Development of a transient expression assay for detecting environmental oestrogens in zebrafish and medaka embryos

This is the final version of the article. Available from BMC via the DOI in this record.BACKGROUND: Oestrogenic contaminants are widespread in the aquatic environment and have been shown to induce adverse effects in both wildlife (most notably in fish) and humans, raising international concern. Available detecting and testing systems are limited in their capacity to elucidate oestrogen signalling pathways and physiological impacts. Here we developed a transient expression assay to investigate the effects of oestrogenic chemicals in fish early life stages and to identify target organs for oestrogenic effects. To enhance the response sensitivity to oestrogen, we adopted the use of multiple tandem oestrogen responsive elements (EREc38) in a Tol2 transposon mediated Gal4ff-UAS system. The plasmid constructed (pTol2_ERE-TATA-Gal4ff), contains three copies of oestrogen response elements (3ERE) that on exposure to oestrogen induces expression of Gal4ff which this in turn binds Gal4-responsive Upstream Activated Sequence (UAS) elements, driving the expression of a second reporter gene, EGFP (Enhanced Green Fluorescent Protein). RESULTS: The response of our construct to oestrogen exposure in zebrafish embryos was examined using a transient expression assay. The two plasmids were injected into 1-2 cell staged zebrafish embryos, and the embryos were exposed to various oestrogens including the natural steroid oestrogen 17ß-oestradiol (E2), the synthetic oestrogen 17α- ethinyloestradiol (EE2), and the relatively weak environmental oestrogen nonylphenol (NP), and GFP expression was examined in the subsequent embryos using fluorescent microscopy. There was no GFP expression detected in unexposed embryos, but specific and mosaic expression of GFP was detected in the liver, heart, somite muscle and some other tissue cells for exposures to steroid oestrogen treatments (EE2; 10 ng/L, E2; 100 ng/L, after 72 h exposures). For the NP exposures, GFP expression was observed at 10 μg NP/L after 72 h (100 μg NP/L was toxic to the fish). We also demonstrate that our construct works in medaka, another model fish test species, suggesting the transient assay is applicable for testing oestrogenic chemicals in fish generally. CONCLUSION: Our results indicate that the transient expression assay system can be used as a rapid integrated testing system for environmental oestrogens and to detect the oestrogenic target sites in developing fish embryos.This work was co-funded by the Natural Environmental Research Council (reference NE/I014470/1) and the University of Exeter (CRT/TK). OL was funded by a studentship from Gwanglim, Korea

Acute Toxicity, Teratogenic, and Estrogenic Effects of Bisphenol A and Its Alternative Replacements Bisphenol S, Bisphenol F, and Bisphenol AF in Zebrafish Embryo-Larvae

This is the author accepted manuscript. The final version is available from American Chemical Society via the DOI in this recordBisphenol A (BPA), a chemical incorporated into plastics and resins, has estrogenic activity and is associated with adverse health effects in humans and wildlife. Similarly structured BPA analogues are widely used but far less is known about their potential toxicity or estrogenic activity in vivo. We undertook the first comprehensive analysis on the toxicity and teratogenic effects of the bisphenols BPA, BPS, BPF, and BPAF in zebrafish embryo-larvae and an assessment on their estrogenic mechanisms in an estrogen-responsive transgenic fish Tg(ERE:Gal4ff)(UAS:GFP). The rank order for toxicity was BPAF > BPA > BPF > BPS. Developmental deformities for larval exposures included cardiac edema, spinal malformation, and craniofacial deformities and there were distinct differences in the effects and potencies between the different bisphenol chemicals. These effects, however, occurred only at concentrations between 1.0 and 200 mg/L which exceed those in most environments. All bisphenol compounds induced estrogenic responses in Tg(ERE:Gal4ff)(UAS:GFP) zebrafish that were inhibited by coexposure with ICI 182 780, demonstrating an estrogen receptor dependent mechanism. Target tissues included the heart, liver, somite muscle, fins, and corpuscles of Stannius. The rank order for estrogenicity was BPAF > BPA = BPF > BPS. Bioconcentration factors were 4.5, 17.8, 5.3, and 0.067 for exposure concentrations of 1.0, 1.0, 0.10, and 50 mg/L for BPA, BPF, BPAF, and BPS, respectively. We thus show that these BPA alternatives induce similar toxic and estrogenic effects to BPA and that BPAF is more potent than BPA, further highlighting health concerns regarding the use of BPA alternatives.Natural Environment Research Council (NERC

