Development and application of cell-based bioassays for the evaluation of AhR-mediated effects on human health

This thesis aims to develop and apply novel in vitro bioassays to evaluate the effect of chemicals and mixtures of chemicals on the AhR with the objective to improve human relevance and optimize the interpretability of bioassay results towards human exposure. The research presented focused on the interaction of different classes of chemicals (dioxins, PAHs and mycotoxins) with the AhR and its public-health relevance. Species-specific differences regarding AhR-signalling in terms of hazard prediction were also discussed because of their potential implications for human health-based hazard assessment. In chapter 2, a human-based AhR luciferase reporter gene assay was developed to provide a human cellular-context needed for the subsequent studies. In this chapter, we applied the newly developed human liver cell-based DRhuman CALUX bioassay to determine relative potency values for PCDD/Fs and PBDD/Fs. The values were compared to values obtained in the rodent-based DR CALUX bioassay and the WHO-TEF values for hazard assessment. Chapter 3 describes an application of the DRhuman CALUX bioassay for the determination of dioxin-like activity in consumer goods, such as plastic toys. An analytical chemistry-based measurement was compared to the activity in vitro determined by the DR CALUX and its human variant, and a first approximation of human hazard assessment based on the ingestion of contaminated plastic from toys by children was performed. Chapter 4 describes a second AhR-mediated bioassay, a high performance (hp) variant of the rodent-based DR CALUX bioassay with improved sensitivity, the DRhp CALUX. In this study, we showed that the bioassay is suitable for highly sensitive detection and quantification of low levels of PAH-mediated AhR activity in human samples. Hence, the DRhp CALUX was applied to quantify the effect of exposure to PAHs via maternal smoking on placental and foetal hepatic PAH-based AhR activity. Chapter 5 describes a study in which in vitro bioassays, including the DRhuman CALUX, were applied to individual and combined hepatotoxicity of carcinogenic mycotoxins aflatoxin B1 and a main fungal precursor, versicolorin A. This study sheds a light on a previously unexplored role of the AhR concerning the carcinogenicity of aflatoxins and discusses hazard assessment related to aflatoxin mixtures. Finally, Chapter 6 summarizes the most important findings of the studies in this thesis, discusses their relevance for hazard assessment, and presents an outlook for future work. The research presented in this thesis was conducted between 2017 and 2020 within the EU Horizon 2020 research and innovation program under the Marie Sklodowska-Curie ITN ProtectED (PROTECTion against Endocrine Disruptors: detection, mixtures, health effects, risk assessment and communication; grant agreement No. 722635). The work presented in chapter 4 & 5 is the result of collaborations from BioDetection Systems B.V. with partners from the ProtectED network, the University of Aberdeen (chapter 4) and the French National Research Institute for Agriculture, Food and the Environment (INRAe, chapter 5). The primary aim of the ProtectED project was to develop innovative and improved analysis capabilities for the risk assessment (and communication) of the impact of endocrine disruptors and their mixtures on human health and the environment.