Biotransformations of bisphenol A in a mammalian model: answers and new questions raised by low-dose metabolic fate studies in pregnant CD1 mice.

We investigated the metabolic fate of a low dose (25 micro g/kg) of bisphenol A [2,2-bis(4-hydroxy-phenyl)propane] (BPA) injected subcutaneously in CD1 pregnant mice using a tritium-labeled molecule. Analytic methods were developed to allow a radio-chromatographic profiling of BPA residues in excreta and tissues, as well as in mothers' reproductive tracts and fetuses, that contained more than 4% of the administered radioactivity. BPA was extensively metabolized by CD1 mice. Identified metabolite structures included the glucuronic acid conjugate of BPA, several double conjugates, and conjugated methoxylated compounds, demonstrating the formation of potentially reactive intermediates. Fetal radioactivity was associated with unchanged BPA, BPA glucuronide, and a disaccharide conjugate. The latter structure, as well as that of a dehydrated glucuronide conjugate of BPA (a major metabolite isolated from the digestive tract), showed that BPA metabolic routes were far more complex than previously thought. The estrogenicity of the metabolites that were identified but not tested for hormonal activity cannot be ruled out; however, in general, conjugated BPA metabolites have significantly lower potency than that of the parent compound. Thus, these data suggest the parental compound is responsible for the estrogenic effects observed in fetuses exposed to BPA during gestation in this mammalian model

Bisphenol A induces otolith malformations during vertebrate embryogenesis

<p>Abstract</p> <p>Background</p> <p>The plastic monomer and plasticizer bisphenol A (BPA), used for manufacturing polycarbonate plastic and epoxy resins, is produced at over 2.5 million metric tons per year. Concerns have been raised that BPA acts as an endocrine disruptor on both developmental and reproductive processes and a large body of evidence suggests that BPA interferes with estrogen and thyroid hormone signaling. Here, we investigated BPA effects during embryonic development using the zebrafish and <it>Xenopus </it>models.</p> <p>Results</p> <p>We report that BPA exposure leads to severe malformations of the otic vesicle. In zebrafish and in <it>Xenopus </it>embryos, exposure to BPA during the first developmental day resulted in dose-dependent defects in otolith formation. Defects included aggregation, multiplication and occasionally failure to form otoliths. As no effects on otolith development were seen with exposure to micromolar concentrations of thyroid hormone, 17-ß-estradiol or of the estrogen receptor antagonist ICI 182,780 we conclude that the effects of BPA are independent of estrogen receptors or thyroid-hormone receptors. Na⁺/K⁺ATPases are crucial for otolith formation in zebrafish. Pharmacological inhibition of the major Na⁺/K⁺ATPase with ouabain can rescue the BPA-induced otolith phenotype.</p> <p>Conclusions</p> <p>The data suggest that the spectrum of BPA action is wider than previously expected and argue for a systematic survey of the developmental effects of this endocrine disruptor.</p

The Dividing Line between Federal and State Promotion of Aeronautics

<p>The model xeno-estrogen bisphenol A (BPA) has been extensively studied over the past two decades, contributing to major advances in the field of endocrine disrupting chemicals research. Besides its well documented adverse effects on reproduction and development observed in rodents, latest studies strongly suggest that BPA disrupts several endogenous metabolic pathways, with suspected steatogenic and obesogenic effects. BPA's adverse effects on reproduction are attributed to its ability to activate estrogen receptors (ERs), but its effects on metabolism and its mechanism(s) of action at low doses are so far only marginally understood. Metabolomics based approaches are increasingly used in toxicology to investigate the biological changes induced by model toxicants and chemical mixtures, to identify markers of toxicity and biological effects. In this study, we used proton nuclear magnetic resonance (¹H-NMR) based untargeted metabolite profiling, followed by multivariate statistics and computational analysis of metabolic networks to examine the metabolic modulation induced in human hepatic cells (HepG2) by an exposure to low and very low doses of BPA (10⁻⁶M, 10⁻⁹M, and 10⁻¹²M), vs. the female reference hormone 17β-estradiol (E2, 10⁻⁹M, 10⁻¹²M, and 10⁻¹⁵M). Metabolomic analysis combined to metabolic network reconstruction highlighted different mechanisms at lower doses of exposure. At the highest dose, our results evidence that BPA shares with E2 the capability to modulate several major metabolic routes that ensure cellular functions and detoxification processes, although the effects of the model xeno-estrogen and of the natural hormone can still be distinguished.</p

