Interactions between chemical and climate stressors: A role for mechanistic toxicology in assessing climate change risks

Incorporation of global climate change (GCC) effects into assessments of chemical risk and injury requires integrated examinations of chemical and nonchemical stressors. Environmental variables altered by GCC (temperature, precipitation, salinity, pH) can influence the toxicokinetics of chemical absorption, distribution, metabolism, and excretion as well as toxicodynamic interactions between chemicals and target molecules. In addition, GCC challenges processes critical for coping with the external environment (water balance, thermoregulation, nutrition, and the immune, endocrine, and neurological systems), leaving organisms sensitive to even slight perturbations by chemicals when pushed to the limits of their physiological tolerance range. In simplest terms, GCC can make organisms more sensitive to chemical stressors, while alternatively, exposure to chemicals can make organisms more sensitive to GCC stressors. One challenge is to identify potential interactions between nonchemical and chemical stressors affecting key physiological processes in an organism. We employed adverse outcome pathways, constructs depicting linkages between mechanism-based molecular initiating events and impacts on individuals or populations, to assess how chemical- and climate-specific variables interact to lead to adverse outcomes. Case examples are presented for prospective scenarios, hypothesizing potential chemicalGCC interactions, and retrospective scenarios, proposing mechanisms for demonstrated chemicalclimate interactions in natural populations. Understanding GCC interactions along adverse outcome pathways facilitates extrapolation between species or other levels of organization, development of hypotheses and focal areas for further research, and improved inputs for risk and resource injury assessments. Environ. Toxicol. Chem. 2013;32:3248. (c) 2012 SETA

Identification of Metabolites of Trenbolone Acetate in Androgenic Runoff from a Beef Feedlot

Little is known concerning the potential ecological effects of hormonally active substances associated with discharges from animal feeding operations. Trenbolone acetate is a synthetic anabolic steroid that is widely used in the United States to promote growth of beef cattle. Metabolites of trenbolone acetate include the stereoisomers 17α- and 17β-trenbolone, both of which are stable in animal wastes and are relatively potent androgens in fish and mammals. Our purpose in this study was to evaluate the occurrence of 17α- and 17β-trenbolone in a beef cattle feedlot discharge and in river water upstream and downstream from the discharge. In conjunction with that effort, we measured in vitro androgenic activity of the discharge using CV-1 cells that had been transiently cotransfected with human androgen receptor and reporter gene constructs. Samples were collected on nine different occasions during 2002 and 2003. Whole-water samples from the discharge caused a significant androgenic response in the CV-1 cells and contained detectable concentrations of 17α- and 17β-trenbolone. Further work is needed to ascertain the degree to which synthetic androgens such as trenbolone contribute to androgenic activity of feedlot discharges

Direct Effects, Compensation, and Recovery in Female Fathead Minnows Exposed to a Model Aromatase Inhibitor

BackgroundSeveral chemicals in the environment have the potential to inhibit aromatase, an enzyme critical to estrogen synthesis.ObjectivesThe objective of this study was to provide a detailed characterization of molecular and biochemical responses of female fathead minnows to a model aromatase inhibitor, fadrozole (FAD).MethodsFish were exposed via water to 0, 3, or 30 microg FAD/L for 8 days and then held in clean water for 8 days, with samples collected at four time points during each 8-day period. We quantified ex vivo steroid production, plasma steroids, and plasma vitellogenin (Vtg) concentrations and analyzed relative transcript abundance of 10 key regulatory genes in ovaries and 3 in pituitary tissue by real-time polymerase chain reaction.ResultsEx vivo 17beta-estradiol (E2) production and plasma E2 and Vtg concentrations were significantly reduced after a single day of exposure to 3 microg or 30 microg FAD/L. However, plasma E2 concentrations recovered by the eighth day of exposure in the 3-microg/L group and within 1 day of cessation of exposure in the 30-microg/L group, indicating concentration- and time-dependent physiologic compensation and recovery. Concentration-dependent increases in transcripts coding for aromatase (A isoform), cytochrome P450 side-chain cleavage, steroidogenic acute regulatory protein, and follicle-stimulating hormone receptor all coincided with increased E2 production and recovery of plasma E2 concentrations.ConclusionsResults of this research highlight the need to consider compensation/adaptation and recovery when developing and interpreting short-term bioassays or biomarkers or when trying to predict the effects of chemical exposures based on mode of action

