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PROTEOMICS IN ECOTOXICOLOGY: PROTEIN EXPRESSION PROFILING TO SCREEN CHEMICALS FOR ENDOCRINE ACTIVITY
Citation:
SALINAS, K., C. C. WALKER, S. S. WILKINSON, J. WATTS, J. T. WINSTEAD, P. S. HARRIS, AND M. J. HEMMER. PROTEOMICS IN ECOTOXICOLOGY: PROTEIN EXPRESSION PROFILING TO SCREEN CHEMICALS FOR ENDOCRINE ACTIVITY. Presented at SETAC Meeting, Baltimore, MD, November 13 - 17, 2005.
Description:
Abstract for poster.
Current endocrine testing methods are animal intensive and lack the throughput necessary to screen large numbers of environmental chemicals for adverse effects. In this study, Matrix Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry MALDI-TOF MS) coupled with a short term fish assay was used to investigate plasma expression protein profiling as a means to screen chemicals for estrogenic activity. Adult male sheepshead minnows (Cyprinodon variegatus) were placed into aquaria consisting of duplicate tanks for vehicle control and the following estrogen agonist treatments: 0.05, 0.1, 0.2, 0.5 and 1.0 µg 17ß-estradiol/L; 6 and 12 µg methoxychlor/L; and 100 and 1000 ug bisphenol-A/L. Treatments with 80 µg chlorpyrifos/L and 0.6 µg endosulfan/L served as non-estrogenic negative controls. Test concentrations were maintained using an intermittent flow-through dosing apparatus supplying exposure water at 20 L/hour. Fish were sampled at 7 or 10 days, plasma diluted, applied to a Ciphergen Biosystems Inc. Gold Chip array and analyzed. Multiple protein peaks, ranging from 2.5-3.5 KDa, were identified as markers of estrogenic effects when comparing estrogen-treated and control fish. A binary classification model was constructed from plasma protein profiles of the vehicle control and 0.1 µg/L 17ß-estradiol treatments and used to evaluate all samples. Self validation of the model was performed using all 17ß-estradiol treatments. The predictive model exhibited 100% sensitivity, specificity and predictive value for all treatments and concentrations used. Treatments with the estrogen agonists 17ß-estradiol, methoxychlor and bisphenol-A generated reproducible diagnostic biomarkers based on the presence of specific estrogen-responsive plasma proteins. However, the controls and non-estrogenic compounds chlorpyrifos and endosulfan did not produce this estrogen responsive protein profile. These results indicate that protein expression profiling can be a useful tool to screen chemicals for mode of action.