Abstract：
Neurotoxic organophosphorus compounds can induce a type of delayed neuropathy in humans and sensitive animals, known as organophosphorus-induced delayed neuropathy (OPIDN). OPIDN is characterized by axonal degeneration akin to Wallerian-like degeneration, which is thought to be caused by increased intra-axonal Ca2+ concentrations. This study was designed to investigate that deregulated cytosolic Ca2+ may function downstream of mitodysfunction in activating Wallerian-like degeneration and necroptosis in OPIDN. Adult hens were administrated a single dosage of 750 mg/kg tri-ortho-cresyl phosphate (TOCP), and then sacrificed at 1 day, 5 day, 10 day and 21 day post-exposure, respectively. Sciatic nerves and spinal cords were examined for pathological changes and proteins expression related to Wallerian-like degeneration and necroptosis. In vitro experiments using differentiated neuro-2a (N2a) cells were conducted to investigate the relationship among mitochondrial dysfunction, Ca2+ influx, axonal degeneration, and necroptosis. The cells were co-administered with the Ca2+-chelator BAPTA-AM, the TRPA1 channel inhibitor HC030031, the RIPK1 inhibitor Necrostatin-1, and the mitochondrial-targeted antioxidant MitoQ along with TOCP. Results demonstrated an increase in cytosolic calcium concentration and key proteins associated with Wallerian degeneration and necroptosis in both in vivo and in vitro models after TOCP exposure. Moreover, co-administration with BATPA-AM or HC030031 significantly attenuated the loss of NMNAT2 and STMN2 in N2a cells, as well as the upregulation of SARM1, RIPK1 and p-MLKL. In contrast, Necrostatin-1 treatment only inhibited the TOCP-induced elevation of p-MLKL. Notably, pharmacological protection of mitochondrial function with MitoQ effectively alleviated the increase in intracellular Ca2+ following TOCP and mitigated axonal degeneration and necroptosis in N2a cells, supporting mitochondrial dysfunction as an upstream event of the intracellular Ca2+ imbalance and neuronal damage in OPIDN. These findings suggest that mitochondrial dysfunction post-TOCP intoxication leads to an elevated intracellular Ca2+ concentration, which plays a pivotal role in the initiation and development of OPIDN through inducing SARM1-mediated axonal degeneration and activating the necroptotic signaling pathway.
摘要：
神经毒性有机磷化合物可在人类和敏感动物中诱发一种迟发性神经病,称为有机磷引起的迟发性神经病（OPIDN）。OPIDN的特征是轴突变性类似于Wallerian样变性，这被认为是由轴突内Ca2+浓度增加引起的。这项研究旨在研究失调的胞浆Ca2可能在OPIDN的线粒体功能障碍下游激活Wallerian样变性和坏死。成年母鸡服用单剂量750mg/kg磷酸三邻甲苯酯（TOCP），然后在1天牺牲，5天,暴露后10天和21天，分别。检查坐骨神经和脊髓的病理变化以及与Wallerian样变性和坏死相关的蛋白表达。使用分化的神经-2a（N2a）细胞进行体外实验，以研究线粒体功能障碍之间的关系，Ca2+流入，轴突变性,和坏死。细胞与Ca2+-螯合剂BAPTA-AM共同施用,TRPA1通道抑制剂HC030031，RIPK1抑制剂Necrostatin-1和线粒体靶向抗氧化剂MitoQ以及TOCP。结果表明，在TOCP暴露后，在体内和体外模型中，胞质钙浓度和与Wallerian变性和坏死相关的关键蛋白均增加。此外，与BATPA-AM或HC030031共同施用可显着减弱N2a细胞中NMNAT2和STMN2的损失，以及SARM1、RIPK1和p-MLKL的上调。相比之下,Necrostatin-1治疗仅抑制TOCP诱导的p-MLKL升高。值得注意的是,线粒体功能的药理学保护与MitoQ有效缓解细胞内Ca2+的增加后TOCP和减轻轴突变性和坏死N2a细胞，支持线粒体功能障碍作为OPIDN细胞内Ca2+失衡和神经元损伤的上游事件。这些发现表明，线粒体功能障碍后TOCP中毒导致细胞内Ca2+浓度升高，通过诱导SARM1介导的轴突变性和激活坏死信号通路，在OPIDN的发生和发展中起关键作用。
