MOLECULAR AND METABOLIC EFFECTS OF ENDOCRINE-DISRUPTING CHEMICALS (EDCs)

There is growing concern about the potential adverse effects on human health of exposure to endocrine-disrupting chemicals (EDCs) in food, air and water. The continuous release of these contaminants, including metals, pesticides and pharmaceuticals into the environment leads to their bioaccumulation and biomagnification through the food chain, raising significant animal welfare and food safety concerns. Humans and wildlife are exposed to complex mixtures of contaminants that may interact and produce cumulative, synergistic or antagonistic effects depending on their nature, mechanisms of action and cellular responses. Among the adverse effects, oxidative stress plays a central role. It results from an imbalance between the production of reactive oxygen species (ROS) and the body's capacity to detoxify them, leading to damage to cellular components such as proteins, lipids and DNA. Despite growing recognition of these issues, the combined effects of multiple pollutants and how they interact in the body remain poorly understood, highlighting a critical gap in current research. In this context, the aim of this thesis was to define a biochemical and molecular approach to assess the cellular and molecular responses to the exposure to EDCs, both as single compounds and in complex mixtures. The study focused on marine fish collected from an area characterised by high levels of contamination by a wide range of EDCs (Chapter 1). Furthermore, two experimental model systems were employed: an in vivo model using Sparus aurata (European sea bass) as key species in marine research (Chapter 2) and an in vitro model using human hepatoma cell line (HepG2) (Chapter 3) to provide a comprehensive understanding of the effects of EDC mixtures in different biological systems. The first study (Chapter 1) investigated the effects of marine contaminants on the food chain, with particular emphasis on the bioaccumulation of mercury (Hg) in fish collected from Augusta Bay. The results showed significant increases in Hg levels and alterations in oxidative stress biomarkers, including reduced lipid content and polyunsaturated fatty acids (PUFAs), along with increased malondialdehyde (MDA), a biomarker of lipid damage. This study demonstrates the usefulness of evaluating bioaccumulation levels in marine organisms, integrated with the study of biochemical and molecular biomarkers related to health, metabolism and quality. This approach could provide insights into how the occurrence of contaminants in the environment affects the welfare and quality of marine organisms, with significant adverse effects at the ecological level and within our food system. The second study (Chapter 2) investigated the in vivo effects of exposure to EDC mixtures including BDE-47, CdCl₂, and CBZ (which are prevalent in the environment) on the liver of Sparus aurata. The fish were exposed to two different doses of the mixture for 15 days, followed by a 15-day detoxification period, to assess both immediate and potential long-term effects, as well as the effects of detoxification. The results showed that chronic exposure to the contaminant mixture altered the oxidative balance, affected the antioxidant defence system, modified fatty acids content and increased lipid peroxidation levels. The observed changes at the molecular and biochemical levels suggest that exposure to EDC mixtures could lead to potential long-term or permanent consequences, affecting individual organisms and possibly extending to population level. The final study (Chapter 3) examined the in vitro effects of 72-hours exposure of human HepG2 cells to sub-lethal mixtures of EDCs, specifically BDE-47, CdCl₂, and CBZ. The results showed that the exposure to mixtures interfered with the enzymatic detoxification and antioxidant system, impairing DNA repair defence mechanisms involved in resistance to oxidative stress. The combined effect of the compounds at sub-lethal doses results in a greater activation of these pathways compared to exposure to each single compound, thereby exacerbating their cytotoxicity. The observed changes at the molecular level could contribute to the identification of early warning biomarkers for environmental monitoring, while providing insight into the toxicity, hazard levels and associated health risks of these substances in the environment. This thesis emphasises the importance of monitoring contaminant levels and oxidative stress biomarkers in marine organisms to evaluate the effects of environmental contaminants on their health and food safety. The observed changes in marine organisms (in vivo) and human cell line (in vitro) exposed to contaminant mixtures demonstrate the usefulness of the experimental model systems, and of the biomarkers used to monitor the effects of single and combined chemical contaminants. The findings contribute to a better understanding of the effects of contaminants on marine organisms and the marine food chain, with direct implications for human health.