Cao Zhuoxiao, Hallur Seema, Qiu Harry Z, Peng Xingxiang, Li Yunbo
Department of Pharmaceutical Sciences, St. John's University College of Pharmacy and Allied Health Professions, 8000 Utopia Parkway, Jamaica, NY 11439, USA.
Biochem Biophys Res Commun. 2004 Apr 16;316(4):1043-9. doi: 10.1016/j.bbrc.2004.02.156.
Substantial evidence suggests that peroxynitrite generated from the bi-radical reaction of nitric oxide and superoxide is critically involved in the pathogenesis of neurodegenerative disorders, such as Parkinson's disease. Reaction with sulfhydryl (SH)-containing molecules has been proposed to be a major detoxification pathway of peroxynitrite in biological systems. This study was undertaken to determine if chemically elevated intracellular reduced glutathione (GSH), a major SH-containing biomolecule, affords protection against peroxynitrite-mediated toxicity in cultured neuronal cells. Incubation of human neuroblastoma SH-SY5Y cells with the unique chemoprotectant, 3H-1,2-dithiole-3-thione (D3T), led to a significant elevation of cellular GSH in a concentration-dependent fashion. To examine the protective effects of D3T-induced GSH on peroxynitrite-mediated toxicity, SH-SY5Y cells were pretreated with D3T and then exposed to either the peroxynitrite generator, 3-morpholinosydnonimine (SIN-1), or the authentic peroxynitrite. We observed that D3T-pretreated cells showed a markedly increased resistance to SIN-1- or authentic peroxynitrite-induced cytotoxicity, as assessed by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium reduction assay. Conversely, depletion of cellular GSH by buthionine sulfoximine (BSO) caused a marked potentiation of SIN-1- or authentic peroxynitrite-mediated cytotoxicity. To further demonstrate the causal role for GSH induction in D3T-mediated cytoprotection, SH-SY5Y cells were co-treated with BSO to abolish D3T-induced GSH elevation. Co-treatment of the cells with BSO was found to significantly reverse the protective effects of D3T on SIN-1- or authentic peroxynitrite-elicited cytotoxicity. Taken together, this study demonstrates for the first time that D3T can induce GSH in cultured SH-SY5Y cells, and that the D3T-augmented cellular GSH defense affords a marked protection against peroxynitrite-induced toxicity in cultured human neuronal cells.
大量证据表明，一氧化氮和超氧化物的双自由基反应产生的过氧亚硝酸盐在神经退行性疾病（如帕金森病）的发病机制中起关键作用。与含巯基（SH）分子的反应被认为是生物系统中过氧亚硝酸盐的主要解毒途径。本研究旨在确定化学方法提高细胞内还原型谷胱甘肽（GSH）（一种主要的含SH生物分子）是否能保护培养的神经元细胞免受过氧亚硝酸盐介导的毒性。用人神经母细胞瘤SH-SY5Y细胞与独特的化学保护剂3H-1,2-二硫醇-3-硫酮（D3T）孵育，导致细胞内GSH以浓度依赖性方式显著升高。为了研究D3T诱导的GSH对过氧亚硝酸盐介导的毒性的保护作用，将SH-SY5Y细胞用D3T预处理，然后暴露于过氧亚硝酸盐发生器3-吗啉代-sydnonimine（SIN-1）或纯过氧亚硝酸盐中。我们观察到，通过3-[4,5-二甲基噻唑-2-基]-2,5-二苯基四氮唑还原试验评估，D3T预处理的细胞对SIN-1或纯过氧亚硝酸盐诱导的细胞毒性表现出明显增强的抗性。相反地，丁硫氨酸亚砜胺（BSO）耗尽细胞内GSH会导致SIN-1或纯过氧亚硝酸盐介导的细胞毒性显著增强。为了进一步证明GSH诱导在D3T介导的细胞保护中的因果作用，将SH-SY5Y细胞与BSO共同处理以消除D3T诱导的GSH升高。发现细胞与BSO共同处理可显著逆转D3T对SIN-1或纯过氧亚硝酸盐引起的细胞毒性的保护作用。综上所述，本研究首次证明D3T可在培养的SH-SY5Y细胞中诱导GSH，并且D3T增强的细胞内GSH防御为培养的人神经元细胞提供了针对过氧亚硝酸盐诱导的毒性的显著保护。
