Effects of common pesticides on prostaglandin D2 (PGD2) inhibition in SC5 mouse sertoli cells, evidence of binding at the cox-2 active site, and implications for endocrine disruption

Background: There are concerns that diminished prostaglandin action in fetal life could increase the risk of congenital malformations. Many endocrine-disrupting chemicals have been found to suppress prostaglandin synthesis, but to our knowledge, pesticides have never been tested for these effects. Objectives: We assessed the ability of pesticides that are commonly used in the European Union to suppress prostaglandin D2 (PGD2) synthesis. Methods: Changes in PGD2 secretion in juvenile mouse Sertoli cells (SC5 cells) were measured using an ELISA. Coincubation with arachidonic acid (AA) was conducted to determine the site of action in the PGD2 synthetic pathway. Molecular modeling studies were performed to assess whether pesticides identified as PGD2-active could serve as ligands of the cyclooxygenase-2 (COX-2) binding pocket. Results: The pesticides boscalid, chlorpropham, cypermethrin, cyprodinil, fenhexamid, fludioxonil, imazalil (enilconazole), imidacloprid, iprodione, linuron, methiocarb, o-phenylphenol, pirimiphos- methyl, pyrimethanil, and tebuconazole suppressed PGD2 production. Strikingly, some of these substances—o-phenylphenol, cypermethrin, cyprodinil, linuron, and imazalil (enilconazole)— showed potencies (IC50) in the range between 175 and 1,500 nM, similar to those of analgesics intended to block COX enzymes. Supplementation with AA failed to reverse this effect, suggesting that the sites of action of these pesticides are COX enzymes. The molecular modeling studies revealed that the COX-2 binding pocket can accommodate most of the pesticides shown to suppress PGD2 synthesis. Some of these pesticides are also capable of antagonizing the androgen receptor. Conclusions: Chemicals with structural features more varied than previously thought can suppress PGD2 synthesis. Our findings signal a need for in vivo studies to establish the extent of endocrinedisrupting effects that might arise from simultaneous interference with PGD2 signaling and androgen action

Sea bass (Dicentrarchus labrax) : a model organism for the screening of estrogenic chemicals in marine surface waters?

Society of Environmental Toxicology and Chemistry - SETAC Europe 14th Annual Meeting, Prague, Czech Republic, April 2004.There is growing concern that aquatic wildlife in surface waters of the European Union is exposed to natural and man-made chemicals that have the ability to mimic estrogens and lead to reproductive dysfunction. Estrogenic responses in fish are the net result of complex chains of events involving the uptake, distribution and metabolism of test agents until they interact with their target sites. Typically these aspects cannot be modelled in short-term cell-based assays, only studies with vertebrates offer the opportunity to assess potential interactions of test compounds at higher organisational levels. However, studies with endocrine disrupting chemicals have been performed mainly with freshwater organisms. The sensitivity of a marine fish species to different estrogenic chemicals was investigated under chronic exposure conditions. This work is part of a study focusing on the combination effects of mixtures of estrogenic chemicals in marine and freshwater organisms (ACE, EVK1-CT-2001-100). As test organism the sea bass (Dicentrarchus labrax) was selected, a common species in European marine systems. Juveniles were exposed under a flow-through system for 14 days for a set of reference chemicals (17Ã -estradiol, ethynylestradiol, nonylphenol, octylphenol, bisphenol A). Effects at subcellular level were analysed using vitellogenesis as endpoint. Its relevance is evaluated by further investigations about the individual fitness (condition factor, hepatossomatic index), as well as the liver cytochrome P450 activity. The general suitability of the sea bass as a model organism for the screening of estrogenic chemicals in the marine environment is discussed.Comissãoo Europeia (CE) - ACE, EVK1-CT-2001-100

Sea bass (Dicentrarchus labrax) : a model organism for assessing multi-level responses to estrogenic chemicals in marine surface waters

