CIIMAR-Interdisciplinary Centre of Marine and Environmental Research, Endocrine Disruptors and Emerging Contaminants Group, University of Porto, Avenida General Norton de Matos, S/N, 4450-208 Matosinhos, Portugal; CITAB - Centre for the Research and Technology of Agro-Environmental and Biological Sciences, University of Trás-os-Montes and Alto Douro (UTAD), Quinta de Prados, Pavilhão 2, 5000-801 Vila Real, Portugal.
CIIMAR-Interdisciplinary Centre of Marine and Environmental Research, Endocrine Disruptors and Emerging Contaminants Group, University of Porto, Avenida General Norton de Matos, S/N, 4450-208 Matosinhos, Portugal.
Sci Total Environ. 2022 Nov 10;846:157361. doi: 10.1016/j.scitotenv.2022.157361. Epub 2022 Jul 14.
Metformin (MET), an anti-diabetic pharmaceutical of large-scale consumption, is increasingly detected in surface waters. However, current knowledge on the long-term effects of MET on non-target organisms is limited. The present study aimed to investigate the effects of MET in the model freshwater teleost Danio rerio, following a full life-cycle exposure to environmentally relevant concentrations (390 to 14 423 ng/L). Considering that the mode of action (MoA) of MET on non-target organisms remains underexplored and that MET may act through similar human pathways, i.e., lipid and energy metabolisms, biochemical markers were used to determine cholesterol and triglycerides levels, as well as mitochondrial complex I activity in zebrafish liver. Also, the hepatosomatic index as an indication of metabolic disruption, and the expression levels of genes involved in MET's putative MoA, i.e. acaca, acadm, cox5aa, idh3a, hmgcra, prkaa1, were determined, the last by qRT-PCR. A screening of mRNA transcripts, associated with lipid and energy metabolisms, and other signaling pathways potentially involved in MET-induced toxicity were also assessed using an exploratory RNA-seq analysis. The findings here reported indicate that MET significantly disrupted critical biochemical and molecular processes involved in zebrafish metabolism, such as cholesterol and fatty acid biosynthesis, mitochondrial electron transport chain and tricarboxylic acid cycle, concomitantly to changes on the hepatosomatic index. Likewise, MET impacted other relevant pathways mainly associated with cell cycle, DNA repair and steroid hormone biosynthesis, here reported for the first time in a non-target aquatic organism. Non-monotonic dose response curves were frequently detected in biochemical and qRT-PCR data, with higher effects observed at 390 and 2 929 ng/L MET treatments. Collectively, the results suggest that environmentally relevant concentrations of MET severely disrupt D. rerio metabolism and other important biological processes, supporting the need to revise the proposed environmental quality standard (EQS) and predicted no-effect concentration (PNEC) for MET.
二甲双胍（MET）是一种大规模消费的抗糖尿病药物，越来越多地在地表水中被检测到。然而，目前对 MET 对非靶标生物的长期影响的了解有限。本研究旨在研究环境相关浓度（390 至 14423ng/L）下全生命周期暴露对模式淡水硬骨鱼斑马鱼的影响。由于 MET 对非靶标生物的作用模式（MoA）仍未得到充分探索，并且 MET 可能通过类似的人类途径（即脂质和能量代谢）起作用，因此使用生化标志物来确定胆固醇和甘油三酯水平，以及斑马鱼肝线粒体复合物 I 活性。此外，还测定了肝体比作为代谢紊乱的指标，以及参与 MET 假定 MoA 的基因的表达水平，即 acaca、acadm、cox5aa、idh3a、hmgcra、prkaa1，最后通过 qRT-PCR 进行测定。还使用探索性 RNA-seq 分析评估了与脂质和能量代谢以及其他可能与 MET 诱导的毒性相关的信号通路相关的 mRNA 转录物的筛选。这里报道的研究结果表明，MET 显著破坏了斑马鱼代谢中涉及的关键生化和分子过程，如胆固醇和脂肪酸生物合成、线粒体电子传递链和三羧酸循环，同时肝体比也发生了变化。同样，MET 还影响了其他与细胞周期、DNA 修复和类固醇激素生物合成相关的重要途径，这是首次在非靶标水生生物中报道。在生化和 qRT-PCR 数据中经常检测到非单调剂量反应曲线，在 390 和 2929ng/L MET 处理中观察到更高的影响。总的来说，这些结果表明，环境相关浓度的 MET 严重破坏了 D. rerio 的新陈代谢和其他重要的生物过程，支持修订 MET 的建议环境质量标准（EQS）和预测无影响浓度（PNEC）的必要性。
