School of Marine and Atmospheric Sciences, Stony Brook University, Stony Brook, NY 11794, USA.
Aquat Toxicol. 2012 Feb;108:60-9. doi: 10.1016/j.aquatox.2011.10.017. Epub 2011 Nov 3.
Fish embryos were used to evaluate the interaction among common environmental and chemical stressors found in urban coastal environments, namely hypoxia, aryl hydrocarbon receptor (AhR) agonists, and estrogenic compounds. At the molecular level, the systems responding to these stressors share common response factors, and evidence exists for cross-talk between them. Biomarkers of exposure to these stressors, cytochrome P4501a (Cyp1a), estrogen receptor alpha (ERα), brain cytochrome P450 aromatase (Cyp19a2 or AromB), and hypoxia inducible factor 1 alpha (Hif-1α) mRNA expression were examined using qRT-PCR simultaneously in embryos of two well studied species, the Atlantic killifish, Fundulus heteroclitus, and the zebrafish Danio rerio. Embryos of both species were exposed to the model Cyp1a inducer β-naphthoflavone (BNF) or 17-β estradiol (E2) under either normoxic or hypoxic (5% oxygen atmosphere) conditions and harvested prior to hatch at 9 days post fertilization (dpf) for the killifish, and 48h post fertilization (hpf) for the zebrafish. BNF significantly induced Cyp1a expression in embryos of both species with killifish embryos being more responsive (700-fold>control) than zebrafish embryos (7-100-fold>control). AromB was also significantly influenced by treatment, but to a lesser extent, with mean expression levels increased by less than two-fold over control values in response to E2, and in one case upregulated by BNF. ERα and Hif-1α were constitutively expressed in embryos of both species, but expression was unaffected by exposure to either BNF or E2. Hypoxic conditions downregulated AromB expression strongly in killifish but not in zebrafish embryos. The impact of hypoxia on expression of other genes in either species was inconsistent, although an interactive effect between hypoxia and BNF on several of the genes evaluated was observed. These data are the first to examine expression patterns of these important environmental response genes together in embryos of two important model fish species. The results support the use of Cyp1a expression as a biomarker of AhR agonists in fish embryos, and indicate that AromB may be more responsive than ERα to estrogenic chemicals at this stage in development. Hif-1α expression was not found to be a good biomarker of hypoxic exposure in either killifish or zebrafish embryos. The interaction observed between BNF and co-exposure to hypoxia warrants further investigation. Finally killifish embryos are generally more sensitive than zebrafish embryos at this stage of development supporting their use in environmental assessments.
利用鱼类胚胎评估城市沿海环境中常见的环境和化学应激源之间的相互作用，这些应激源包括缺氧、芳香烃受体（AhR）激动剂和雌激素化合物。在分子水平上，对这些应激源作出反应的系统共享共同的反应因子，并且存在它们之间相互作用的证据。使用 qRT-PCR 同时检测两种研究较多的物种，即大西洋鲦鱼（Fundulus heteroclitus）和斑马鱼（Danio rerio）胚胎中这些应激源的暴露生物标志物，细胞色素 P4501a（Cyp1a）、雌激素受体α（ERα）、脑细胞色素 P450 芳香化酶（Cyp19a2 或 AromB）和缺氧诱导因子 1α（Hif-1α）mRNA 表达。两种物种的胚胎均在正常氧（21%氧气）或缺氧（5%氧气）条件下暴露于模型 Cyp1a 诱导剂β-萘黄酮（BNF）或 17-β 雌二醇（E2）下，然后在受精后 9 天（killifish）或受精后 48 小时（zebrafish）收获，以进行孵化。BNF 显著诱导两种物种的 Cyp1a 表达，其中鲦鱼胚胎的反应更敏感（700 倍>对照），而斑马鱼胚胎的反应（7-100 倍>对照）。AromB 也受到处理的显著影响，但程度较小，E2 处理下平均表达水平仅比对照值增加不到两倍，而 BNF 处理下增加了一个。ERα 和 Hif-1α 在两种物种的胚胎中均持续表达，但暴露于 BNF 或 E2 均不影响其表达。缺氧条件强烈下调鲦鱼胚胎中的 AromB 表达，但不影响斑马鱼胚胎。在这两种物种中，缺氧对其他基因表达的影响不一致，尽管观察到在评估的几个基因中存在缺氧与 BNF 之间的相互作用。这些数据首次在两种重要的模式鱼类胚胎中同时检查这些重要的环境反应基因的表达模式。结果支持将 Cyp1a 表达作为鱼类胚胎中 AhR 激动剂的生物标志物，并表明在该发育阶段，AromB 可能比 ERα 对雌激素化学物质更敏感。Hif-1α 表达未被发现是鲦鱼或斑马鱼胚胎缺氧暴露的良好生物标志物。在 BNF 与共暴露于缺氧之间观察到的相互作用需要进一步研究。最后，在这个发育阶段，鲦鱼胚胎通常比斑马鱼胚胎更敏感，支持将其用于环境评估。