Resumo apresentado sob poster ao 5th International Symposium of Fish Endocrinology, CAstellon, Spain, Setember 5-9, 2004.There is growing concern that aquatic wildlife in surface waters of the European Union is exposed to natural and man-made chemicals that have the ability to mimic estrogens and lead to reproductive dysfunction. Estrogenic responses in fish are the net result of complex chains of events involving the uptake, distribution and metabolism of test agents until they interact with their target sites. Typically these aspects cannot be modelled in short-term cell-based assays, only studies with vertebrates offer the opportunity to assess potential interactions of test compounds at higher organisational levels. The most widely studied biological response in fish to environmental estrogens is the production of vitellogenin (Vtg). However, few studies have attempted to link this endpoint with effects on xenobiotic biotransformation enzymes and genotoxic responses. This work is part of a study focusing on the combination effects of mixtures of estrogenic chemicals in marine and freshwater organisms. As test organism the sea bass (Dicentrarchus labrax) was selected, a common species in European marine systems. Juveniles were exposed under a flow-through system for 14 days to the natural estrogen 17ß-estradiol and ethynylestradiol. Actual chemical concentrations in the water-column were determined by gas chromatography with ion trap detection. Effects at subcellular level were analysed using Vtg as a reference endpoint [1]. Its relevance is evaluated by further investigations on liver 7-ethoxyresorufin-O-deethylase (EROD) activity and erythrocytic nuclear abnormalities [2]. These measurements were integrated with organism level endpoints (i.e. condition factor, hepatossomatic index) to provide evidence for cause-effect of estrogenic contamination. The general suitability of the sea bass as a model organism for the screening of estrogenic chemicals in the marine environment is discussed.Comissão Europeia (CE) - ACE, EVK1-CT-2001-100

Genotoxic mixtures and dissimilar action: Concepts for prediction and assessment

This article has been made available through the Brunel Open Access Publishing Fund. This article is distributed under the terms of the creative commons Attribution license which permits any use, distribution, and reproduction in any medium, provided the original author(s)and the source are credited.Combinations of genotoxic agents have frequently been assessed without clear assumptions regarding their expected (additive) mixture effects, often leading to claims of synergisms that might in fact be compatible with additivity. We have shown earlier that the combined effects of chemicals, which induce micronuclei (MN) in the cytokinesis-block micronucleus assay in Chinese hamster ovary-K1 cells by a similar mechanism, were additive according to the concept of concentration addition (CA). Here, we extended these studies and investigated for the first time whether valid additivity expectations can be formulated for MN-inducing chemicals that operate through a variety of mechanisms, including aneugens and clastogens (DNA cross-linkers, topoisomerase II inhibitors, minor groove binders). We expected that their effects should follow the additivity principles of independent action (IA). With two mixtures, one composed of various aneugens (colchicine, flubendazole, vinblastine sulphate, griseofulvin, paclitaxel), and another composed of aneugens and clastogens (flubendazole, doxorubicin, etoposide, melphalan and mitomycin C), we observed mixture effects that fell between the additivity predictions derived from CA and IA. We achieved better agreement between observation and prediction by grouping the chemicals into common assessment groups and using hybrid CA/IA prediction models. The combined effects of four dissimilarly acting compounds (flubendazole, paclitaxel, doxorubicin and melphalan) also fell within CA and IA. Two binary mixtures (flubendazole/paclitaxel and flubendazole/doxorubicin) showed effects in reasonable agreement with IA additivity. Our studies provide a systematic basis for the investigation of mixtures that affect endpoints of relevance to genotoxicity and show that their effects are largely additive.UK Food Standards Agenc

