Citation:

Bencic, D C., D. C. Volz, P. Chen, A Biales, J M. Lazorchak, D. E. Hinton, AND S. W. Kullman. ALTERED GENE EXPRESSION AND DEVELOPMENT OF POTENTIAL BIOMARKERS IN MEDAKA (ORYZIAS LATIPES) BRAIN, LIVER AND TESTIS FOLLOWING EXPOSURE TO FIBRATE PHARMACEUTICALS. Presented at Society of Toxicology, New Orleans, LA, March 06 - 10, 2005.

Impact/Purpose:

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

Description:

To help address the consequences of increasing levels of environmental contaminants and to identify potentially novel markers of toxicity, we examined gene expression profiles from medaka (Oryzias latipes) exposed to a prototypical fibrate pharmaceutical. Changes in gene expression in male medaka brain, liver and testis following 48 h exposure to 15 ug (injected) of the antihyperlipidemic agent ciprofibrate (CF) were analyzed using suppressive subtractive hydridization. From a total of 1152 clones, 288 were sequenced and 100 were identified as differentially expressed. Furthermore, only one gene was shared among all three organs and less than 5% between any two organs, suggesting organ specific responses at the level of gene expression. The results of this CF and other xenobiotic induced gene expression studies were used to develop macroarrays specific for brain, liver or testis containing 200-300 non-redundant cDNAs. Following 7 day water exposures to 10, 100 and 1000 ug/l of CF or clofibrate, compound, dose and organ specific altered gene expression was determined. Highly responsive genes were further validated using real-time QPCR. In order to associate changes in gene expression with altered phenotype, compound- and dose-dependent histopathology samples were also examined. A combination of macroarray, real-time QPCR and histopathological techniques will aid in linking molecular events and biochemical alterations following exposure and provide a mechanism to evaluate exposure and effects at the individual and population levels.

Record Details: