De nombreux sites industriels émettent, non-intentionnellement, depuis des années des particules ultra-fines dans l’air. De nombreuses questions se posent sur leurs effets sur les écosystèmes et la santé humaine. Dans ce contexte, nous avons axé nos recherches autour de nanoparticules de fer manufacturées (Fe-NP), représentatives de celles que l’on retrouve dans les fumées des industries métallurgiques dunkerquoises et nous avons mené en parallèle des études d’écotoxicité et de toxicité. L’objectif principal de ces recherches était d’étudier l’impact de Fe-NP manufacturées exposées par voie aérienne sur deux modèles biologiques : Physcomitrella patens (Hedw.) Bruch & Schimp. et des cultures primaires de cellules épithéliales bronchiques humaines (HBEC). Pour répondre à ces objectifs, nous avons dû passer par l’étape indispensable qu’est la caractérisation fine de notre modèle de NP. En effet, la caractérisation physico-chimique doit être la plus complète possible (8 paramètres) de façon à déterminer, au préalable, leurs propriétés de surface. Puis, nous avons vérifié leur pénétration au sein de nos modèles biologiques. Puis, des biomarqueurs liés au stress oxydant ont été dosés chez la bryophyte, exposée à des concentrations faibles en Fe-NP. Premièrement, aucune perte de vitalité chez notre plante n’a pu être observée au cours du temps aux doses testées. Nos études n’ont pas permis de mettre en évidence une augmentation significative des espèces réactives de l’oxygène et du malondialdéhyde ; ainsi qu’une modulation significative du ratio GSSG/GSH, même si un phénomène de « sur-compensation » peut être évoqué sur le long terme, conduisant à la production de GSH au sein de notre plante témoignant d’une adaptation de la plante au stress. Enfin une analyse toxicogénomique a montré des modulations de l’expression (non significatives) de tous les isoformes des gènes d’intérêt étudiés aux doses testées. Dans le cadre d’études de toxicité, nous avons caractérisé notre modèle cellulaire par coloration immunocytologique. Puis, un test de viabilité nous a permis de choisir notre dose d’exposition : 2 μg.cm-2. Les travaux sur le stress oxydant et la modulation de l’expression génique ont été réalisés sur des cultures de cellules issues de trois patients différents pour prendre en compte la variabilité interindividuelle. Contrairement à certaines publications, nous n’avons pas montré une augmentation dose-dépendante des ROS. Puis, notre étude pangénomique, nous a permis de sélectionner 10 gènes d’intérêt dont l’étude approfondie a mis en évidence des effets précoces (dès 6 h d’exposition) sur des gènes impliquées dans des phénomènes inflammatoires. Néanmoins, les Fe-NP n'ont pas causé d’augmentation significative du MDA et du ratio GSSG/GSH après plusieurs jours d'exposition. Suite à ces résultats, il est maintenant envisageable de mener des recherches sur les impacts des nanoparticules d’origine environnementale à l’aide de nos deux modèles biologiques et d’améliorer les connaissances concernant leur danger potentiel pour l’environnement et la santé humaine. / Many industries emit, unintentionally, ultra-fine particles in the air, for many years. Many questions arise about their effects on ecosystems and human health. In this context, we focused our research on the iron-engineered nanoparticles (Fe-NP), representative of industrial smoke emitted by metallurgical industries and we conducted, in parallel, toxicity and ecotoxicity studies. The main objective of this work was to study the impact of Fe-NP exposed by air in two biological models: Physcomitrella patens (Hedw.) Bruch & Schimp. and primary cultures of human bronchial epithelial cells (HBEC). To meet these objectives, we had to characterize our NP model. Indeed, the physic-chemical characterization must be as complete as possible (8 parameters) to determine, firstly, their surface properties. Then, we checked their penetration within our biological models. And, oxidative stress biomarkers were measured in the bryophyte, exposed to low concentrations of Fe-NP. Firstly, any loss of vitality could be observed over time at the doses tested. Our studies have failed to demonstrate a significant increase of reactive oxygen species and malondialdehyde, and a significant modulation of the ratio GSSG/GSH, although the phenomenon of "over-compensation" can be discussed, over the long term, leading to the production of GSH in our plant showing a plant adaptation to stress. Finally a toxicogenomic analysis showed modulation of expression (not significant) of all isoforms of the genes of interest studied at the doses tested. For toxicity studies, we characterized our cellular model by immunocytological staining. Then, a viability test allowed us to choose the exposure dose: 2 μg.cm-2. The research on oxidative stress and the modulation of gene expression were performed on cells derived from three different patients to take into account individual variability. Unlike some publications, we did not show a dose-dependent increase of ROS. Then, the pangenomic study has allowed us to select 10 genes. A detailed study of this genes showed early effects (from 6 h of exposure) in genes involved in inflammation. However, Fe-NP did not cause any significant increase of MDA and GSSG/GSH ratio after several days of exposure. Following these results, it is now possible to conduct research on the impacts of ultra-fine particles using our two biological models and improve knowledge about their potential danger on the environment and human health.

