Laboratory for Ecotoxicology, Department of Environment and Agronomy, INIA-CSIC (National Institute for Agricultural Research and Food Research and Technology-CSIC), A Coruña, km 7.5. 28040 Madrid, Spain.
Laboratory for Ecotoxicology, Department of Environment and Agronomy, INIA-CSIC (National Institute for Agricultural Research and Food Research and Technology-CSIC), A Coruña, km 7.5. 28040 Madrid, Spain.
Aquat Toxicol. 2023 Aug;261:106612. doi: 10.1016/j.aquatox.2023.106612. Epub 2023 Jun 15.
The toxicity of ZnO nanoparticles (ZnO NPs) in aquatic organisms has been extensively studied, but little information is available on the effects associated with their interaction with other contaminants. In this context, the in vitro effects of co-exposure of chlorpyrifos (CPF) and ZnO NPs on fish-derived cells were investigated. A selection of concentrations was tested in single and binary exposures: CPF (0.312 - 75 mg/L) and ZnO NPs (10 - 100 mg/L). Cytotoxicity was measured using commonly used cellular endpoints: Alamar Blue/CFDA-AM for viability and plasma membrane integrity, NRU for lysosomal disruption and MTT for mitochondrial function. In addition, specific mechanisms of toxicity for CPF and ZnO NPs were tested: acetylcholinesterase (AChE) activity and ROS generation, respectively. AChE was by far the most sensitive assay for single exposure to CPF. There was no concentration-response relationship for ROS after single exposure to ZnO NPs, but 10 mg/L produced significant effects only for this cellular endpoint. Co-exposure of CPF with 10 m/L of ZnO NPs produced significant effects in almost all endpoints tested, which were enhanced by co-exposure with 100 mg/L of ZnO NPs. AChE testing of additional co-exposures with bulk ZnO, together with the application of the Independent Action (IA) prediction model, which allowed us to draw more in-depth conclusions on the toxicological behavior of the mixture. Synergism was observed at 0.625 mg/L CPF concentration and antagonism at 5 mg/L CPF in mixtures containing 100 mg/L of both ZnO NPs and bulk ZnO. However, more cases of synergism between CPF and ZnO NPs occurred at intermediate CPF concentrations, demonstrating that nano-sized particles have a more toxic interaction with CPF than bulk ZnO. Therefore it can be argued that in vitro assays allow the identification of interaction profiles of NP-containing mixtures by achieving multiple endpoints with a large number of concentration combinations.
氧化锌纳米粒子（ZnO NPs）在水生生物中的毒性已得到广泛研究，但有关其与其他污染物相互作用相关影响的信息却很少。在这种情况下，研究了毒死蜱（CPF）和 ZnO NPs 共暴露对鱼类细胞的体外影响。在单一和二元暴露中测试了一系列浓度：CPF（0.312-75 mg/L）和 ZnO NPs（10-100 mg/L）。使用常用的细胞终点测量细胞毒性：Alamar Blue/CFDA-AM 用于测定细胞活力和质膜完整性，NRU 用于测定溶酶体破裂，MTT 用于测定线粒体功能。此外，还测试了 CPF 和 ZnO NPs 的特定毒性机制：乙酰胆碱酯酶（AChE）活性和 ROS 生成。在单一 CPF 暴露中，AChE 是迄今为止最敏感的测定方法。在单一 ZnO NPs 暴露后，ROS 没有浓度-反应关系，但 10 mg/L 仅对该细胞终点产生显著影响。CPF 与 10 mg/L 的 ZnO NPs 共暴露在几乎所有测试终点都产生了显著影响，而与 100 mg/L 的 ZnO NPs 共暴露则增强了这些影响。对含有大量 ZnO 的其他共暴露进行 AChE 测试，并应用独立作用（IA）预测模型，使我们能够更深入地得出混合物的毒理学行为结论。在混合物中含有 0.625 mg/L CPF 浓度时观察到协同作用，在含有 5 mg/L CPF 浓度时观察到拮抗作用，混合物中同时含有 100 mg/L 的 ZnO NPs 和大量 ZnO。然而，在中间 CPF 浓度下，CPF 和 ZnO NPs 之间发生更多协同作用的情况表明，纳米颗粒与 CPF 的相互作用比大量 ZnO 更具毒性。因此，可以说体外测定法通过实现具有大量浓度组合的多个终点，允许鉴定含有纳米粒子的混合物的相互作用谱。
