PARENT SESSION

PH14 Ecotoxicogenomics
Exhibit Hall
8:00 AM - Thursday

(PH156) Plasma protein-profiling as a high throughput tool for chemical screening using a small fish model.

Hudson, Robert¹, Hemmer, Michael¹, Salinas, Kimberly¹, Wilkinson, Sherry ¹, Watts, James¹, Winstead, James¹, Harris, Peggy ¹, Kirkpatrick, Amy ¹, Walker, Calvin ¹, ¹ EPA/ORD/NHEERL/Gulf Ecology Division, Gulf Breeze, FL, USA

ABSTRACT- Current endocrine testing methods are animal intensive and lack the rapid throughput necessary to screen large numbers of environmental chemicals for adverse effects. Surface Enhanced Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (SELDI-TOF MS) was used to investigate plasma protein profiling as a possible means for screening chemical inventories for pathway specific toxicity using a small fish model. Protein profiling is a technique for identifying protein biomarker fingerprints indicative of a ′disease′ state when compared with a ′normal′ state. Comparably sized adult male sheepshead minnows, (Cyprinodon variegatus), were placed into flow-through aquaria (30 fish each) consisting of duplicate tanks for seawater control, vehicle control, and 17-estradiol treatments of 50 ng/mL and 200 ng/mL. Test concentrations were maintained by intermittent seawater flow and injection of test stock solutions for the chemical and carrier control treatments. Fish were sampled at 7 days for plasma and tissues. The plasma was applied to ProteinChip© arrays, and the arrays were analyzed using a SELDI-TOF instrument. Estrogen specific spectral fingerprints were identified using pattern recognition software. No difference was found between protein profiles of seawater control and carrier control (triethylene glycol) treated fish. A number of protein peaks, ranging from 1-25 kDa, were identified as markers of estrogenic effects when comparing estrogen treated and control fish. We show that an estrogenic compound produces distinct biomarkers, based on the presence or absence of plasma proteins. We are also investigating protein patterns of other compounds, representing receptor-mediated and non-receptor mediated pathways of estrogenic toxicity. Comparison of these protein signatures to 17-estradiol will be accomplished using a predictive estrogen-responsive pattern recognition model. Although the patterns generated will be specific to the species tested, the protocol should be readily transferrable to any species or chemical toxicity mechanism of interest.

Key words: EDC, SELDI, protein-profiling, sheepshead minnow