异泽兰黄素对脑出血小鼠的神经保护作用及氧化应激和神经元凋亡的影响

Neuroprotective Effect of Eupatilin on Mice with Cerebral Hemorrhage and Influences on Oxidative Stress and Neurocytes Apoptosis

目的:探究异泽兰黄素对脑出血(intracerebral hemorrhage,ICH)小鼠的神经保护作用及其机制。方法:采用自体血注射法复制小鼠ICH模型,同时设立假手术组。分别给予ICH小鼠1、3、10 mg/kg异泽兰黄素灌胃给药3 d。Longa评分法评估小鼠神经缺损情况。称重小鼠脑组织,测定含水量,免疫荧光染色观察脑组织神经元改变及应激诱导蛋白Sestrin2(Sestrin2)表达,ELISA测定脑组织丙二醛(MDA)水平及超氧化物歧化酶(SOD)、过氧化氢酶(CAT)活性,Western blot测定脑组织B淋巴细胞瘤-2基因(Bcl-2)、Bcl-2相关X蛋白(Bax)、天冬氨酸特异半胱氨酸蛋白酶-3(caspase-3)、AMP依赖性蛋白激酶(AMPK)、磷酸化AMPK(p-AMPK)、肝醣合成酶激酶3β(GSK-3β)、磷酸化GSK-3β(p-GSK-3β)蛋白表达。结果:异泽兰黄素干预后,ICH小鼠第1天、第3天神经缺损症状减轻(P<0.05或P<0.01),具有时间-剂量效应;此外,ICH小鼠脑部水肿程度降低(P<0.05或P<0.01),脑组织神经元细胞凋亡减少、尼氏小体数量增加(P<0.05或P<0.01),Sestrin2荧光表达下降(P<0.01),MDA水平降低(P<0.01),SOD、CAT活性提高(P<0.01),Bax、caspase-3蛋白表达下降(P<0.05或P<0.01),Bcl-2、p-AMPK/AMPK、p-GSK-3β/-GSK-3β表达升高(P<0.05或P<0.01),具有剂量效应。结论:异泽兰黄素对ICH小鼠具有神经保护作用,抑制氧化应激和细胞凋亡是其部分作用机制。

Objective:To explore the neuroprotective effect and mechanism of Eupatilin on intracerebral hemorrhage(ICH)mice.Methods:The ICH model mice were established by autologous blood injection me-thod,and sham operation mice were set at the same time.ICH mice were orally administered 1,3,10 mg/kg Eupatilin for 3 days.Longa score was used to evaluate the neurological deficit.Mice brain tissue were collected,weighted to determine the moisture content,staining to observe neuron changes and Sestrin2 expression,ELISA was used to detect MDA level and SOD,CAT activities,Western blot was used to determine Sestrin2,Bcl-2,Bax,caspase-3,AMPK,p-AMPK,GSK-3β,p-GSK-3βprotein expressions.Results:Eupatilin could alleviate nerve defect of ICH mice(P<0.05 or P<0.01),with time-dose dependent;as well decreased brain tissue moisture content(P<0.05 or P<0.01),reduced neuronal apoptosis(P<0.01),increased the number of nissl bodies(P<0.05 or P<0.01),reduced MDA level(P<0.01),increased SOD and CAT activities(P<0.01),and decreased Bax and caspase-3 protein expressions(P<0.05 or P<0.01),and increased the expressions of Sestrin2,Bcl-2,p-AMPK/AMPK,and p-GSK-3β/-GSK-3βprotein expressions(P<0.05 or P<0.01)with dose dependent.Conclusion:Eupatilin exerts a neuro-protective effect on ICH mice by inhibiting oxidative stress and neuronal apoptosis.