法舒地尔通过抑制NogoA/NgR信号通路减轻H_(2)O_(2)诱导的SH-SY5Y人神经母细胞瘤细胞凋亡并增强突触可塑性  

Fasudil reduces apoptosis of SH-SY5Y cells induced by H2O2 and promotes synaptic plasticity by inhibiting neurite outgrowth inhibitor A and its receptor (NogoA/NgR) signaling pathway

作　　者：郭敏芳[1] 张慧宇 于婧文[1] 章培军 王玉银 魏文悦 宋丽娟 柴智[2] 尉杰忠 马存根[1,2,3] GUO Minfang;ZHANG Huiyu;YU Jingwen;ZHANG Peijun;WANG Yuyin;WEI Wenyue;SONG Lijuan;CHAI Zhi;YU Jiezhong;MA Cungen(Institute of Brain Science/Shanxi Key Laboratory of Inflammatory Neurodegenerative Diseases,Shanxi Datong University,Datong 037009;The Key Research Laboratory of Benefiting Qi for Acting Blood Circulation Method to Treat Multiple Sclerosis of State Administration of Traditional Chinese Medicine/Research Center for Neurobiology,Shanxi University of Chinese Medicine,Jinzhong 030619;Dept.of Neurology,First Affiliated Hospital,Shanxi Medical University,Taiyuan 030001;Dept.of Neurology,Datong Fourth People's Hospital,Datong 037009,China)

机构地区：[1]山西大同大学脑科学研究所/神经炎症及变性疾病基础与应用研究山西省重点实验室,山西大同037009 [2]山西中医药大学国家中医药管理局多发性硬化益气活血重点研究室/神经生物学研究中心,山西晋中030619 [3]山西医科大学生理学系/第一临床医学院,山西太原030001 [4]大同市第四人民医院神经科,山西大同037009

出　　处：《细胞与分子免疫学杂志》2022年第7期625-631,共7页Chinese Journal of Cellular and Molecular Immunology

基　　金：山西省平台基地专项(201805D131005,201805D111009);山西省高等学校科技创新项目(2020L0484);大同市平台基地计划项目(2019198);神经炎症及变性疾病基础与应用研究山西省重点实验室开放课题(KF2019007);山西省卫健委医学科技领军团队(2020TD05);基于炎性反应的重大疾病创新药物山西省重点实验室项目(202105D121011);山西中医药大学青年科学家培育项目(2021-PY-QN-09)。

摘　　要：目的探讨法舒地尔(Fasudil)对H(2)O_(2)诱导的SY5Y人神经母细胞瘤细胞凋亡和突触可塑性的影响及其机制。方法细胞分为PBS对照组、250μmol/L H(2)O_(2)处理组和250μmol/L H(2)O_(2)联合15μg/mL Fasudil处理组。噻唑蓝(MTT)法检测细胞活性,原位末端转移酶标记技术(TUNEL)检测细胞凋亡,免疫荧光细胞化学染色检测神经突生长抑制因子A(NogoA)、NogoA受体(NgR)和突触小泡蛋白(Syn)的表达,Western blot法检测NogoA、NgR、p75神经营养因子受体(p75NTR)和含富含亮氨酸重复序列免疫球蛋白结构域的Nogo相互作用蛋白1(LINGO-1)、Syn和突触后致密区蛋白95(PSD-95)的表达。结果与PBS组比较,H(2)O_(2)组细胞活性降低,细胞凋亡率升高,Fasudil处理可显著提高细胞活性,降低细胞凋亡率。与H(2)O_(2)模型组比较,Fasudil处理能使Syn和PSD-95的表达增加。与PBS组比较,H(2)O_(2)组NogoA及其受体复合物NgR/p75NTR/LINGO-1的表达明显增加,而Fasudil处理可以抑制NogoA及其受体复合物NgR/p75NTR/LINGO-1的表达。结论Fasudil可能通过抑制NogoA/NgR信号通路的激活,从而减轻H(2)O_(2)诱导的SH-SY5Y细胞凋亡以及增强突触的可塑性。Objective To investigate the effect of Fasudil on H(2)O_(2)-induced apoptosis and synaptic plasticity in human neuroblastoma SY5Y cells and its mechanism.Methods The cells were divided into three groups:PBS control group,H(2)O_(2)model group(250μmol/L H(2)O_(2)treatment)and Fasudil intervention group(250μmol/L H(2)O_(2)combined with 15μg/mL Fasudil treatment).MTT assay was applied to detect cell activity and TUNEL was performed to detect cell apoptosis respectively.Immunofluorescence cytochemical staining was used to determine the expression of neurite outgrowth inhibitor A(NogoA),Nogo receptor(NgR)and synaptophysin(Syn).Western blotting was then conducted to detect the expression of NogoA,NgR,p75 neurotrophin receptor(p75NTR),leucine-rich repeat Ig domain-containing Nogo-interacting protein 1(LINGO-1),Syn and postsynaptic density protein-95(PSD-95).Results Compared with the PBS group,the H(2)O_(2)group showed decreased cell viability and increased apoptosis rate while Fasudil treatment significantly increased the cell viability and reduced the apoptosis rate.Compared with the H(2)O_(2)model group,Fasudil intervention increased expressions of Syn and PSD-95.Compared with the PBS group,the expression of NogoA and its receptor complex NgR/p75NTR/LINGO-1 grew significantly in the H(2)O_(2)group,suggesting Fasudil treatment could inhibit the expression of NogoA and its receptor complex NgR/p75NTR/LINGO-1.ConclusionFasudil may inhibit the activation of the NogoA/NgR signaling pathway,therefore reducing the apoptosis induced by H(2)O_(2)in SH-SY5Y cells and enhancing the plasticity of the synapses.

关 键 词：法舒地尔(Fasudil) 神经突生长抑制因子A(NogoA) Nogo受体(NgR) 细胞凋亡 突触可塑性 

分 类 号：Q255[生物学—细胞生物学] R965[医药卫生—药理学] R392-33[医药卫生—药学]