Endocrine disruption and obesity: an in vitro cell model for the assessment of metabolism disrupting chemicals and the evaluation of beneficial properties of nutraceuticals.

Metabolic diseases like metabolic syndrome, obesity and type II diabetes are nowadays matters of very high concern for human health. The incidence of these diseases is increasing constantly in both developing and developed countries. The causes of this epidemic are undoubtedly genetic background, lifestyle factors and high-fat diet. However, it has been recently observed that some environmental contaminants, such as the Endocrine Disrupting Chemicals (EDCs), can contribute to the development of these metabolic disorders. An EDC is an exogenous substance, or a mixture of substances, able to interfere with the proper functioning of the endocrine system at different levels. Metabolism Disrupting Chemicals (MDCs) are a subclass of EDCs that, inducing adipogenesis and lipid accumulation in target cells like adipocytes, promote obesity, diabetes, and fatty liver. Despite the increasing need to identify MDCs, an accepted standardized protocol to test them in vitro is not yet available. The 3T3-L1 murine pre-adipocyte cell line is an established cell culture model for screening MDCs because chemical treatment could affect pre-adipocytes differentiation into mature adipocytes. However, recent studies have highlighted great variability of the responses to various compounds, depending on cell sources and culture protocols used. The first aim of my thesis work was thus to compare diverse differentiation protocols utilizing 3T3-L1 pre-adipocyte cells coming from different suppliers. Three well-known MDCs were tested, namely Pyraclostrobin, Tributyltin chloride and Bisphenol A (BPA). This work showed indeed low reproducibility among the different conditions, though this largely did not impact determination of chemical activity classification (active/inactive). The standardization of the differentiation protocol would therefore be feasible and very advantageous to increase reproducibility and data comparability among laboratories. Then, I used the 3T3-L1 cell culture model to evaluate the possible pro-obesogenic effects of environmental EDCs mixtures. In this regard, I analyzed the effects of microplastic organic extracts containing several EDCs, sampled in the Adriatic Sea at different distances from the coastline. A strong increase in triglyceride accumulation was observed, highlighting the obesogenic activity of environmental pollutants carried by microplastics present in the sea. The last point I focused on in my thesis work is related to the investigation of the beneficial properties of nutraceuticals on reducing metabolic disruption driven by MDCs. Anti-obesogenic properties of (E)-β-caryophyllene (BCP) have been recently described, therefore I tested whether simultaneous exposure of 3T3-L1 cells to BPA and BCP could reduce the well-known obesogenic effect of BPA. The preliminary results obtained from these last experiments are worth to be expanded and confirmed in future studies.