Indiscriminate overuse or occupational exposure to agricultural chemicals can lead to neurotoxicity. Many pesticides act to impair mitochondrial function which can lead to exacerbation of neurodegeneration. Triazole fungicides are applied to grain, fruit, and vegetable crops to combat mold and fungi and their use is increasing worldwide. Here, we assessed the in vitro toxicity of two widely used triazole fungicides, propiconazole and tebuconazole, to mitochondria using differentiated SH-SY5Y neuroblastoma cells as an in vitro cell model used in Parkinson’s disease research. Cell viability (based on ATP levels), mitochondrial membrane potential, oxidative respiration, and reactive oxygen species (ROS) were measured following fungicide treatments. Cell viability was decreased with 100 μM propiconazole after 24 and 48 h, while tebuconazole required higher doses to affect viability (−200 μM at 24 h). Mitochondrial membrane potential (MMP) was reduced with 50 μM propiconazole after 24 h while 200 μM tebuconazole reduced MMP. Oxidative respiration of SH-SY5Y cells was then measured using a XFe24 Flux analyzer and 100 μM propiconazole reduced basal respiration, oligomycin-induced ATP production, and FCCP-induced maximum respiration by −40−50%, while tebuconazole did not affect mitochondrial bioenergetics at the concentrations tested. Acute exposure to 100 μM propiconazole over 4 h did not immediately affect oxidative respiration in SH-SY5Y cells. ROS were not induced by propiconazole and tebuconazole up to 100 and 300 μM respectively. Based on these results, we focused our lipidomics investigations on SH-SY5Y exposed only to propiconazole, as lipid dysregulation is associated with mitochondrial dysfunction. Both 50 and 100 μM propiconazole altered the abundance of some ceramides, specifically reducing glucosylceramide non-hydroxyfatty acid-sphingosine (HexCer-NS) and increasing N-stearoyl-phytosphingosine (CerNP). Moreover, a recently discovered bioactive lipid called fatty acid ester of hydroxy fatty acid (FAHFA) was increased 5-fold, hypothesized to be a neuroprotective mechanism that has been demonstrated in other studies of human diseases. Additional lipids reduced in abundance included oxidized phosphatidylcholine (OxPC) and oxidized phosphatidylethanolamine (OxPE). There were no changes in cellular triacylglycerols nor total lipids with exposure to propiconazole. Taken together, this study provides insight into the toxicity of triazole fungicides in neuronal cells, which has implications for neurodegenerative diseases that involve the mitochondria such as Parkinson’s disease.

不加选择的过度使用或职业性接触农药会导致神经毒性。许多农药会破坏线粒体功能，从而导致神经退行性恶化。三唑类杀菌剂应用于谷物，水果和蔬菜作物，以对抗霉菌和真菌，并且在世界范围内其使用量正在增加。在这里，我们评估了在体外两种广泛使用的三唑类杀菌剂，丙环唑和戊唑醇的毒性，使用线粒体分化SH-SY5Y成神经瘤细胞作为体外细胞模型用于帕金森氏病研究。杀菌剂处理后，测量细胞活力（基于ATP含量），线粒体膜电位，氧化呼吸和活性氧（ROS）。24和48小时后，使用100μM的丙环唑降低细胞活力，而戊唑醇需要更高的剂量才能影响活力（24小时为-200μM）。24 h后，用50μM丙环唑降低线粒体膜电位（MMP），而200μM戊唑醇则降低MMP。然后使用XFe24通量分析仪测量SH-SY5Y细胞的氧化呼吸，100μM丙环唑使基础呼吸，寡霉素诱导的ATP产生和FCCP诱导的最大呼吸降低-40％至50％，而戊唑醇并不影响线粒体的生物能。测试的浓度。急性暴露于100μM丙环唑4小时并没有立即影响SH-SY5Y细胞的氧化呼吸。分别高达100和300μM的丙环唑和戊唑醇未诱导ROS。基于这些结果，我们将脂质组学的研究重点放在仅暴露于丙环唑的SH-SY5Y上，因为脂质失调与线粒体功能障碍有关。50和100μM的丙环唑都改变了一些神经酰胺的丰度，特别是降低了葡萄糖基神经酰胺非羟基脂肪酸-鞘氨醇（HexCer-NS）和增加了N-硬脂酰-植物鞘氨醇（CerNP）。此外，最近发现的称为羟基脂肪酸脂肪酸酯（FAHFA）的生物活性脂质增加了5倍，据推测是一种神经保护机制，已在其他人类疾病研究中得到证实。大量减少的其他脂质包括氧化磷脂酰胆碱（OxPC）和氧化磷脂酰乙醇胺（OxPE）。丙环唑暴露后细胞三酰基甘油和总脂质均无变化。两者合计，这项研究提供了对三唑类杀菌剂在神经元细胞中毒性的见解，这对涉及线粒体的神经退行性疾病（如帕金森氏病）有影响。