Relative perversity

We define and study a relative perverse $t$-structure associated with any finitely presented morphism of schemes $f: X\to S$, with relative perversityequivalent to perversity of the restrictions to all geometric fibres of $f$. The existence of this $t$-structure is closely related to perverse $t$-exactness properties of nearby cycles. This $t$-structure preserves universally locally acyclic sheaves, and one gets a resulting abelian category $\mathrm{Perv}^{\mathrm{ULA}}(X/S)$ with many of the same properties familiar in the absolute setting (e.g., noetherian, artinian, compatible with Verdier duality). For $S$ connected and geometrically unibranch with generic point $\eta$, the functor $\mathrm{Perv}^{\mathrm{ULA}}(X/S)\to$\mathrm{Perv}(X_\eta)$ is exact and fully faithful, and its essential image is stable under passage to subquotients. This yields a notion of "good reduction" for perverse sheaves

Transgenic Expression of the Anti-parasitic Factor TEP1 in the Malaria Mosquito Anopheles gambiae

Mosquitoes genetically engineered to be resistant to Plasmodium parasites represent a promising novel approach in the fight against malaria. The insect immune system itself is a source of anti-parasitic genes potentially exploitable for transgenic designs. The Anopheles gambiae thioester containing protein 1 (TEP1) is a potent anti-parasitic protein. TEP1 is secreted and circulates in the mosquito hemolymph, where its activated cleaved form binds and eliminates malaria parasites. Here we investigated whether TEP1 can be used to create malaria resistant mosquitoes. Using a GFP reporter transgene, we determined that the fat body is the main site of TEP1 expression. We generated transgenic mosquitoes that express TEP1r, a potent refractory allele of TEP1, in the fat body and examined the activity of the transgenic protein in wild-type or TEP1 mutant genetic backgrounds. Transgenic TEP1r rescued loss-of-function mutations, but did not increase parasite resistance in the presence of a wild-type susceptible allele. Consistent with previous reports, TEP1 protein expressed from the transgene in the fat body was taken up by hemocytes upon a challenge with injected bacteria. Furthermore, although maturation of transgenic TEP1 into the cleaved form was impaired in one of the TEP1 mutant lines, it was still sufficient to reduce parasite numbers and induce parasite melanization. We also report here the first use of Transcription Activator Like Effectors (TALEs) in Anopheles gambiae to stimulate expression of endogenous TEP1. We found that artificial elevation of TEP1 expression remains moderate in vivo and that enhancement of endogenous TEP1 expression did not result in increased resistance to Plasmodium. Taken together, our results reveal the difficulty of artificially influencing TEP1-mediated Plasmodium resistance, and contribute to further our understanding of the molecular mechanisms underlying mosquito resistance to Plasmodium parasites

The ACE Project: a synopsis of in vivo studies to predict estrogenic mixture effects in freshwater and marine fish

Society of Environmental Toxicology and Chemistry - SETAC Europe 15th Annual Meeting, Lille, France, May 2005.This work is part of the ACE project (ACE, EVK1-CT-2001-100) which aim is to investigate multi-component mixtures of estrogenic compounds in aquatic ecosystems. Here we present a synopsis of in vivo data related with the joint estrogenic action of five estrogenic compounds (17ß-estradiol, ethynylestradiol, nonylphenol, octylphenol and bisphenol-A) on vitellogenesis in fathead minnow (Pimephales promelas) and sea bass (Dicentrarchus labrax). The studies were conducted with freshwater adult males and marine juveniles under flow through exposure conditions for two weeks. In the first step, fish were exposed to the five compounds individually in order to generate concentration- response curves. Therefore mixture effects were predicted on the basis of the potency of each compound by using the model of concentration addition (CA). Finally, the compounds were tested as a mixture at equipotent concentrations, and the observed mixture effects were compared to the predictions. The mixture studies showed an good agreement between observed and predicted effects and provided evidence that CA can be used as a predictive tool for the effect assessment of mixtures of (xeno)estrogens in freshwater or marine ecosystems. The differences/limitations of running in vivo mixture studies with freshwater and marine species will be discussed.Comissão Europeia (CE) - ACE project - ACE, EVK1-CT-2001-100

(Meta-)stable reconstructions of the diamond(111) surface: interplay between diamond- and graphite-like bonding

