In Vitro Genotoxicity Evaluation of PAHs in Mixtures Using Experimental Design

International audienceSettled dusts are sinks for environmental pollutants, including Polycyclic Aromatic Hydrocarbons (PAHs) that are ubiquitous, persistent, and carcinogenic. To assess their toxicity in mixtures, Toxic Equivalent Factors (TEFs) are routinely used and based on the hypothesis of additive effects, although PAH interactions may occur and remain an open issue. This study investigated genotoxic binary interaction effects for six PAHs in mixtures using two in vitro assays and estimated Genotoxic Equivalent Factors (GEFs) to roughly predict the genotoxicity of PAH in mixtures. The Design of the Experiment approach was used with the micronucleus assay for cytostasis and micronuclei frequency and the alkaline comet assay for DNA damage. GEFs were determined for each PAH independently and in a mixture. For the cytostasis endpoint, no PAHs interaction was noted. BbF and BaP had a synergistic effect on DNA damage. All the PAH interacted between them regarding chromosomal damage. Although the calculated GEFs were similar to the TEFs, the latter may underestimate the genotoxic potential of a PAH mixture. GEFs calculated for PAH alone were lower than GEFs for PAHs in mixtures; thus, mixtures induce greater DNA/chromosomal damage than expected. This research helps to advance the challenging issue of contaminant mixtures’ effects on human health

Toward an interdisciplinary approach to assess the adverse health effects of dust-containing polycyclic aromatic hydrocarbons (PAHs) and metal(loid)s on preschool children

International audienceSettled dust can function as a pollutant sink for compounds, such as polycyclic aromatic hydrocarbons (PAHs) and metal(loid)s (MMs), which may lead to health issues. Thus, dust represents a hazard specifically for young children, because of their vulnerability and hand-to-mouth behavior favoring dust ingestion.The aim of the present study was to explore the influence of the season and the microenvironment on the concentrations of 15 PAHs and 17 MMs in indoor and outdoor settled dust in three preschools (suburban, urban, and industrial). Second, the potential sources and health risks among children associated with dust PAHs and MMs were assessed. Third, domestic factors (risk perception, knowledge and parental style) were described to explore protective parental behaviors toward dust hazards.The suburban preschool had the lowest concentrations of dust PAHs and MMs, while the industrial and urban preschools had higher but similar concentrations. Seasonal tendencies were not clearly observed. Indoor dusts reflected the outdoor environment, even if specific indoor sources were noted. Source analysis indicated mainly vehicular emissions, material release, and pyrogenic or industrial sources. The non-cancer health risks were nonexistent, but potential cancer health risks (between 1.10(-6) and 1.10(-4)) occurred at all sampling locations. Notably, the highest cancer risk was observed in a playground area (>1.10(-4)) and material release should be further addressed. Whereas we assessed higher risk indoors, parents perceived a higher risk in the open-air environment and at the preschool than at home. They also perceived a lower risk for their own children, revealing an optimism bias, which reduces parental anxiety

Potentially harmful elements in house dust from Estarreja, Portugal: characterization and genotoxicity of the bioaccessible fraction

International audienceDue to their behavioral characteristics, young children are vulnerable to the ingestion of indoor dust, often contaminated with chemicals that are potentially harmful. Exposure to potentially harmful elements (PHEs) is currently exacerbated by their widespread use in several industrial, agricultural, domestic and technological applications. PHEs cause adverse health effects on immune and nervous systems and can lead to cancer development via genotoxic mechanisms. The present study is an integrated approach that aims at assessing the genotoxicity of bioaccessible PHEs following ingestion of contaminated house dust. A multidisciplinary methodology associating chemical characterization of five house dust samples, extraction of the bioaccessible PHEs in gastric extracts by the unified BARGE method, determination of the bioaccessible fraction and in vitro genotoxicity of gastric extracts in adenocarcinoma gastric human (AGS) cells was developed. The five gastric extracts induced dose-dependent genotoxicity in AGS cells. Copper (bioaccessible concentration up to 111 mg/kg) was probably the prevalent PHE inducing primary DNA damage (up to 5.1-fold increase in tail DNA at 0.53 g/l of gastric extract). Lead (bioaccessible concentration up to 245 mg/kg) was the most prevalent PHE inducing chromosome-damaging effects (r = 0.55; p \ 0.001 for micronucleated cells induction). The association of principal component analysis and Spearman's correlations was decisive to understand the chromosome-damaging properties of the bioacces-sible PHEs in AGS cells. This methodology could be used on a larger-scale study to provide useful information for science-based decision-making in regulatory policies, and a better estimation of human exposure and associated health risks

