Pesticides, which are used in agriculture and forestry to eliminate insects, are a major cause of environmental pollution. Among them, diflubenzuron (DFB), 1-(4-chlorophenyl)-3-(2,6-difluorobenzoyl) urea, is a common benzoylurea insecticide that hinders larval development, primarily in Aedes aegypti larvae. Many experts have announced the biological toxicity of DFB in various species. However, the toxicity of benzoylurea pesticides, including DFB, to bovine mammary epithelial cells (MAC-T) is unclear. Therefore, in this study, we confirmed the cytotoxic effects of DFB on the viability and proliferation of MAC-T cells. Additionally, we observed that DFB induced lipid peroxidation through reactive oxygen species (ROS) production, resulting in an increase in transcriptional gene expression related to inflammatory response. Moreover, we demonstrated mitochondrial dysfunction including depolarization of the mitochondrial membrane, perturbation of calcium homeostasis, and, eventually, apoptosis. Furthermore, we identified DFB-triggered signaling pathways related to ROS generation and cell proliferation, as well as their interactions, by treating the cells with pharmacological inhibitors in combination with DFB. DFB attenuated the phosphorylation of AKT, P70S6K, S6, and ERK1/2 and facilitated the phosphorylation of JNK and c-Jun. These results show that DFB can induce apoptotic cell death via ROS generation and mitochondrial dysfunction in MAC-T cells.

在农业和林业中用于消灭昆虫的农药是造成环境污染的主要原因。其中，除虫脲(DFB)，1-(4-氯苯基)-3-(2,6-二氟苯甲酰)脲，是一种常见的苯甲酰脲类杀虫剂，主要是埃及伊蚊，阻碍幼虫发育。幼虫。许多专家已经公布了DFB在不同物种中的生物毒性。然而，苯甲酰脲类农药（包括 DFB）对牛乳腺上皮细胞 (MAC-T) 的毒性尚不清楚。因此，在本研究中，我们证实了 DFB 对 MAC-T 细胞活力和增殖的细胞毒性作用。此外，我们观察到 DFB 通过产生活性氧 (ROS) 诱导脂质过氧化，导致与炎症反应相关的转录基因表达增加。此外，我们证明了线粒体功能障碍，包括线粒体膜的去极化、钙稳态的扰动，以及最终的细胞凋亡。此外，我们确定了与 ROS 生成和细胞增殖相关的 DFB 触发的信号通路，以及它们的相互作用，通过用药物抑制剂与 DFB 组合处理细胞。DFB 减弱 AKT、P70S6K、S6 和 ERK1/2 的磷酸化并促进 JNK 和 c-Jun 的磷酸化。这些结果表明DFB可以诱导细胞凋亡通过MAC-T 细胞中的 ROS 生成和线粒体功能障碍。