Perinatal Exposure to Environmentally Relevant Levels of Bisphenol A Decreases Fertility and Fecundity in CD-1 Mice

Bac k g r o u n d: Perinatal exposure to low-doses of bisphenol A (BPA) results in alterations in the ovary, uterus, and mammary glands and in a sexually dimorphic region of the brain known to be important for estrous cyclicity. Objectives: We aimed to determine whether perinatal exposure to environmentally relevant doses of BPA alters reproductive capacity. Met h o d s: Female CD-1 mice that were exposed to BPA at 0, 25 ng, 250 ng, or 25 µg/kg body weight (BW)/day or diethylstilbestrol (DES) at 10 ng/kg BW/day (positive control) from gestational day 8 through day 16 of lactation were continuously housed with proven breeder males for 32 weeks starting at 2 months of age. At each delivery, pups born to these mating pairs were removed. The cumulative number of pups, number of deliveries, and litter size were recorded. The purity of the BPA used in this and our previous studies was assessed using HPLC, mass spectrometry, and nuclear magnetic resonance. Res u l t s: The forced breeding experiment revealed a decrease in the cumulative number of pups, observed as a nonmonotonic dose–response effect, and a decline in fertility and fecundity over time in female mice exposed perinatally to BPA. The BPA was 97 % pure, with no evidence of contaminatio

Use of reconstituted metabolic networks to assist in metabolomic data visualization and mining

Metabolomics experiments seldom achieve their aim of comprehensively covering the entire metabolome. However, important information can be gleaned even from sparse datasets, which can be facilitated by placing the results within the context of known metabolic networks. Here we present a method that allows the automatic assignment of identified metabolites to positions within known metabolic networks, and, furthermore, allows automated extraction of sub-networks of biological significance. This latter feature is possible by use of a gap-filling algorithm. The utility of the algorithm in reconstructing and mining of metabolomics data is shown on two independent datasets generated with LC–MS LTQ-Orbitrap mass spectrometry. Biologically relevant metabolic sub-networks were extracted from both datasets. Moreover, a number of metabolites, whose presence eluded automatic selection within mass spectra, could be identified retrospectively by virtue of their inferred presence through gap filling

Development of new approach methods for the identification and characterization of endocrine metabolic disruptors-a PARC project

In past times, the analysis of endocrine disrupting properties of chemicals has mainly been focused on (anti-)estrogenic or (anti-)androgenic properties, as well as on aspects of steroidogenesis and the modulation of thyroid signaling. More recently, disruption of energy metabolism and related signaling pathways by exogenous substances, so-called metabolism-disrupting chemicals (MDCs) have come into focus. While general effects such as body and organ weight changes are routinely monitored in animal studies, there is a clear lack of mechanistic test systems to determine and characterize the metabolism-disrupting potential of chemicals. In order to contribute to filling this gap, one of the project within EU-funded Partnership for the Assessment of Risks of Chemicals (PARC) aims at developing novel in vitro methods for the detection of endocrine metabolic disruptors. Efforts will comprise projects related to specific signaling pathways, for example, involving mTOR or xenobiotic-sensing nuclear receptors, studies on hepatocytes, adipocytes and pancreatic beta cells covering metabolic and morphological endpoints, as well as metabolism-related zebrafish-based tests as an alternative to classic rodent bioassays. This paper provides an overview of the approaches and methods of these PARC projects and how this will contribute to the improvement of the toxicological toolbox to identify substances with endocrine disrupting properties and to decipher their mechanisms of action