Potentially harmful elements in house dust from Estarreja, Portugal: characterization and genotoxicity of the bioaccessible fraction

International audienceDue to their behavioral characteristics, young children are vulnerable to the ingestion of indoor dust, often contaminated with chemicals that are potentially harmful. Exposure to potentially harmful elements (PHEs) is currently exacerbated by their widespread use in several industrial, agricultural, domestic and technological applications. PHEs cause adverse health effects on immune and nervous systems and can lead to cancer development via genotoxic mechanisms. The present study is an integrated approach that aims at assessing the genotoxicity of bioaccessible PHEs following ingestion of contaminated house dust. A multidisciplinary methodology associating chemical characterization of five house dust samples, extraction of the bioaccessible PHEs in gastric extracts by the unified BARGE method, determination of the bioaccessible fraction and in vitro genotoxicity of gastric extracts in adenocarcinoma gastric human (AGS) cells was developed. The five gastric extracts induced dose-dependent genotoxicity in AGS cells. Copper (bioaccessible concentration up to 111 mg/kg) was probably the prevalent PHE inducing primary DNA damage (up to 5.1-fold increase in tail DNA at 0.53 g/l of gastric extract). Lead (bioaccessible concentration up to 245 mg/kg) was the most prevalent PHE inducing chromosome-damaging effects (r = 0.55; p \ 0.001 for micronucleated cells induction). The association of principal component analysis and Spearman's correlations was decisive to understand the chromosome-damaging properties of the bioacces-sible PHEs in AGS cells. This methodology could be used on a larger-scale study to provide useful information for science-based decision-making in regulatory policies, and a better estimation of human exposure and associated health risks

Biological properties of a neutralized 2.5% sodium hypochlorite solution

International audienceObjectives. The objective of this study was to evaluate the influence of neutralizing a 2.5% NaOCl solution on its cytotoxicity, genotoxicity, and tissue-dissolving potential.Study design. The cytotoxicity and the genotoxicity of Dakin, a 2.5% NaOCl solution, and a neutralized 2.5% NaOCl solution were assessed according to ISO 10993 standards. The weight of palatal mucosa samples placed in neutralized 2.5% NaOCl, 2.5% NaOCl was recorded over time as well as the pH of the solutions.Results. The neutralized 2.5% NaOCl solution was 10-fold more cytotoxic than the 2.5% NaOCl solution. None of the solutions was genotoxic. The 2.5% NaOCl solution had a better tissue-dissolving capacity than the neutralized 2.5% NaOCl solution. The pH of the 2.5% NaOCl solution and neutralized 2.5% NaOCl solution decreased from 12 to 9 and from 7.5 to 5.6, respectively.Conclusion. Neutralizing a 2.5% NaOCl solution increased its cytotoxicity, did not induce any genotoxic effect, and reduced its tissue-dissolving ability

Effects of environmental concentrations of chlordecone (insecticide), glyphosate-based herbicide and imazalil (fungicide) in mixture on reproductive rate, morphology and DNA damage in the freshwater Hydra vulgaris

