Early embryo development is one of the most sensitive stages to environmental chemicals during the whole life. Prenatal exposures to many environmental chemicals have been shown to impact fetal development and be associated with adverse health outcomes in later life stages. However, the effects of chemical mixture exposure on developing embryos, especially in early developmental stages, have yet to be fully studied. To fulfill this research gap, my thesis was divided into three data chapters and mainly aimed at investigating the effects of a chemical mixture on human early-stage embryo development. In Chapter 2, I chose methylmercury (MeHg) as the main study toxicant to establish procedures for embryotoxicity testing using human embryonic stem cells (hESCs). I then characterized the effects of low doses of MeHg on this stem cell model by screening a set of cell fate decision-related makers and found MeHg is embryotoxic, which is consistent with epidemiological and in vivo findings. In Chapter 3, I studied the embryotoxicity of a chemical mixture that consists of 23 individual environmental chemicals (including MeHg) detected from the maternal blood samples of pregnant women in Nunavik, labelled as Nunavik Contaminant Mixture (NCM), using the same cell model. The effects of NCM exposure on hESCs were compared to MeHg exposure alone. NCM exposure adversely affected cell viability and adhesion, induced apoptosis, disrupted the cell cycle, altered the expression of cytoskeleton and autophagy proteins, and changed the levels of lineage marker gene and protein expressions in a dose-dependent manner. Some distinct effects on hESCs between NCM exposure and MeHg alone exposure were noticed, and the potential interactions among the chemical components within a chemical mixture were indicated. In Chapter 4, I studied the effects of MeHg exposure during the formation of definitive endoderm (DE) cells from hESCs and compared that to MeHg's effects on undifferentiated hESCs. I found that cell specification towards endoderm could be affected by MeHg exposure, mainly through disrupting calcium homeostasis and over-generating reactive oxygen species, leading to increased ribosome biogenesis and protein synthesis. Moreover, MeHg effects are state-dependent; MeHg enhances pluripotency in undifferentiated hESCs, but it promotes differentiation during DE induction. Taken together, this thesis verifies the value of hESCs in testing the embryotoxicity and developmental toxicity of environmental chemicals, enriches the understanding of the toxicity of MeHg and NCM, emphasizes the necessity of evaluating the effects of chemical mixtures and provides new directions in studying environmental chemical toxicity using stem cells. Findings from my thesis could hopefully contribute to predicting the potential effects of prenatal environmental chemical exposures and aid in developing evidence-based public health policy.

L’Homme est exposé tout au long de sa vie à de nombreux contaminants présents dans l’environnement et l’alimentation généralement à faibles doses et en mélanges. L’évaluation des risques pose problème dans la mesure où il est bien établi qu’il existe des différences de réponse entre l’homme et l’animal. Quelle que soit la voie d’exposition, de par son rôle majeur dans la biotransformation des xénobiotiques, le foie est considéré comme un organe cible pour de nombreuses classes de produits chimiques potentiellement cytotoxiques, génotoxiques voire cancérogènes. Nous avons utilisé la lignée de cellules hépatiques humaines HepaRG non transformées pour évaluer la toxicité chronique et/ou le pouvoir cancérogène de pesticides et de composés génotoxiques présents dans l’environnement. Cette lignée est la seule connue pour avoir conservé des propriétés proches des hépatocytes humains en culture primaire. Dans une première partie nous avons confirmé le maintien de ses capacités fonctionnelles à confluence par l’analyse du transcriptome et de la biocinétique de 4 médicaments, après traitements quotidiens pendant 14 jours. Nous avons ensuite recherché les effets de mélanges de pesticides après des expositions aiguës et répétées. Nous avons ainsi montré que : 1. l’isomalathion, une impureté majeure du malathion, joue un rôle prépondérant sur la toxicité hépatique de ce dernier et qu’il inhibe la carboxylesterase impliquée dans le métabolisme des deux composés; 2. l’endosulfan et le méthoxychlore, deux organochlorés métabolisés par les CYP3A4 et 2B6, agissent de manière synergique sur leur cytotoxicité après une exposition unique ou répétée. De plus, alors que l'activité du CYP3A4 est inhibée de manière réversible par l’endosulfan, le méthoxychlore l’augmente. En revanche, l’activité du CYP2B6 est induite par les deux pesticides. Lorsqu’ils sont en mélange équimolaire un effet additif ou antagoniste est observé sur l'activité du CYP3A4 et du CYP2B6 respectivement quelle que soit la durée de l’exposition. Enfin, dans une troisième partie, nous avons exposé des cellules HepaRG pendant une quinzaine de passages à de faibles doses de deux contaminants génotoxiques nécessitant une bioactivation, l’aflatoxine B1 et une amine aromatique hétérocyclique, le PhiP, et démontré l’acquisition de propriétés de cellules transformées (par exemple croissance sur agar, migration dans le test de griffure et surexpression de gènes associés au cancer). Au total, nos résultats démontrent tout l’intérêt que représente la lignée hépatique humaine HepaRG métaboliquement compétentes pour l’étude de la toxicité chronique et/ou le potentiel cancérogène des contaminants de l’environnement. Ils ont permis de mettre en évidence d’interactions entre des pesticides en mélanges binaires et pour la première fois d’analyser le potentiel cancérogène de contaminants génotoxiques dans une lignée hépatique humaine. / Humans are exposed throughout their life to many environmental and food contaminants, usually at low doses and in mixtures. Risk assessment remains questionable as it is well established that there are differences in the response to chemicals between humans and animals. Regardless of the route of exposure, due to its major role in xenobiotic biotransformation, the liver is considered as a target organ for many classes of chemicals potentially cytotoxic, genotoxic or carcinogenic. We used the HepaRG cell line to evaluate chronic toxicity and/or carcinogenicity of pesticides and genotoxic compounds. This cell line is the only one known to exhibit properties similar to those of human hepatocytes in primary culture. In the first part we confirmed the maintenance of functional capacities of these cells at confluence by transcriptomic and biokinetic analysis of several drugs after a 14-day treatment. We then investigated the effects of mixtures of pesticides after acute and repeated exposures. We showed that : 1. Isomalathion, a major impurity of malathion, played a leading role on liver toxicity and inhibited carboxylesterase that is involved in the metabolism of these two compounds; 2. Endosulfan and methoxychlor, two organochlorines, metabolized by CYP3A4 and CYP2B6, acted synergistically on their cytotoxicity after single or repeated exposure. Moreover, whereas activity of CYP3A4 was reversibly inhibited by endosulfan and increased by methoxychlor. By contrast, CYP2B6 activity was induced by these two pesticides while in equimolar mixtures, they caused additive or antagonistic effects on CYP3A4 and CYP2B6 activities respectively, regardless of the duration of exposure. Finally, in the third part, we exposed HepaRG cells for up to 15 passages to low doses of two genotoxic contaminants which required bioactivation, aflatoxin B1 and heterocyclic aromatic amine, PhIP, and demonstrated the appearance of properties of transformed cells (e.g. growth on agar, cell migration in the wound healing test and overexpression of a number of genes associated with cancer). Altogether, our results demonstrate the great potential interest that represents the metabolically competent human liver cell line HepaRG for the study of chronic toxicity and/or carcinogenic potential of environmental contaminants. They highlight possible interactions between pesticides in binary mixtures and for the first time, demonstrate that the carcinogenic potential of genotoxic contaminants can be analyzed in an human hepatic cell line.