中国神经再生研究(英文版)

中国神经再生研究(英文版) ›› 2023, Vol. 18 ›› Issue (6): 1277-1285.doi: 10.4103/1673-5374.357906

• 原著:脑损伤修复保护与再生 • 上一篇    下一篇

神经元型一氧化氮合酶-活性氧通路参与癫痫中的细胞凋亡和焦亡

  

  • 出版日期:2023-06-15 发布日期:2023-01-05
  • 基金资助:
    国家自然科学基金项目（81971212，81601129）；西南医科大学心血管研究所教育部医学电生理学重点实验室和四川省医学电生理重点实验室开放基金项目（KeyME-2018-07）；辽宁省兴辽人才计划项目（XLYC1907164）

Neuronal nitric oxide synthase/reactive oxygen species pathway is involved in apoptosis and pyroptosis in epilepsy

Xiao-Xue Xu1, 2, #, Rui-Xue Shi1, #, Yu Fu1, #, Jia-Lu Wang2, #, Xin Tong1, Shi-Qi Zhang1, Na Wang3, Mei-Xuan Li1, Yu Tong1, Wei Wang4, Miao He1, Bing-Yang Liu5, *, Gui-Lan Chen3, *, Feng Guo1, *#br#   

  1. 1Department of Pharmaceutical Toxicology, School of Pharmaceutical Science, China Medical University, Shenyang, Liaoning Province, China; 2Department of Neurology, the First Affiliated Hospital of China Medical University, Shenyang, Liaoning Province, China; 3Key Laboratory of Medical Electrophysiology, Ministry of Education & Medical Electrophysiological Key Laboratory of Sichuan Province, Institute of Cardiovascular Research, Southwest Medical University, Luzhou, Sichuan Province, China; 4Department of Endocrinology and Metabolism, the Fourth Affiliated Hospital of China Medical University, Shenyang, Liaoning Province, China; 5Department of Endocrinology, Shengjing Hospital of China Medical University, Shenyang, Liaoning Province, China
  • Online:2023-06-15 Published:2023-01-05
  • Contact: Feng Guo, MD, PhD, blueforest611@hotmail.com; Gui-Lan Chen, PhD, chenguilan@swmu.edu.cn; Bing-Yang Liu, MD, PhD, Liuliuby@sj-hospital.org.
  • Supported by:
    This work was supported by the Natural Science Foundation of China, Nos. 81971212 (to FG), 81601129 (to XXX); the Open Fund of the Key Laboratory of Medical Electrophysiology, Ministry of Education & Medical Electrophysiological Key Laboratory of Sichuan Province, Institute of Cardiovascular Research, Southwest Medical University, No. KeyME-2018-07 (to FG); and Liaoning Province Xingliao Talent Program Project, No. XLYC1907164 (to FG).

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摘要：

神经元一氧化氮合酶的功能障碍会导致神经毒性，从而引发癫痫等神经疾病中的细胞死亡。有研究发现抑制神经元型一氧化氮合酶活性可增大癫痫阈值，即存在抗惊厥作用。尽管如此，神经元型一氧化氮合酶在癫痫发作中的确切作用和潜在机制仍不清楚。此次实验利用遗传性癫痫大鼠模型-震颤大鼠海马的RNA测序表达谱，通过加权基因共表达网络分析、功能富集分析发现一氧化氮合酶生物合成与震颤大鼠癫痫发生高度相关。同时发现在震颤大鼠海马中存在神经元型一氧化氮合酶的高表达。接下来，他们发现震颤大鼠海马线粒体结构的受损以及功能酶（琥珀酸脱氢酶和Na+-K+-ATP酶）水平的异常。此外，以匹罗卡品诱导的N2a细胞模型模拟癫痫样损伤，经神经元型一氧化氮合酶抑制剂7-硝基吲唑干预后，氧化应激相关指标丙二醛、乳酸脱氢酶和超氧化物歧化酶的变化可被逆转，且活性氧水平的升高可被7-硝基吲唑或活性氧阻断剂N-乙酰半胱氨酸反转。而7-硝基吲唑或N-乙酰半胱氨酸下调了模型中caspase-3和细胞色素c的高表达，并逆转癫痫细胞凋亡；同时，7-硝基吲唑或N-乙酰半胱氨酸还能下调模型中NLRP3、消皮素D、白细胞介素1β、白细胞介素18的异常高表达，提示其逆转癫痫细胞焦亡。上述结果提示神经元型一氧化氮合酶-活性氧通路可参与癫痫细胞死亡，而抑制神经元型一氧化氮合酶活性或 其诱导的氧化应激可能在癫痫中发挥神经保护作用。

https://orcid.org/0000-0001-9833-9691 (Feng Guo)

关键词: 癫痫, 一氧化氮合酶, 氧化应激, 震颤大鼠, 细胞凋亡, 细胞焦亡, 细胞死亡, RNA测序, 生物信息学分析, 加权基因共表达网络分析

Abstract: Dysfunction of neuronal nitric oxide synthase contributes to neurotoxicity, which triggers cell death in various neuropathological diseases, including epilepsy. Studies have shown that inhibition of neuronal nitric oxide synthase activity increases the epilepsy threshold, that is, has an anticonvulsant effect. However, the exact role and potential mechanism of neuronal nitric oxide synthase in seizures are still unclear. In this study, we performed RNA sequencing, functional enrichment analysis, and weighted gene coexpression network analysis of the hippocampus of tremor rats, a rat model of genetic epilepsy. We found damaged hippocampal mitochondria and abnormal succinate dehydrogenase level and Na+-K+-ATPase activity. In addition, we used a pilocarpine-induced N2a cell model to mimic epileptic injury. After application of neuronal nitric oxide synthase inhibitor 7-nitroindazole, changes in malondialdehyde, lactate dehydrogenase and superoxide dismutase, which are associated with oxidative stress, were reversed, and the increase in reactive oxygen species level was reversed by 7-nitroindazole or reactive oxygen species inhibitor N-acetylcysteine. Application of 7-nitroindazole or N-acetylcysteine downregulated the expression of caspase-3 and cytochrome c and reversed the apoptosis of epileptic cells. Furthermore, 7-nitroindazole or N-acetylcysteine downregulated the abnormally high expression of NLRP3, gasdermin-D, interleukin-1β and interleukin-18. This indicated that 7-nitroindazole and N-acetylcysteine each reversed epileptic cell death. Taken together, our findings suggest that the neuronal nitric oxide synthase/reactive oxygen species pathway is involved in pyroptosis of epileptic cells, and inhibiting neuronal nitric oxide synthase activity or its induced oxidative stress may play a neuroprotective role in epilepsy.

Key words: apoptosis, bioinformatics analysis, cell death, epilepsy, nitric oxide synthase, oxidative stress, pyroptosis, RNA sequencing, Tremor rat, weighted gene co-expression network analysis