The goliath project: Towards an internationally harmonised approach for testing metabolism disrupting compounds

Copyright © 2020 by the authors. The purpose of this project report is to introduce the European “GOLIATH” project, a new research project which addresses one of the most urgent regulatory needs in the testing of endocrine-disrupting chemicals (EDCs), namely the lack of methods for testing EDCs that disrupt metabolism and metabolic functions. These chemicals collectively referred to as “metabolism disrupting compounds” (MDCs) are natural and anthropogenic chemicals that can promote metabolic changes that can ultimately result in obesity, diabetes, and/or fatty liver in humans. This project report introduces the main approaches of the project and provides a focused review of the evidence of metabolic disruption for selected EDCs. GOLIATH will generate the world’s first integrated approach to testing and assessment (IATA) specifically tailored to MDCs. GOLIATH will focus on the main cellular targets of metabolic disruption—hepatocytes, pancreatic endocrine cells, myocytes and adipocytes—and using an adverse outcome pathway (AOP) framework will provide key information on MDC-related mode of action by incorporating multi-omic analyses and translating results from in silico, in vitro, and in vivo models and assays to adverse metabolic health outcomes in humans at real-life exposures. Given the importance of international acceptance of the developed test methods for regulatory use, GOLIATH will link with ongoing initiatives of the Organisation for Economic Development (OECD) for test method (pre-)validation, IATA, and AOP development

Retardateurs de flamme bromés et pertubation des équilibres endocriniens (métabolisme comparé in vitro du tétrabromobisphénol A chez l'homme et chez le rat)

BORDEAUX2-BU Santé (330632101) / SudocSudocFranceF

Complementarity of phosphorylated histones H2AX and H3 quantification in different cell lines for genotoxicity screening

The in vitro micronucleus assay is broadly used, but is not per se able to discriminate aneugenic from clastogenic compounds, and cytotoxicity can be a confounding factor. In vitro genotoxicity assays generally rely on cell lines with limited metabolic capabilities. Recently, the use of histone H2AX and H3 phosphorylation markers (γH2AX and p-H3) was proposed to discriminate aneugenic from clastogenic chemicals. The aim of the present study was to develop a new genotoxic screening strategy based on the use of the γH2AX and p-H3 biomarkers in combination with cell lines with distinct biotransformation properties. First, we tested a training set of 20 model chemicals comprised of 10 aneugens, five clastogens and five cytotoxics on three human cell lines (HepG2, LS-174T and ACHN). Our data confirm the robustness of these two biomarkers to discriminate efficiently clastogens, aneugens and misleading cytotoxic chemicals in HepG2 cells. Aneugenic compounds induced either an increase or a decrease in p-H3 depending on their mode of action. Clastogens induced γH2AX, and cytotoxic compounds generated a marked decrease in these two biomarkers. Moreover, the use of different cell lines permits to discriminate direct from bioactivated genotoxins without the need of an exogenous metabolic activation system. Finally, we further evaluated this strategy using a test set of 13 chemicals with controversial genotoxic potential. The resulting data demonstrate that the combined analysis of γH2AX and p-H3 is an efficient strategy. Notably, we demonstrated that three compounds (fisetin, hydroquinone and okadaic acid) display both aneugenic and clastogenic properties

Toxicité neurocomportementale à court et à long-terme du BDE-99 chez le rat adulte ou en développement (Etude des effets de l'administration quotidienne par voie orale de doses représentatives de l'exposition humaine pendant 90 jours)