International audienceBetween 1972 and 1993, 300 of tons of the organochlorine insecticide chlordecone (C10Cl10O, CLD) were dumped in the banana plantations of the French West Indies (FWI) to eradicate the black banana weevil (Cosmopolites sordidus). Today, almost all of their water and soil is contaminated and will remain so until remedial action is taken, if at all. Also, in 1999, massive water pollution in FWI was revealed as well as contamination of the majority of the population. The results of a monitoring conducted by the French Water Board showed that 130 phytosanitary products were detected: 40% herbicides, 30% insecticides and 23% fungicides. Among them, the herbicide glyphosate and the fungicide imazalil (IMAZ) were the still frequently detected (Wintz & Pak, 2021). In rivers, CLD concentrations was up to 100 µg/L due not only to erosion and runoff but also to a widely used herbicide, glyphosate. Indeed, glyphosate causes increased soil erosion and leads to the release of stable CLD stored in the soils of polluted fields (Sabatier et al., 2021). Imazalil, a possible human carcinogen according to the U.S. EPA, is a fungicide that was notably used in post-harvest treatment to control storage diseases. Banned for this use as of May 2019, it appears to be initiating a slow decline in impacted rivers (Wintz & Pak, 2021). The objective of this work was to evaluate, in the cnidarian Hydra vulgaris, Pallas, 1766, the effects of chronic exposure to mixtures of CLD, IMAZ, and a glyphosate-based herbicide (GBH) at environmental concentrations found in FWI rivers. Because multiscale studies evaluating multiple biological responses from the molecular to the population level should be emphasized in environmental toxicology, reproductive rates, morphological damage, and DNA integrity were determined after 14-day exposure. Three concentrations of CLD (0.1, 10, and 20 μg/L) were combined with a cocktail of IMAZ and GBH at two concentration levels: the lowest level (1 µg/L IMAZ and 2 µg/L active ingredient (a.i.) glyphosate GBH) and the highest level (1.5 µg/L IMAZ and 3 µg a.i./L GBH). All concentrations used in the mixtures were also tested alone. The impacts on morphology suggest a coalition of these three pesticides in mixtures at the lowest concentrations tested here. This result emphasizes the importance of testing mixtures at low environmental concentrations for which the compounds tested separately have no effect on the biological endpoint being evaluated. This work also allowed the development of a Comet assay that demonstrated primary DNA damage in hydra exposed to CLD, GBH, IMAZ and their various mixtures. The results of the comet assay are difficult to interpret and the fact that all conditions induce primary DNA damage does not necessarily mean that the compounds alone or in mixtures are genotoxic. The lesions observed after 14 days may be due to indirect factors, e.g., oxidative stress and the poor cellular state of the hydra, whose morphology is very altered. Further investigation is required before a conclusion can be reached on the issue of genotoxicity. These preliminary results, obtained on hydra, suggest that CLD in these mixtures is the agent that induce most of the toxicity observed and that remediation programs aimed at eliminating the CLD in situ would be valuable for the quality of the environment in the West Indies.References.- Sabatier P, Mottes C, Cottin N, Evrard O, Comte I, Piot C, Gay B, Arnaud F, Lefevre I, Develle AL, Deffontaines L, Plet J, Lesueur-Jannoyer M, Poulenard J (2021) Evidence of chlordecone resurrection by glyphosate in French West Indies. Environ Sci Technol 55:2296-2306. https://dx.doi.org/10.1021/acs.est.0c05207.- Wintz L, Pak LT (2021) Appui au pilotage et au suivi du dispositif d’instrumentation pour la caractérisation des pesticides du bassin versant pilote martiniquais du Galion. Caractérisation des évolutions des concentrations en pesticides dans les eaux de surface du bassin versant du Galion en Martinique : résultats sur quatre années de suivi. ODE/CIRAD 060-03-2019. 78 page

In vitro genotxicity evaluation of PAHs in mixtures using experimental design.

International audienc

In vitro genotxicity evaluation of PAHs in mixtures using experimental design.

International audienc

Gaining insight into genotoxicity with the comet assay in inhomogenoeous exposure scenarios: The effects of tritiated steel and cement particles on human lung cells in an inhalation perspective

International audienceThe comet assay was recently applied for the first time to test the genotoxicity of micrometric stainless steel and cement particles, representative of those produced in the dismantling of nuclear power plants. A large dataset was obtained from in vitro exposure of BEAS-2B lung cells to different concentrations of hydrogenated (non-radiative control) and tritiated particles, to assess the impact of accidental inhalation. Starting from the distributions of the number of nuclei scored at different extent of DNA damage (% tail DNA values), we propose a new comet data treatment designed to consider the inhomogeneity of the action of such particles. Indeed, due to particle behavior in biological media and concentration, a large fraction of cells remains undamaged, and standard averaging of genotoxicity indicators leads to a misinterpretation of experimental results. The analysis we propose reaches the following goals: genotoxicity in human lung cells is assessed for stainless steel and cement microparticles; the role of radiative damage due to tritium is disentangled from particulate stress; the fraction of damaged cells and their average level of DNA damage are assessed separately, which is essential for carcinogenesis implications and sets the basis for a better-informed risk management for human exposure to radioactive particles