Gene expression responses in male fathead minnows exposed to binary mixtures of an estrogen and antiestrogen

<p>Abstract</p> <p>Background</p> <p>Aquatic organisms are continuously exposed to complex mixtures of chemicals, many of which can interfere with their endocrine system, resulting in impaired reproduction, development or survival, among others. In order to analyze the effects and mechanisms of action of estrogen/anti-estrogen mixtures, we exposed male fathead minnows (<it>Pimephales promelas</it>) for 48 hours via the water to 2, 5, 10, and 50 ng 17α-ethinylestradiol (EE₂)/L, 100 ng ZM 189,154/L (a potent antiestrogen known to block activity of estrogen receptors) or mixtures of 5 or 50 ng EE₂/L with 100 ng ZM 189,154/L. We analyzed gene expression changes in the gonad, as well as hormone and vitellogenin plasma levels.</p> <p>Results</p> <p>Steroidogenesis was down-regulated by EE₂as reflected by the reduced plasma levels of testosterone in the exposed fish and down-regulation of genes in the steroidogenic pathway. Microarray analysis of testis of fathead minnows treated with 5 ng EE₂/L or with the mixture of 5 ng EE₂/L and 100 ng ZM 189,154/L indicated that some of the genes whose expression was changed by EE₂were blocked by ZM 189,154, while others were either not blocked or enhanced by the mixture, generating two distinct expression patterns. Gene ontology and pathway analysis programs were used to determine categories of genes for each expression pattern.</p> <p>Conclusion</p> <p>Our results suggest that response to estrogens occurs via multiple mechanisms, including canonical binding to soluble estrogen receptors, membrane estrogen receptors, and other mechanisms that are not blocked by pure antiestrogens.</p

Investigating Alternatives to the fish early-life stage test: A strategy for discovering and annotating adverse outcome pathways for early fish development

The fish early-life stage (FELS) test (Organisation for Economic Co-operation and Development [OECD] test guideline 210) is the primary test used internationally to estimate chronic fish toxicity in support of ecological risk assessments and chemical management programs. As part of an ongoing effort to develop efficient and cost-effective alternatives to the FELS test, there is a need to identify and describe potential adverse outcome pathways (AOPs) relevant to FELS toxicity. To support this endeavor, the authors outline and illustrate an overall strategy for the discovery and annotation of FELS AOPs. Key events represented by major developmental landmarks were organized into a preliminary conceptual model of fish development. Using swim bladder inflation as an example, a weight-of-evidence–based approach was used to support linkage of key molecular initiating events to adverse phenotypic outcomes and reduced young-of-year survival. Based on an iterative approach, the feasibility of using key events as the foundation for expanding a network of plausible linkages and AOP knowledge was explored and, in the process, important knowledge gaps were identified.Given the scope and scale of the task, prioritization of AOP development was recommended and key research objectives were defined relative to factors such as current animal-use restrictions in the European Union and increased demands for fish toxicity data in chemical management programs globally. The example and strategy described are intended to guide collective efforts to define FELS-related AOPs and develop resource-efficient predictive assays that address the toxicological domain of the OECD 210 test.Keywords: Risk assessment, Aquatic toxicology, Adverse outcome pathways, Mode of action, Animal alternative, Fish early-life stage toxicity, Swim bladderKeywords: Risk assessment, Aquatic toxicology, Adverse outcome pathways, Mode of action, Animal alternative, Fish early-life stage toxicity, Swim bladde

Use of chemical mixtures to differentiate mechanisms of endocrine action in a small fish model