Les PBDEs appartiennent à la famille des retardateurs de flamme bromés, substances massivement utilisés ces dernières décennies. Ces molécules sont aujourd hui retrouvées de façon ubiquitaire dans l environnement extérieur et intérieur. L Homme est exposé à ces polluants par le biais de l alimentation et de l ingestion/inhalation de poussières ménagères ou industrielles contaminées. Leurs propriétés lipophiles et leur persistance sont à l origine de leur bioaccumulation dans les matrices abiotiques et biologiques (lait, sérum ). Les travaux de recherche de cette thèse ont eu pour objectif d évaluer l impact d une exposition à un polluant environnemental toxique et très répandu, le BDE-99, administré dans des conditions reflétant l exposition humaine. Ainsi, les effets d une administration de longue durée à des doses réalistes de BDE-99 (0,15, 1,5 et 15 g/kg/j) ont été évalués au plan neurocomportemental et physiologique chez le rat mâle de souche Sprague-Dawley. Aucune altération de l activité, de l anxiété et des fonctions cognitives n a été mise en évidence que les animaux aient été traités à l âge adulte, à partir du sevrage ou à partir du 8ème jour de vie postnatale. Les seules variations comportementales significatives observées ont été des troubles transitoires de la coordination locomotrice ainsi qu un retard d une journée de l ouverture des yeux chez les animaux traités avec le BDE-99 depuis l âge de 8 jours. Au plan physiologique, aucune variation significative de l évolution pondérale, des prises hydrique et alimentaire ainsi que du poids de plusieurs organes (foie, cerveau, reins, rate et thymus) n a été observée quelle que soit la dose de BDE-99 administrée et la période d exposition considérée. Ces résultats montrent donc que ce polluant administré chez le rat à des doses réalistes, reflétant celles auxquelles l Homme est réellement exposé, n est pas à même d induire une toxicité neurocomportementale que les animaux aient contaminés à l âge adulte ou bien lors de différentes phases du développement postnatal. Le fait d avoir utilisé des doses correspondantes au niveau d exposition environnemental est sans aucun doute à la base des discordances observées entre les résultats de ce travail et ceux des études publiées par ailleurs, soulignant ainsi la nécessité de réaliser des modèles d exposition les plus pertinents possibles de la réalité humaine pour pouvoir conclure au mieux quant au risque lié à cette famille de contaminants pour la santé humaine.PBDEs belong to flame retardants, substances heavily used in recent decades. These molecules are now found ubiquitously in the environment outside and inside. Humans are exposed to these pollutants through diet and ingestion/inhalation of household dust or contaminated industrial dust. Their lipophilic properties and their persistence are the cause of their bioaccumulation in abiotic and biological matrices (milk, serum ...). The research works of this thesis was aimed to assess the impact of exposure to a toxic environmental pollutant and very prevalent, BDE-99, administered in conditions reflecting the human exposure. Thus, the effects of long-term exposure to realistic doses of BDE-99 (0.15, 1.5 and 15 g/kg/day) were evaluated in terms neurobehavioral and physiological in male Sprague-Dawley rats. No alteration of the activity, anxiety and cognitive function has been highlighted that the animals were treated in adulthood, from weaning or from the 8th day of postnatal life. The only significant behavioral variations observed were transient locomotor coordination disorders and a delay of one day of eye opening in animals treated with BDE-99 from the age of 8 days. On a physiological level, no significant variation in body weight change, food and water consumption and weight of several organs (liver, brain, kidneys, spleen and thymus) was observed whatever the dose of BDE-99 administered and the exposure period considered. These results show that this pollutant administered to rats at realistic doses, reflecting those to which humans are actually exposed, is not able to induce neurobehavioral toxicity that the animal are contaminated in adulthood or at different phases of postnatal development. The fact to have used corresponding the level of environmental exposure is undoubtedly the basis of discrepancies observed between the results of this work and those of the published studies elsewhere, underscoring the necessity to realize exposure models the most possible relevant in order to able to conclude at best about risk associated with this family of contaminants to human health.NANCY-INPL-Bib. électronique (545479901) / SudocSudocFranceF