Off-lattice Grand Canonical Monte Carlo simulations of the clean diamond (111) surface, based on the effective many-body Brenner potential, yield the $(2\times1)$ Pandey reconstruction in agreement with \emph{ab-initio} calculations and predict the existence of new meta-stable states, very near in energy, with all surface atoms in three-fold graphite-like bonding. We believe that the long-standing debate on the structural and electronic properties of this surface could be solved by considering this type of carbon-specific configurations.Comment: 4 pages + 4 figures, Phys. Rev. B Rapid Comm., in press (15Apr00). For many additional details (animations, xyz files) see electronic supplement to this paper at http://www.sci.kun.nl/tvs/carbon/meta.htm

A novel behavioral fish model of nociception for testing analgesics

Pain is a major symptom in many medical conditions, and often interferes significantly with a person's quality of life. Although a priority topic in medical research for many years, there are still few analgesic drugs approved for clinical use. One reason is the lack of appropriate animal models that faithfully represent relevant hallmarks associated with human pain. Here we propose zebrafish (Danio rerio) as a novel short-term behavioral model of nociception, and analyse its sensitivity and robustness. Firstly, we injected two different doses of acetic acid as the noxious stimulus. We studied individual locomotor responses of fish to a threshold level of nociception using two recording systems: a video tracking system and an electric biosensor (the MOBS system). We showed that an injection dose of 10% acetic acid resulted in a change in behavior that could be used to study nociception. Secondly, we validated our behavioral model by investigating the effect of the analgesic morphine. In time-course studies, first we looked at the dose-response relationship of morphine and then tested whether the effect of morphine could be modulated by naloxone, an opioid antagonist. Our results suggest that a change in behavioral responses of zebrafish to acetic acid is a reasonable model to test analgesics. The response scales with stimulus intensity, is attenuated by morphine, and the analgesic effect of morphine is blocked with naloxone. The change in behavior of zebrafish associated with the noxious stimulus can be monitored with an electric biosensor that measures changes in water impedance. © 2011 by the authors; licensee MDPI, Basel, Switzerland

Dispelling urban myths about default uncertainty factors in chemical risk assessment - Sufficient protection against mixture effects?

© 2013 Martin et al.; licensee BioMed Central LtdThis article has been made available through the Brunel Open Access Publishing Fund.Assessing the detrimental health effects of chemicals requires the extrapolation of experimental data in animals to human populations. This is achieved by applying a default uncertainty factor of 100 to doses not found to be associated with observable effects in laboratory animals. It is commonly assumed that the toxicokinetic and toxicodynamic sub-components of this default uncertainty factor represent worst-case scenarios and that the multiplication of those components yields conservative estimates of safe levels for humans. It is sometimes claimed that this conservatism also offers adequate protection from mixture effects. By analysing the evolution of uncertainty factors from a historical perspective, we expose that the default factor and its sub-components are intended to represent adequate rather than worst-case scenarios. The intention of using assessment factors for mixture effects was abandoned thirty years ago. It is also often ignored that the conservatism (or otherwise) of uncertainty factors can only be considered in relation to a defined level of protection. A protection equivalent to an effect magnitude of 0.001-0.0001% over background incidence is generally considered acceptable. However, it is impossible to say whether this level of protection is in fact realised with the tolerable doses that are derived by employing uncertainty factors. Accordingly, it is difficult to assess whether uncertainty factors overestimate or underestimate the sensitivity differences in human populations. It is also often not appreciated that the outcome of probabilistic approaches to the multiplication of sub-factors is dependent on the choice of probability distributions. Therefore, the idea that default uncertainty factors are overly conservative worst-case scenarios which can account both for the lack of statistical power in animal experiments and protect against potential mixture effects is ill-founded. We contend that precautionary regulation should provide an incentive to generate better data and recommend adopting a pragmatic, but scientifically better founded approach to mixture risk assessment. © 2013 Martin et al.; licensee BioMed Central Ltd.Oak Foundatio