Miao Wanying, Qu Zhongwei, Shi Kejie, Zhang Dengyue, Zong Yanyan, Zhang Gongliang, Zhang Guangyi, Hu Shuqun
Center for Biochemistry and Molecular Biology; Jiangsu Key Laboratory of Brain Disease and Bioinformation, Xuzhou Medical College, Xuzhou, Jiangsu 221002, China.
Department of Physiology, College of Basic Medical Science, Anhui Medical University, 81 Meishan Road, Hefei, Anhui 230032, China.
Brain Res. 2015 Nov 19;1627:165-76. doi: 10.1016/j.brainres.2015.08.020. Epub 2015 Aug 28.
Cerebral ischemia-reperfusion is associated with NMDA receptor-mediated calcium influx which activates neuronal nitric oxide synthase (nNOS) and consequently induces NO production. NO S-nitrosylates cellular protein and aggravates neuronal injury. Receptor-interacting protein 3 (RIP3) is a sensor molecule regulating cell apoptosis and necrosis. However, the roles of RIP3 in cerebral ischemic injury remain elusive. In this study, we reported that RIP3 could be S-nitrosylated by the exogenous NO donor GSNO in HEK293 cells and the Cys(119) residue was the key nitrosylation site. In addition, we found that cerebral ischemia induced RIP3 S-nitrosylation at different time points of reperfusion, which was coupling with RIP3 phosphorylation (which is associated with its activation) and its interaction with receptor-interacting protein 1 (RIP1), and this process facilitated cerebral ischemic injury. Treatment with NMDA receptor antagonist MK801, or nNOS inhibitor 7NI, diminished RIP3 S-nitrosylation and reduced neuronal damage. Taken together, these data demonstrated that NMDAR-dependent RIP3 S-nitrosylation induced by ischemia facilitated its activation in the early stages of ischemia, blocking this process could reduce the ischemia neuronal injury.
脑缺血再灌注与N-甲基-D-天冬氨酸（NMDA）受体介导的钙内流有关，该钙内流激活神经元型一氧化氮合酶（nNOS），从而诱导一氧化氮（NO）生成。NO使细胞蛋白发生S-亚硝基化，加重神经元损伤。受体相互作用蛋白3（RIP3）是一种调节细胞凋亡和坏死的传感分子。然而，RIP3在脑缺血损伤中的作用仍不清楚。在本研究中，我们报道在HEK293细胞中，RIP3可被外源性NO供体GSNO进行S-亚硝基化，半胱氨酸（Cys）119残基是关键的亚硝基化位点。此外，我们发现脑缺血在再灌注的不同时间点诱导RIP3 S-亚硝基化，这与RIP3磷酸化（与其激活有关）及其与受体相互作用蛋白1（RIP1）的相互作用相关联，并且这一过程促进了脑缺血损伤。用NMDA受体拮抗剂MK801或nNOS抑制剂7NI处理，可减少RIP3 S-亚硝基化并减轻神经元损伤。综上所述，这些数据表明缺血诱导的NMDAR依赖性RIP3 S-亚硝基化在缺血早期促进其激活，阻断这一过程可减轻缺血性神经元损伤。
