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EVALUATION OF BIOLOGICAL ACTIVITY OF VITELLOGENIN EXPRESSION IN DIFFERENT AQUATIC MESOCOSM TROPIC LEVELS
Gordon, D. A., D. L. Lattier, J. M. Lazorchak, T. V. Reddy, L. W. Chang, G. P. Toth, D. W. Graham, F. J. deNoyelles, S. E. Campbell, D. E. Williams, AND B. E. Wiechman. EVALUATION OF BIOLOGICAL ACTIVITY OF VITELLOGENIN EXPRESSION IN DIFFERENT AQUATIC MESOCOSM TROPIC LEVELS. Presented at Society of Environmental Toxicology and Chemistry, Baltimore, MD, November 11-15, 2001.
Aquatic mesocosms were dosed with an environmentally relevant concentration of 17-a-ethinyl estradiol (EE2) to study the significance of trophic status (N, P levels) on the attenuation and bioavailability of synthetic estrogens in aquatic ecosystems. Estrogenic activity was assessed by the measurement of vitellogenin gene (Vg) expression in the liver of caged adult male fathead minnows (Pimephales promelas) using synthetic oligonucleotide primers which detect Vg messenger RNA in a quantitative reverse transcription-polymerase chain reaction. Caged male adult fathead minnows (Pimephales promelas) were maintained in outdoor mesocosms at the University of Kansas' Nelson Field Station and exposed to 0 or 20 ng/L EE2 at oligotrophic (total N and P, 0.77 and 0.015 mg/L), mesotrohpic (total N and P, 1.09 and 0.022 mg/L), or eutrophic (total N and P, 1.99 and 0.032 mg/L) nutrient levels. Randomly selected individuals were sacrificed at timepoints of 0, hr, 8 hr, 24 hr, 4 days, 7 days and 14 days post exosure and the livers were harvested for total RNA preparation. Vg expression was observed as early as 8 hr post exposure in all three trophic levels. Trend differences were observed among the three levels at the various timepoints. These results indicate that trophic level is a critical factor for assessing bioavailability of synthetic estrogens to aquatic species as part of a complete ecological risk assessment. These results will also provide data for the systhesis of environmental compartmental models for the fate and transport of model EDCs in aquatic ecosystems.
The indeterminate condition of exposure indicator research stands to change markedly with the ability to connect molecular biological technologies with cellular or tissue effects and outcomes. Three focal areas of ecological research aim to develop a sequence of approaches where "the earliest recognizable signatures of exposure" (i.e., unique patterns of up- and down-regulated genes and proteins) are identified for numerous stressors, demonstrable in case studies and incorporated into Agency, State and Regional studies supported by EMAP and other programs.
Area 1, Computational Toxicology Research: Exposure assessment has historically been based on use of chemical analysis data to generate exposure models. While biological activity of chemicals has been recognized to be important for exposure risk assessments, measurement of such activity has been limited to whole organism toxicity tests. Use of molecular approaches will:
improve extrapolation between components of source-to-outcome continuum (source , exposure , dose , effect , outcome)
Using a systems modeling approach, gene and protein expression data, in small fish models (fathead minnow and zebrafish), will be integrated with metabolomic and histopathological data. This will assist in prediction of environmental transformation and chemical effects based on structural characteristics, and enhance quantitative risk assessments, including areas of uncertainty such as a basis for extrapolation of effects of endocrine disrupting chemicals, interspecies extrapolation, complex chemical mixtures and dose-response assessment.
Area 2, Ecological Research-Environmental Diagnostics: Development of molecular diagnostic indicators contributes to several of the GPRA Diagnostic Research Goals. Methods will employ DNA microarray technology and expression proteomics, focusing on species of relevance to aquatic ecosystem risk assessment. Significantly, these diagnostic indicators will open the door to understanding subcellular interactions resulting from exposure to complex chemical mixtures.
define relationship between genetic disposition of populations and degree/specificity of stressor-specific gene transcriptional response in aquatic organisms (fish and invertebrates)
identify of chemical mixture induced transcriptional "patterns" using microarrays and hyperspectral scanning - via collaboration with DOE Sandia National Labs
apply molecular indicators to watershed level stressor study, including pilot studies with targeted pesticides and toxins indicators
develop molecular indicators of exposure for invertebrates (Daphnia, Lumbriculus, Chironomus)
Area 3, Exposure Research in Endocrine Disruptors:
Subobjective 1: Develop exposure methods, measurement protocols, and models for assessment of risk management practices of endocrine disrupting compounds. As risk management approaches are identified and developed, there will be a need to identify, adapt and develop bioassay screening tools and other analytical methods to assess their efficacy. Measurements research will be performed to define management needs. This effort will entail cross-lab participation from NRMRL, NERL and NHEERL.
Subobjective 2: Determine extent of environmental and human exposures to EDCs, characterize sources and factors influencing these exposures, develop and evaluate risk management strategies to reduce exposures. In order to develop effective risk management strategies, it is important to understand the extent of exposures to endocrine disrupting compounds and factors influencing source-to-exposure-to-dose relationships.
apply molecular indicators of exposure to estrogenic compounds in selected wastewater treatment plants located in ten USEPA Regions
identify differential gene expression following exposure of fathead minnows to environmental androgens and androgen-like compounds
apply molecular indicators of exposu
Record Details:Record Type: DOCUMENT (PRESENTATION/ABSTRACT)
Organization:U.S. ENVIRONMENTAL PROTECTION AGENCY
OFFICE OF RESEARCH AND DEVELOPMENT
NATIONAL EXPOSURE RESEARCH LAB
ECOLOGICAL EXPOSURE RESEARCH DIVISION
MOLECULAR ECOLOGY RESEARCH BRANCH