<p>Generation of DNA damage is regarded to be an important initial event in the development cancer. Consequently, a battery of tests have been developed to detect different types of genotoxic effects in order to be able to predict the potential genotoxicity and mutagenicity of chemicals, including both pharmaceutical drugs and various types of environmental and occupational agents, as well as dietary factors. The aim of this thesis was to evaluate whether the combination of the comet assay and the extended-term cultures of human lymphocytes (ETC) can be used as an alternative <i>in vitro</i> system to more commonly used transformed mammalian cell lines, and primary cell cultures from humans, when testing the potential genotoxicity of chemicals. </p><p>Using the comet assay, a panel of reference compounds showed that the ETC were found to detect the DNA-damaging effects with no remarkable difference to what has been reported in other cell types. Moreover, in comparison with a well-established rodent cell line, the mouse lymphoma L5178Y cells, the ETC showed similar sensitivity to the DNA damaging effects of the genotoxic agents hydrogen peroxide and catechol. Although there was an interindividual variation in induced DNA damage and the subsequent repair when using ETC from different blood donors, it did not seem to be of crucial importance for the identification of DNA-damaging agents. The demonstrated difference in sensitivity to catechol-induced DNA damage between freshly isolated peripheral lymphocytes and ETC may very well be due to their different proliferative status but despite this difference, both <i>in vitro</i> systems were able to identify catechol as a DNA-damaging agent at the same concentration.</p><p>Based on these results, it is proposed that the ETC and the comet assay are a useful combination when testing for the potential DNA damaging effects of chemicals. Representing easily cultivated cells possessing the normal human karyotype, where one blood sample can be used for numerous experiments performed over a long time, extended-term cultures appear to be a useful alternative, both to transformed mammalian cell lines, and primary cell cultures from humans. In fact, the extended-term lymphocytes, with or without S9 and/or lesion specific DNA repair enzymes, should be used more frequently when screening for the potential genotoxicity of chemicals.</p>

Generation of DNA damage is regarded to be an important initial event in the development cancer. Consequently, a battery of tests have been developed to detect different types of genotoxic effects in order to be able to predict the potential genotoxicity and mutagenicity of chemicals, including both pharmaceutical drugs and various types of environmental and occupational agents, as well as dietary factors. The aim of this thesis was to evaluate whether the combination of the comet assay and the extended-term cultures of human lymphocytes (ETC) can be used as an alternative in vitro system to more commonly used transformed mammalian cell lines, and primary cell cultures from humans, when testing the potential genotoxicity of chemicals. Using the comet assay, a panel of reference compounds showed that the ETC were found to detect the DNA-damaging effects with no remarkable difference to what has been reported in other cell types. Moreover, in comparison with a well-established rodent cell line, the mouse lymphoma L5178Y cells, the ETC showed similar sensitivity to the DNA damaging effects of the genotoxic agents hydrogen peroxide and catechol. Although there was an interindividual variation in induced DNA damage and the subsequent repair when using ETC from different blood donors, it did not seem to be of crucial importance for the identification of DNA-damaging agents. The demonstrated difference in sensitivity to catechol-induced DNA damage between freshly isolated peripheral lymphocytes and ETC may very well be due to their different proliferative status but despite this difference, both in vitro systems were able to identify catechol as a DNA-damaging agent at the same concentration. Based on these results, it is proposed that the ETC and the comet assay are a useful combination when testing for the potential DNA damaging effects of chemicals. Representing easily cultivated cells possessing the normal human karyotype, where one blood sample can be used for numerous experiments performed over a long time, extended-term cultures appear to be a useful alternative, both to transformed mammalian cell lines, and primary cell cultures from humans. In fact, the extended-term lymphocytes, with or without S9 and/or lesion specific DNA repair enzymes, should be used more frequently when screening for the potential genotoxicity of chemicals.