Abstract

Styrene 7,8-oxide (SO) is the principal metabolite of styrene, an industrial neurotoxic compound which causes various neurodegenerative disorders. The present study aimed to explore the mechanisms of SO cytotoxicity (0.5 − 4 mM) in primary cortical neurons and to evaluate the neuroprotective potential of quercetin (QUER). Our results showed that exposure to SO decreased viability of cortical neurons in a concentration-dependent manner. In the presence of QUER, cell viability was increased significantly. The neuroprotective effects of QUER were associated with the reduction of intracellular Reactive Oxygen Species (ROS), the decrease in calcium overload and the restoration of mitochondrial membrane depolarization caused by SO. Additionally, to evaluate neuronal death mechanisms triggered by SO, cells were incubated with Ac-DEVD-CHO, Calpeptin and Necrostatin-1, pharmacological inhibitors of caspase-3, calpains and necroptosis respectively. The data showed that the three inhibitors reduced cell death induced by SO and suggested the implication of apoptotic, necrotic and necroptotic pathways. However, western blot analysis showed that QUER attenuated the activation of caspase-3 but did not prevent calpain activity. Taken together, these data indicated that the cytotoxicity of SO was mediated by oxidative stress and apoptosis, necrosis and necroptosis mechanisms, while the neuroprotection provided by QUER against SO depended mainly on its anti-apoptotic activity.

摘要

苯乙烯 7,8-氧化物 (SO) 是苯乙烯的主要代谢物，苯乙烯是一种工业神经毒性化合物，可导致各种神经退行性疾病。本研究旨在探索 SO 细胞毒性的机制 (0.5 - 4 mM）在初级皮层神经元中并评估槲皮素（QUER）的神经保护潜力。我们的结果表明，暴露于 SO 会以浓度依赖性方式降低皮质神经元的活力。在QUER的存在下，细胞活力显着增加。QUER的神经保护作用与细胞内活性氧（ROS）的减少、钙超载的减少和SO引起的线粒体膜去极化的恢复有关。此外，为了评估 SO 触发的神经元死亡机制，细胞与 Ac-DEVD-CHO、Calpeptin 和 Necrostatin-1、caspase-3、钙蛋白酶和坏死性凋亡的药理学抑制剂分别孵育。数据显示，这三种抑制剂减少了 SO 诱导的细胞死亡，并暗示了细胞凋亡、坏死和坏死通路。然而，蛋白质印迹分析表明，QUER 减弱了 caspase-3 的激活，但并未阻止钙蛋白酶的活性。总之，这些数据表明 SO 的细胞毒性是由氧化应激和细胞凋亡、坏死和坏死性凋亡机制介导的，而 QUER 对 SO 的神经保护主要取决于其抗凋亡活性。