外源性NO供体对脑缺血/再灌注神经元的保护作用

Protective effect of exogenous NO donors on brain ischemic/reperfused neurons

目的观察外源性一氧化氮(NO)供体硝普钠(SNP)和亚硝基谷胱甘肽(GSNO)对脑缺血/再灌注过程中c-jun N末端激酶3(JNK3)磷酸化的影响,及其对再灌注海马CA1区锥体神经元的作用。方法采用四动脉结扎法建立大鼠全脑缺血模型。SNP(5 mg/kg)溶于生理盐水,间隔1.5 h分别腹腔注射3次,第1次注射时间在全脑缺血前30 min。GSNO(100μg/kg)溶于生理盐水,缺血前20 min于侧脑室注射给药。大鼠随机分为假手术组、缺血/再灌注组、缺血/再灌注溶剂对照组、缺血/再灌注给予SNP组和缺血/再灌注给予GSNO组,缺血15 min后分别灌注1天或5天。利用免疫印迹、免疫沉淀和焦油紫染色法检测相关蛋白的表达、磷酸化水平及神经元细胞的存活。结果给予SNP组和GSNO组与生理盐水对照组相比,能够显著抑制脑缺血/再灌注过程中JNK3的磷酸化,并且能显著提高再灌注过程中海马CA1区锥体神经元的存活。结论外源性NO供体能够抑制脑缺血/再灌注过程中JNK3的磷酸化,进一步对海马CA1区锥体神经元发挥了保护作用。本研究为外源性NO对脑卒中的有效治疗提供了参考价值。

Objective To observe the effect of exogenous nitric oxide （NO） donors sodium nitropreside （SNP） and S -nitrosoglutathione （GSNO） on the phosphorylation of c -jun N -terminal kinase 3 （JNK3） during brain ischemia/ reperfusion and on the reperfused hippocampal CA1 pyramidal neurons. Methods Rats were induced by the fourvessel occlusion method （4 - VO） to establish rat model of global ischemla. The rats were injected with SNP dissolved in saline at a dose of 5 mg/kg, three times at an internal of 1.5 h, i.p. , with the first injection given 30 min before ischemia. GSNO 100 μg/kg, dissolved in saline, was administrated intracerebroventricularly 20 min prior to ischemia. Rats were randomly assigned to 5 groups ： sham group, ischemia/reperfusion （I/R） group, I/R solvent （saline） control group, I/R drug test （SNP） group and I/R drug test （GSNO） group. Reperfusion was allowed for 1 day or 5 days fol- lowing transient brain ischemia of 15 min. Immunoblot （IB）, immunoprecipitation （IP） and Cresyl Violet staining were performed to examine the expression and phosphorylation of relevant proteins and the survival of hippocampal neurons. Results Compared with saline group, treatment with SNP and GSNO groups dramatically depressed the phosphorylation of JNK3 and remarkably improved the survival of hippocampal CA1 pyramidal neurons, following reperfusion. Conclu- sions Exogenous NO donors can inhibit phosphorylation of JNK3 during brain ischemia/reperfusion, hence the neuro- protective effect on reperfused hippocampal CA1 pyramidal neurons.