Clathrate hydrates as a sink of noble gases in Titan's atmosphere

We use a statistical thermodynamic approach to determine the composition of clathrate hydrates which may form from a multiple compound gas whose composition is similar to that of Titan's atmosphere. Assuming that noble gases are initially present in this gas phase, we calculate the ratios of xenon, krypton and argon to species trapped in clathrate hydrates. We find that these ratios calculated for xenon and krypton are several orders of magnitude higher than in the coexisting gas at temperature and pressure conditions close to those of Titan's present atmosphere at ground level. Furthermore we show that, by contrast, argon is poorly trapped in these ices. This trapping mechanism implies that the gas-phase is progressively depleted in xenon and krypton when the coexisting clathrate hydrates form whereas the initial abundance of argon remains almost constant. Our results are thus compatible with the deficiency of Titan's atmosphere in xenon and krypton measured by the {\it Huygens} probe during its descent on January 14, 2005. However, in order to interpret the subsolar abundance of primordial Ar also revealed by {\it Huygens}, other processes that occurred either during the formation of Titan or during its evolution must be also invoked.Comment: Astronomy & Astrophysics Letters, in pres

Biosensor zebrafish provide new insights into potential health effects of environmental estrogens

This is the final version of the article. Available from NIEHS via the DOI in this record.BACKGROUND: Environmental estrogens alter hormone signaling in the body that can induce reproductive abnormalities in both humans and wildlife. Available testing systems for estrogens are focused on specific systems such as reproduction. Crucially, however, the potential for significant health impacts of environmental estrogen exposures on a variety of body systems may have been overlooked. OBJECTIVE: Our aim was to develop and apply a sensitive transgenic zebrafish model to assess real-time effects of environmental estrogens on signaling mechanisms in a whole body system for use in integrated health assessments. METHODS: We created a novel transgenic biosensor zebrafish containing an estrogen-inducible promoter derived with multiple tandem estrogen responsive elements (EREs) and a Gal4ff-UAS system for enhanced response sensitivity. RESULTS: Using our novel estrogen-responsive transgenic (TG) zebrafish, we identified target tissues for environmental estrogens; these tissues have very high sensitivity even at environmentally relevant concentrations. Exposure of the TG fish to estrogenic endocrine-disrupting chemicals (EDCs) induced specific expression of green fluorescent protein (GFP) in a wide variety of tissues including the liver, heart, skeletal muscle, otic vesicle, forebrain, lateral line, and ganglions, most of which have not been established previously as targets for estrogens in fish. Furthermore, we found that different EDCs induced GFP expression with different tissue response patterns and time trajectories, suggesting different potential health effects. CONCLUSION: We have developed a powerful new model for understanding toxicological effects, mechanisms, and health impacts of environmental estrogens in vertebrates

New nonuniform black string solutions

We present nonuniform vacuum black strings in five and six spacetime dimensions. The conserved charges and the action of these solutions are computed by employing a quasilocal formalism. We find qualitative agreement of the physical properties of nonuniform black strings in five and six dimensions. Our results offer further evidence that the black hole and the black string branches merge at a topology changing transition. We generate black string solutions of the Einstein-Maxwell-dilaton theory by using a Harrison transformation. We argue that the basic features of these solutions can be derived from those of the vacuum black string configurations.Comment: 30 pages, 12 figures; v2: more details on numerical method, references added; v3: references added, minor revisions, version accepted by journa

Thermal phases of D1-branes on a circle from lattice super Yang-Mills

We report on the results of numerical simulations of 1+1 dimensional SU(N) Yang-Mills theory with maximal supersymmetry at finite temperature and compactified on a circle. For large N this system is thought to provide a dual description of the decoupling limit of N coincident D1-branes on a circle. It has been proposed that at large N there is a phase transition at strong coupling related to the Gregory-Laflamme (GL) phase transition in the holographic gravity dual. In a high temperature limit there was argued to be a deconfinement transition associated to the spatial Polyakov loop, and it has been proposed that this is the continuation of the strong coupling GL transition. Investigating the theory on the lattice for SU(3) and SU(4) and studying the time and space Polyakov loops we find evidence supporting this. In particular at strong coupling we see the transition has the parametric dependence on coupling predicted by gravity. We estimate the GL phase transition temperature from the lattice data which, interestingly, is not yet known directly in the gravity dual. Fine tuning in the lattice theory is avoided by the use of a lattice action with exact supersymmetry.Comment: 21 pages, 8 figures. v2: References added, two figures were modified for clarity. v3: Normalisation of lattice coupling corrected by factor of two resulting in change of estimate for c_cri

