摘要目的：探讨尼可地尔对深低温低流量(deep hypothermic low flow, DHLF)小鼠缺血再灌注模型的神经保护作用及其可能机制。方法105只3周龄雄性C57/BL-6小鼠随机分为假手术组、模型组、尼可地尔5 mg/kg 组、尼可地尔10 mg/kg 组、尼可地尔20 mg/kg 组、尼可地尔20 mg/kg +LY294002组以及 LY294002组,每组15只。建立 DHLF 模型,在再灌注24 h后取小鼠脑组织进行 HE和 TUNEL 染色,观察大脑皮质神经元病理学改变和细胞凋亡,应用蛋白质印迹法检测总 Akt、磷酸化Akt(phospho-Akt, p-Akt)、Bcl-2和 Bax 蛋白表达。结果 HE 病理学染色显示,尼可地尔组皮质神经元损伤减轻,胞膜凹陷、胞核固缩、浓染、核仁模糊不清等现象明显减少,神经元细胞形态基本恢复正常。 TUNEL 染色显示,各剂量尼可地尔组凋亡指数均较模型组显著降低(P 均<0．05)。蛋白质印迹分析显示,与模型组相比,各剂量尼可地尔组 p-Akt 和 Bcl-2蛋白表达水平显著升高(P 均<0．05),而Bax 蛋白表达水平则显著降低(P 均<0．05)。加入磷脂酰肌醇3激酶(phosphatidylinositol 3-kinase, PI3K)特异性抑制剂 LY294002后,脑皮质神经元病理学损伤较模型组无明显差异,凋亡指数和p-Akt、Bcl-2和 Bax 蛋白表达水平较模型组差异无统计学意义。结论尼可地尔对 DHLF 小鼠模型具有一定的神经保护作用,其机制可能与激活 PI3K/Akt 信号通路进而调控下游蛋白 Bcl-2和 Bax 表达有关。

abstractsObjective To investigate the neuroprotective effect and possible mechanism of nicorandil in mice under deep hypothermic low flow (DHLF). Methods A total of 105 3-week-old male C57/BL-6 mice were randomly divided into 7 groups: sham operation, model, nicorandil (5, 10, and 20 mg/kg), nicorandil 20 mg/kg + LY294002, and LY294002 groups (n = 15 in each group). A DHLF model was induced. At 24 h after reperfusion, the brain tissues of mice were taken out for HE and TUNEL staining. The pathological changes of cerebral cortical neurons and apoptosis were observed. Western blot was used to detect the expression levels of the total Akt, phospho-Akt (p-Akt), Bcl-2, and Bax. Results HE pathological staining showed that cortical neuronal injury was reduced, the phenomena of cel membrane depression, nuclear condensation, concentrated dye, and the blurring of the nucleus were decreased significantly in nicorandil group. The morphology of neurons was basicaly restored to normal. TUNEL staining showed that the apoptosis index in various dose groups of nicorandil was decreased significantly compared with the model group (al P < 0. 05). Western blot analysis showed that the expression levels of p-Akt and Bcl-2 proteins increased significantly in various dose groups of nicorandil compared with the model group (al P < 0. 05), and the expression level of Bax protein was decreased significantly (al P < 0. 05 ). After adding the phosphatidylinositol 3-kinase (PI3K) specific inhibitor LY294002, there was no significant difference in neurons pathological injury in the cortex compared with the model group. There was no significant difference in the apoptosis index, and the expression levels of p-Akt, Bcl-2, and Bax compared with the model group. Conclusions Nicorandil has a certain neuroprotective effect in mice under DHLF. Its mechanism may be associated with the activation of PI3K/Akt signaling pathway, and then further regulation of the downstream protein Bcl-2 and Bax expression.