Wei T, Chen C, Hou J, Xin W, Mori A
Department of Molecular and Cellular Biophysics, Institute of Biophysics, Chinese Academy of Sciences, Beijing, PR China.
Biochim Biophys Acta. 2000 Oct 20;1498(1):72-9. doi: 10.1016/s0167-4889(00)00078-1.
Within the central nervous system and under normal conditions, nitric oxide (NO) is an important physiological signaling molecule. When produced in large excess, NO also displays neurotoxicity. In our previous report, we have demonstrated that the exposure of neuronal cells to NO donors induced apoptotic cell death, while pretreatment with free radical scavengers L-ascorbic acid 2-[3, 4-dihydro-2,5,7,8-tetramethyl-2-(4,8, 12-trimethyltridecyl)-2H-1-benzopyran-6-yl-hydrogen phosphate] potassium salt (EPC-K1) or superoxide dismutase attenuated apoptosis effectively, suggesting that reactive oxygen species (ROS) may be involved in the cascade of events leading to apoptosis. In the present investigation, we directly studied the kinetic generation of ROS in NO-treated neuronal cells by flow cytometry using 2', 7'-dichloro-fluorescein diacetate and dihydrorhodamine 123 as redox-sensitive fluorescence probes. The results indicated that exposure of cerebellar granule cells to the NO donor S-nitroso-N-acetylpenicillamine (SNAP) induced oxidative stress, which was characterized by the accumulation of cytosolic and mitochondrial ROS, the increase in the extracellular hydrogen peroxide level, and the formation of lipid peroxidation products. SNAP treatment also induced apoptotic cell death as confirmed by the formation of cytosolic mono- and oligonucleosomes. Pretreating cells with the novel antioxidant EPC-K1 effectively prevented oxidative stress induced by SNAP, and attenuated cells from apoptosis.
在中枢神经系统内且在正常情况下，一氧化氮（NO）是一种重要的生理信号分子。当大量产生时，NO也表现出神经毒性。在我们之前的报告中，我们已经证明，将神经元细胞暴露于NO供体可诱导凋亡性细胞死亡，而用自由基清除剂L-抗坏血酸2-[3,4-二氢-2,5,7,8-四甲基-2-(4,8,12-三甲基十三烷基)-2H-1-苯并吡喃-6-基-氢磷酸酯]钾盐（EPC-K1）或超氧化物歧化酶进行预处理可有效减轻凋亡，这表明活性氧（ROS）可能参与了导致凋亡的一系列事件。在本研究中，我们使用2',7'-二氯荧光素二乙酸酯和二氢罗丹明123作为氧化还原敏感荧光探针，通过流式细胞术直接研究了经NO处理的神经元细胞中ROS的动力学产生情况。结果表明，小脑颗粒细胞暴露于NO供体S-亚硝基-N-乙酰青霉胺（SNAP）会诱导氧化应激，其特征为胞质和线粒体ROS的积累、细胞外过氧化氢水平的升高以及脂质过氧化产物的形成。SNAP处理还诱导了凋亡性细胞死亡，这通过胞质单核小体和寡核小体的形成得到证实。用新型抗氧化剂EPC-K1预处理细胞可有效预防SNAP诱导的氧化应激，并减轻细胞凋亡。