New Phases of Near-Extremal Branes on a Circle

We study the phases of near-extremal branes on a circle, by which we mean near-extremal branes of string theory and M-theory with a circle in their transverse space. We find a map that takes any static and neutral Kaluza-Klein black hole, i.e. any static and neutral black hole on Minkowski-space times a circle M^d x S^1, and map it to a corresponding solution for a near-extremal brane on a circle. The map is derived using first a combined boost and U-duality transformation on the Kaluza-Klein black hole, transforming it to a solution for a non-extremal brane on a circle. The resulting solution for a near-extremal brane on a circle is then obtained by taking a certain near-extremal limit. As a consequence of the map, we can transform the neutral non-uniform black string branch into a new non-uniform phase of near-extremal branes on a circle. Furthermore, we use recently obtained analytical results on small black holes in Minkowski-space times a circle to get new information about the localized phase of near-extremal branes on a circle. This gives in turn predictions for the thermal behavior of the non-gravitational theories dual to these near-extremal branes. In particular, we give predictions for the thermodynamics of supersymmetric Yang-Mills theories on a circle, and we find a new stable phase of (2,0) Little String Theory in the canonical ensemble for temperatures above its Hagedorn temperature.Comment: 72 pages, 5 figures. v2: Typos fixed, refs. added. v3: Sec. 3.2 fixe

Sequences of Bubbles and Holes: New Phases of Kaluza-Klein Black Holes

We construct and analyze a large class of exact five- and six-dimensional regular and static solutions of the vacuum Einstein equations. These solutions describe sequences of Kaluza-Klein bubbles and black holes, placed alternately so that the black holes are held apart by the bubbles. Asymptotically the solutions are Minkowski-space times a circle, i.e. Kaluza-Klein space, so they are part of the (\mu,n) phase diagram introduced in hep-th/0309116. In particular, they occupy a hitherto unexplored region of the phase diagram, since their relative tension exceeds that of the uniform black string. The solutions contain bubbles and black holes of various topologies, including six-dimensional black holes with ring topology S^3 x S^1 and tuboid topology S^2 x S^1 x S^1. The bubbles support the S^1's of the horizons against gravitational collapse. We find two maps between solutions, one that relates five- and six-dimensional solutions, and another that relates solutions in the same dimension by interchanging bubbles and black holes. To illustrate the richness of the phase structure and the non-uniqueness in the (\mu,n) phase diagram, we consider in detail particular examples of the general class of solutions.Comment: 71 pages, 22 figures, v2: Typos fixed, comment added in sec. 5.

On non-uniform smeared black branes

We investigate charged dilatonic black $p$-branes smeared on a transverse circle. The system can be reduced to neutral vacuum black branes, and we perform static perturbations for the reduced system to construct non-uniform solutions. At each order a single master equation is derived, and the Gregory-Laflamme critical wavelength is determined. Based on the non-uniform solutions, we discuss thermodynamic properties of this system and argue that in a microcanonical ensemble the non-uniform smeared branes are entropically disfavored even near the extremality, if the spacetime dimension is $D \le 13 +p$, which is the critical dimension for the vacuum case. However, the critical dimension is not universal. In a canonical ensemble the vacuum non-uniform black branes are thermodynamically favorable at $D > 12+p$, whereas the non-uniform smeared branes are favorable at $D > 14+p$ near the extremality.Comment: 24 pages, 2 figures; v2: typos corrected, submitted to Class.Quant.Gra

Naked shell singularities on the brane

By utilizing non-standard slicings of 5-dimensional Schwarzschild and Schwarzschild-AdS manifolds based on isotropic coordinates, we generate static and spherically symmetric braneworld spacetimes containing shell-like naked null singularities. For planar slicings, we find that the brane-matter sourcing the solution is a perfect fluid with an exotic equation of state and a pressure singularity where the brane crosses the bulk horizon. From a relativistic point of view, such a singularity is required to maintain matter infinitesimally above the surface of a black hole. From the point of view of the AdS/CFT conjecture, the singular horizon can be seen as one possible quantum correction to a classical black hole geometry. Various generalizations of planar slicings are also considered for a Ricci-flat bulk, and we find that singular horizons and exotic matter distributions are common features.Comment: REVTeX4, 13 pages, 6 figures, references and comments adde