五味子乙素对缺氧缺血新生大鼠的脑保护作用及机制

Brain protective effect of schisandrin B on neonatal rats with hypoxia-ischemia and its mechanism

目的研究五味子乙素(Sch B)对缺氧缺血新生大鼠的脑保护作用。方法SD新生大鼠48只随机均分为假手术组、模型组、Sch B组(Sch B,55.4 mg/kg)及3-甲基腺嘌呤(3-MA)组(自噬抑制剂3-MA,10 mg/kg),后3组大鼠采用结扎左侧颈总动脉建立缺氧缺血新生大鼠模型,后两组大鼠在造模同时灌胃或注射相应药物、假手术组和模型组灌胃生理盐水,共7 d;采用Longa分级法对各组大鼠进行神经功能缺失评分,HE染色检测各组大鼠海马神经元组织学变化,免疫荧光染色检测脑组织线粒体形态,免疫印迹实验检测脑组织自噬蛋白(Beclin-1、LC3Ⅱ/LC3Ⅰ)及PINK1、Parkin蛋白表达,试剂盒测定各组大鼠脑组织超氧化物歧化酶(SOD)和丙二醛(MDA)水平。结果与假手术组比较,模型组大鼠海马神经元萎缩、变性,数量减少,出现严重病理损伤,脑神经细胞线粒体长度、脑组织SOD水平明显降低(P<0.05),MDA水平、神经功能缺失评分及脑组织Beclin-1、LC3Ⅱ/LC3Ⅰ、PINK1、Parkin蛋白表达水平明显升高(P<0.05);与模型组比较,Sch B组大鼠脑海马神经元损伤减轻,脑神经细胞线粒体长度、脑组织SOD水平、脑组织自噬蛋白(Beclin-1、LC3II/LC3I)及PINK1、Parkin蛋白表达水平升高(P<0.05),MDA水平、神经功能缺失评分降低(P<0.05);3-MA组大鼠海马神经元损伤加重,脑神经细胞线粒体长度、脑组织SOD水平、脑组织Beclin-1、LC3II/LC3I及PINK1、Parkin蛋白表达水平降低(P<0.05),MDA水平、神经功能缺失评分升高(P<0.05)。结论Sch B降低缺氧缺血大鼠神经功能缺损症状,其机制可能与抑制大鼠脑组织氧化应激、促进线粒体自噬作用有关。

Objective To study the protective effect of schisandrin B(Sch B)on the brain of neonatal rats with hypoxia-ischemia.Methods Forty-eight SD neonatal rats were randomly divided into sham operation group,model group,Sch B group(Sch B 55.4 mg/kg),and 3-methyladenine(3-MA)group(autophagy inhibitor 3-MA,10 mg/kg).The rats in the last three groups were subjected to left common carotid artery ligation to establish hypoxic-ischemic neonatal rat models.The rats in the last two groups were orally administered or injected with corresponding drugs during modeling while rats in the sham surgery group and model group were orally administered with physiological saline for a total of 7 days.The neurological deficits of rats in each group was evaluated by Longa grading method;the pathological changes of hippocampal neurons in each group were detected by HE staining;the morphology of brain mitochondria was detected by immunofluorescence staining;the expressions of autophagy protein(Beclin-1,LC3Ⅱ/LC3Ⅰ),PINK1 and Parkin were examined by Western blotting;the levels of superoxide dismutase(SOD)and malondialdehyde(MDA)in brain tissue of rats in each group were measured by the kit.Results Compared with the sham operation group,the hippocampal neurons in the model group were atrophic and degenerated,its number was reduced,and severe pathological damage occurred.Also,the mitochondrial length in brain nerve cells and the level of SOD in brain tissue were significantly decreased(P<0.05)while the level of MDA,neurological deficit score and expression levels of Beclin-1,LC3Ⅱ/LC3Ⅰ,PINK1 and Parkin protein in brain tissue were significantly increased(P<0.05).Compared with the model group,rats in the Sch B group showed reduced damage to hippocampal neurons,with increased mitochondrial length,SOD levels in brain tissue,autophagic proteins(Beclin-1,LC3Ⅱ/LC3Ⅰ),PINK1,and Parkin protein expression levels in brain tissue(P<0.05)and decreased MDA levels and neurological deficit scores(P<0.05);the damage of hippocampal neurons in 3-MA group was aggravated,with the decreased mitochondrial length in brain nerve cells,SOD levels in brain tissue,the protein expression levels of Beclin-1,LC3II/LC3I,PINK1 and Parkin in brain tissue(P<0.05)and the increased MDA levels and neurological deficit score(P<0.05).Conclusion Sch B can reduce the symptoms of neurological deficit in hypoxic-ischemic rats,and its mechanism may be related to the inhibition of oxidative stress in rat brain tissue and the promotion of mitochondrial autophagy.