrated that the same compounds show genotoxicity in a human T-cell leukemia cell line (Jurkat). Moreover, we observed an increased production of reactive oxygen species in Jurkat cells in the presence of the four methyl-pyrazoles. These results demonstrate that tebufenpyrad, bixafen, fenpyroximat and tolfenpyrad induce DNA damage in human cell lines, very likely by a mode of action that involves oxidative stress. Nonetheless, additional in vivo data are required before a definitive conclusion can be drawn regarding hazard prediction to humans.

In Vitro Genotoxicity of Fipronil Sister Chromatid Exchange, Cytokinesis Block Micronucleus Test, and Comet Assay

“…It is indicated that fenproximate tested with various test systems (Ames test, DNA repair test, chromosomal aberration test, unsheduled DNA synthesis test) has no genotoxic potential in studies performed on living systems, including procaryotic and eucaryotic organisms (EFSA, 2010). Graillot et al (2012) demonstrated that bixapen was genotoxic in jurkat cells with the same genotoxic potential as observed in SH-SY%Y cells. Huang et al (2008) and Xia et al (2008) indicated that different cells can show same effects against xenobiotics, including pesticides.…”

“…Graillot et al (2012) observed an increased production of reactive oxygen species (ROS) in jurkat cells in the presence of four methylpyrazoles and demostrated that the four methylpyrazole pesticides (tebufenpyrad, bixafen, fenpyroximat, and tolfenpyrad) induced DNA damage in human cell lines. The relationship between the increase of DNA damage level in comet assay in this study and the increase in ROS level obtained in the study performed by Graillot is redolent of oxidative damage of DNA.…”

Combined Genotoxic Effects of a Polycyclic Aromatic Hydrocarbon (B(a)P) and an Heterocyclic Amine (PhIP) in Relation to Colorectal Carcinogenesis

“…The H2AX In Cell Western technique was performed as previously described with the following modifications [27], [28], [29], [30], [34]. Briefly, cells were dispensed in 96-well cell culture plate (40×10 3 cells, 200 µL/well) and were treated in duplicate, 16 h later, with the model compounds or vehicle (DMSO) in serum free medium.…”

“…Raw absorbance data was averaged for the duplicate, corrected for background; the relative fluorescence units from the scanning allowed a quantitative analysis. The ICW technique allows the determination of cytotoxicity and genotoxicity in a single experiment [27], [28], [30], [34]. For determination of genotoxicity, relative fluorescent units for γH2AX per cell (as determined by γH2AX divided by DNA content) were divided by the respective controls (vehicle only), in order to determine the change in phosphorylation of H2AX levels relative to control.…”

“…In order to evaluate the genotoxic potential of these two compounds, alone or in mixture on the two cell lines, we quantified B( a )P and PhIP derived DNA adducts by HPLC-MS/MS. Moreover, we used a genotoxic assay based on the quantification of the phosphorylation of the histone H2AX (named γH2AX) that reflects a global genotoxic insult resulting from diverse types of DNA damages, notably DNA adducts and oxidative DNA lesions [27], [28], [29], [30], [31], [32], [33], [34].…”