Screening has revealed that aquafeeds with high inclusion of plant material may contain small amounts of endocrine disrupting agricultural pesticides. In this work, bisphenol A (BPA) and genistein (GEN) were selected as model endocrine disrupting toxicants with impact on DNA methylation in fish. Atlantic salmon hepatocytes were exposed in vitro to four concentrations of BPA and GEN (0.1, 1.0, 10 and 100 μM) for 48 h. Toxicity endpoints included cytotoxicity, global DNA methylation, targeted transcriptomics and metabolomic screening (100 μM). GEN was not cytotoxic in concentrations up to 100 μM, whereas one out of two cell viability assays indicated a cytotoxic response to 100 μM BPA. Compared to the control, significant global DNA hypomethylation was observed at 1.0 μM BPA. Both compounds upregulated cyp1a1 transcription at 100 μM, while estrogenic markers esr1 and vtg1 responded strongest at 10 μM Dnmt3aa transcription was downregulated by both compounds at 100 μM. Metabolomic screening showed that BPA and GEN resulted in significant changes in numerous biochemical pathways consistent with alterations in carbohydrate metabolism, indicating perturbation in glucose homeostasis and energy generation, and glutamate metabolism. Pathway analysis showed that while the superpathway of methionine degradation was among the most strongly affected pathways by BPA, GEN induced changes to uridine and pyrimidine biosynthesis. In conclusion, this mechanistic study proposes metabolites associated with glucose and glutamate metabolism, glucuronidation detoxification, as well as transcriptional markers cyp1a1, vtg1, esr1, ar, dnmt3aa, cdkn1b and insig1 as markers for BPA and GEN exposure in fish liver cells.

中文翻译：

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大西洋鲑鱼肝细胞中双酚A和染料木黄酮的代谢特征

筛选显示，含有高含量植物原料的水产饲料可能含有少量破坏内分泌的农用农药。在这项工作中，双酚A（BPA）和染料木黄酮（GEN）被选作模型内分泌干扰物，对鱼类DNA甲基化有影响。大西洋鲑鱼肝细胞在体外暴露于四种浓度的BPA和GEN（0.1、1.0、10和100μM）48小时。毒性终点包括细胞毒性，总体DNA甲基化，靶向转录组学和代谢组学筛选（100μM）。GEN浓度高达100μM时无细胞毒性，而两种细胞活力分析中有一项表明对100μMBPA有细胞毒性反应。与对照相比，在1.0μMBPA下观察到明显的总体DNA低甲基化。两种化合物均上调CYP1A1转录为100微米，而雌激素标志物ESR1和vtg1回应最强在10μM Dnmt3aa两种化合物都以100μM下调了转录。代谢组学筛选显示，BPA和GEN导致许多生物化学途径的显着变化，与碳水化合物代谢的变化一致，表明葡萄糖稳态和能量产生以及谷氨酸代谢受到干扰。途径分析表明，尽管甲硫氨酸降解的超级途径是受BPA影响最大的途径之一，但GEN诱导了尿苷和嘧啶生物合成的变化。总之，该机制研究提出了与葡萄糖和谷氨酸代谢，葡萄糖醛酸化解毒有关的代谢物，以及转录标记cyp1a1，vtg1，esr1，ar，dnmt3aa，cdkn1b和insig1作为鱼肝细胞中BPA和GEN暴露的标志物。