Department of Biological Sciences, University of Bergen, Bergen, Norway.
Computational Biology Unit, Department of Informatics, University of Bergen, Bergen, Norway.
Aquat Toxicol. 2018 Aug;201:174-186. doi: 10.1016/j.aquatox.2018.06.003. Epub 2018 Jun 7.
Polycyclic aromatic hydrocarbons such as benzo[a]pyrene (BaP) that activate the aryl hydrocarbon receptor (Ahr) pathway, and endocrine disruptors acting through the estrogen receptor pathway are among environmental pollutants of major concern. In this work, we exposed Atlantic cod (Gadus morhua) precision-cut liver slices (PCLS) to BaP (10 nM and 1000 nM), ethynylestradiol (EE2) (10 nM and 1000 nM), and equimolar mixtures of BaP and EE2 (10 nM and 1000 nM) for 48 h, and performed RNA-Seq based transcriptome mapping followed by systematic bioinformatics analyses. Our gene expression analysis showed that several genes were differentially expressed in response to BaP and EE2 treatments in PCLS. Strong up-regulation of genes coding for the cytochrome P450 1a (Cyp1a) enzyme and the Ahr repressor (Ahrrb) was observed in BaP treated PCLS. EE2 treatment of liver slices strongly up-regulated genes coding for precursors of vitellogenin (Vtg) and eggshell zona pellucida (Zp) proteins. As expected, pathway enrichment and network analysis showed that the Ahr and estrogen receptor pathways are among the top affected by BaP and EE2 treatments, respectively. Interestingly, two genes coding for fibroblast growth factor 3 (Fgf3) and fibroblast growth factor 4 (Fgf4) were up-regulated by EE2 in this study. To our knowledge, the fgf3 and fgf4 genes have not previously been described in relation to estrogen signaling in fish liver, and these results suggest the modulation of the FGF signaling pathway by estrogens in fish. The signature expression profiles of top differentially expressed genes in response to the single compound (BaP or EE2) treatment were generally maintained in the expression responses to the equimolar binary mixtures. However, in the mixture-treated groups, BaP appeared to have anti-estrogenic effects as observed by lower number of differentially expressed putative EE2 responsive genes. Our in-depth quantitative analysis of changes in liver transcriptome in response to BaP and EE2, using PCLS tissue culture provides further mechanistic insights into effects of the compounds. Moreover, the analyses demonstrate the usefulness of PCLS in cod for omics experiments.
多环芳烃，如苯并[a]芘（BaP），激活芳香烃受体（Ahr）途径，以及通过雌激素受体途径发挥作用的内分泌干扰物，是主要关注的环境污染物之一。在这项工作中，我们将大西洋鳕鱼（Gadus morhua）的精密肝切片（PCLS）暴露于 BaP（10 nM 和 1000 nM）、乙炔雌二醇（EE2）（10 nM 和 1000 nM）和 BaP 和 EE2 的等摩尔混合物（10 nM 和 1000 nM）中 48 小时，并进行了基于 RNA-Seq 的转录组图谱绘制和系统的生物信息学分析。我们的基因表达分析表明，PCLS 中几种基因对 BaP 和 EE2 处理有差异表达。在 BaP 处理的 PCLS 中，观察到编码细胞色素 P450 1a（Cyp1a）酶和 Ahr 抑制剂（Ahrrb）的基因强烈上调。EE2 处理肝切片强烈上调卵黄蛋白原（Vtg）和卵壳透明带（Zp）蛋白前体的基因编码。正如预期的那样，途径富集和网络分析表明，Ahr 和雌激素受体途径分别是受 BaP 和 EE2 处理影响最大的途径。有趣的是，在这项研究中，两个编码成纤维细胞生长因子 3（Fgf3）和成纤维细胞生长因子 4（Fgf4）的基因被 EE2 上调。据我们所知，在鱼类肝脏中，fgf3 和 fgf4 基因以前没有与雌激素信号相关的描述，这些结果表明雌激素对鱼类中 FGF 信号通路的调节。对单一化合物（BaP 或 EE2）处理的差异表达基因的签名表达谱通常保持在对等摩尔二元混合物的表达反应中。然而，在混合物处理组中，BaP 似乎表现出抗雌激素作用，因为具有不同表达的推定 EE2 反应基因的数量较少。我们使用 PCLS 组织培养对 BaP 和 EE2 引起的肝转录组变化进行了深入的定量分析，为这些化合物的作用提供了进一步的机制见解。此外，分析表明 PCLS 在鳕鱼中的组学实验中是有用的。
