Hook Sharon E, Skillman Ann D, Small Jack A, Schultz Irvin R
Battelle Pacific Northwest Division, Sequim, WA 98382, USA.
Mar Environ Res. 2006 Jul;62 Suppl(Suppl):S151-5. doi: 10.1016/j.marenvres.2006.04.056. Epub 2006 Apr 18.
Determining how gene expression profiles change with toxicant dose will improve the utility of arrays in identifying biomarkers and modes of toxic action. Isogenic rainbow trout, Oncorhyncus mykiss,were exposed to 10, 50 or 100 ng/L ethynylestradiol (a xeno-estrogen) for 7 days. Following exposure hepatic RNA was extracted. Fluorescently labeled cDNA were generated and hybridized against a commercially available Atlantic Salmon/Trout array (GRASP project, University of Victoria) spotted with 16,000 cDNAs. Transcript expression in treated vs control fish was analyzed via Genespring (Silicon Genetics) to identify genes with altered expression, as well as to determine gene clustering patterns that can be used as "expression signatures". Array results were confirmed via qRT PCR. Our analysis indicates that gene expression profiles varied somewhat with dose. Established biomarkers of exposure to estrogenic chemicals, such as vitellogenin, vitelline envelope proteins, and the estrogen receptor alpha, were induced at every dose. Other genes were dose specific, suggesting that different doses induce distinct physiological responses. These findings demonstrate that cDNA microarrays could be used to identify both toxicant class and relative dose.
确定基因表达谱如何随毒物剂量变化，将提高阵列在识别生物标志物和毒性作用模式方面的效用。将同基因的虹鳟（Oncorhyncus mykiss）暴露于10、50或100 ng/L乙炔雌二醇（一种外源性雌激素）中7天。暴露后提取肝脏RNA。生成荧光标记的cDNA，并与点有16,000个cDNA的市售大西洋鲑鱼/鳟鱼阵列（维多利亚大学GRASP项目）杂交。通过Genespring（Silicon Genetics）分析处理组与对照组鱼的转录本表达，以鉴定表达改变的基因，并确定可用作“表达特征”的基因聚类模式。通过qRT PCR确认阵列结果。我们的分析表明，基因表达谱随剂量略有变化。在每个剂量下均诱导了暴露于雌激素化学物质的既定生物标志物，如卵黄蛋白原、卵黄膜蛋白和雌激素受体α。其他基因具有剂量特异性，表明不同剂量诱导不同的生理反应。这些发现表明，cDNA微阵列可用于识别毒物类别和相对剂量。