Various assays with adult fish have been developed to identify potential endocrine-disrupting chemicals (EDCs) which may cause toxicity via alterations in the hypothalamic-pituitary-gonadal (HPG) axis. These assays can be sensitive and highly diagnostic for key mechanisms such as agonism of the estrogen and androgen receptors (ERs, ARs) and inhibition of steroid synthesis. However, most of the tests do not unambiguously identify AR antagonists. The purpose of this work was to explore the utility of a mixture test design with the fathead minnow (Pimephales promelas) for detecting different classes of EDCs including AR antagonists. Adults of both sexes were exposed via the water to EDCs with diverse mechanisms of action in the absence or presence of 17beta-trenbolone (TB), a potent AR agonist which masculinizes female fathead minnows. Similar to previous studies with the model AR antagonists flutamide and vinclozolin, exposure of females to the AR antagonist cyproterone acetate in the presence of TB decreased expression of an easily-observed masculinization response, nuptial tubercle formation. Mixture studies with TB and the model ER agonists, 17alpha-ethinylestradiol and bisphenol A, also showed inhibition of tubercle formation in the females, but unlike the AR antagonists, the estrogens markedly induced synthesis of vitellogenin (VTG: egg yolk protein), particularly in males. The ER agonists also offset TB-induced depressions in plasma VTG concentrations in female fish. Additional mixture experiments were conducted with TB and triclocarban, an anti-microbial reported to enhance AR-mediated responses, or ammonia, a "negative control" with no known direct effects on HPG function. Neither chemical affected VTG status in males or females in the absence or presence of TB; however, both slightly enhanced TB-induced tubercle formation in females. Based on studies described herein and elsewhere with the fathead minnow, a TB co-exposure assay appears to be an effective approach for clearly identifying AR antagonists as well as potential EDCs with other relevant mechanisms of action

Integrating Omic Technologies into Aquatic Ecological Risk Assessment and Environmental Monitoring: Hurdles, Achievements, and Future Outlook

Background: In this commentary we present the findings from an international consortium on fish toxicogenomics sponsored by the U.K. Natural Environment Research Council (Fish Toxicogenomics—Moving into Regulation and Monitoring, held 21–23 April 2008 at the Pacific Environmental Science Centre, Vancouver, BC, Canada). Objectives: The consortium from government agencies, academia, and industry addressed three topics: progress in ecotoxicogenomics, regulatory perspectives on roadblocks for practical implementation of toxicogenomics into risk assessment, and dealing with variability in data sets. Discussion: Participants noted that examples of successful application of omic technologies have been identified, but critical studies are needed to relate molecular changes to ecological adverse outcome. Participants made recommendations for the management of technical and biological variation. They also stressed the need for enhanced interdisciplinary training and communication as well as considerable investment into the generation and curation of appropriate reference omic data. Conclusions: The participants concluded that, although there are hurdles to pass on the road to regulatory acceptance, omics technologies are already useful for elucidating modes of action of toxicants and can contribute to the risk assessment process as part of a weight-of-evidence approach

Pharmaceuticals and personal care products in the environment: What are the big questions?

Background: Over the past 10-15 years, a substantial amount of work has been done by the scientific, regulatory, and business communities to elucidate the effects and risks of pharmaceuticals and personal care products (PPCPs) in the environment. Objective: This review was undertaken to identify key outstanding issues regarding the effects of PPCPs on human and ecological health in order to ensure that future resources will be focused on the most important areas. Data sources: To better understand and manage the risks of PPCPs in the environment, we used the "key question" approach to identify the principle issues that need to be addressed. Initially, questions were solicited from academic, government, and business communities around the world. A list of 101 questions was then discussed at an international expert workshop, and a top-20 list was developed. Following the workshop, workshop attendees ranked the 20 questions by importance. Data synthesis: The top 20 priority questions fell into seven categories: a) prioritization of substances for assessment, b) pathways of exposure, c) bioavailability and uptake, d) effects characterization, e) risk and relative risk, f) antibiotic resistance, and g) risk management. Conclusions: A large body of information is now available on PPCPs in the environment. This exercise prioritized the most critical questions to aid in development of future research programs on the topic.Centro de Investigaciones del Medioambient