左乙拉西坦对小鼠局灶性脑缺血-再灌注损伤的神经保护作用  

The neuro-protection effect of levetiracetam against focal cerebral ischemia-reperfusion injury in mice

作　　者：周广平 郝俊杰[2] 李颖[2] 左联[2] 李刚[1,2] 

机构地区：[1]新疆喀什地区莎车县人民医院脑系科,844700 [2]同济大学附属东方医院神经内科脑血管病中心,上海200125

出　　处：《中华脑科疾病与康复杂志（电子版）》2016年第2期83-88,共6页Chinese Journal of Brain Diseases and Rehabilitation（Electronic Edition）

基　　金：新疆自然科学基金支持项目(201442137-13)

摘　　要：目的研究突触囊泡蛋白2A(SV2A)配体左乙拉西坦(LEV)对小鼠急性局灶性脑缺血-再灌注损伤的神经保护作用及其机制。方法实验第一部分:将20只SPF级C57BL/6小鼠分为三组:手术组(WT组,n=6),腹腔注射(150 mg/kg)LEV 60 min后手术组(WT-LEV组,n=8),假手术组(sham组,n=6),采用线栓法制作小鼠大脑中动脉闭塞缺血-再灌注模型后检测脑组织损伤情况。第二部分:体外进行C57BL/6小鼠海马神经元培养后通过氧糖剥夺建立缺血神经元模型,A组不添加上清液正常培养,作为对照组(n=3),B组添加野生型C57BL/6小鼠的CD8+T淋巴细胞激活后的上清液进行体外培养(n=3),C组添加穿孔素基因敲除C57BL/6 Prf1-/-小鼠的CD8+T淋巴细胞激活后的上清液进行体外培养(n=3),D组培养液中加入穿孔素抗体(n=3),分别检测各组神经元活性。结果 (1)脑缺血-再灌注后24 h:WT-LEV组小鼠神经行为评分(Longa评分)低于WT组(1.82±0.74 vs.2.81±1.03,P<0.05),WT-LEV组小鼠脑梗死体积百分比小于WT组[(11.29±2.30)%vs.(25.21±5.31)%,P<0.05],WT-LEV组小鼠脑组织凋亡细胞数比率低于WT组[(49.05±15.49)%vs.(77.73±12.77)%,P<0.05]。(2)离体培养缺血海马神经元24、36、48 h时,B组的MTT标记的阳性细胞相对数和A组相比均明显减少(24 h:76.93±4.48 vs.94.57±4.17,P<0.05;36 h:63.10±12.80 vs.94.57±4.45,P<0.05;48 h:41.33±9.76 vs.93.07±4.54,P<0.05);D组的MTT标记的阳性细胞相对数和A组相比在共同培养24 h和36 h均没有明显降低,在48 h时出现明显降低(76.53±6.73 vs.93.07±4.54,P<0.05);在共同培养从0 h开始至48 h时,C组的MTT标记的阳性细胞相对数和A组相比均没有明显减少。结论 SV2A配体LEV对小鼠急性脑缺血-再灌注的神经损伤有保护作用,其机制可能与通过抑制脑内穿孔素的释放有关。Objective To explore the neuro-protection effect and mechanism of levetiracetam （LEV） [ a ligand of synaptic vesicle protein 2A（SV2A） ] against acute focal cerebral ischemia-reperfusion injury in mice. Methods The first part：20 SPF C57BL/6 mice were divided into three groups,the surgical group（WT group,n = 6）,the surgical group of wild type C57BL/6 mice that given intraperitoneal injection （ 150 mg/kg） of LEV （ WT-LEV group, n = 8 ）, the control group （ sham group, n = 6 ）, line plug method was used to make a mouse model of isehemia-reperfasion caused by middle cerebra/artery occlusion, then mice brain tissue injury were detected. The second part： in vitro, ischemic neurones model was prepared with oxygen and glucose deprivation（OGD） method in C57BL/6 mice hippocampal neurons, CD8 ＋ T lymphocytes were isolated separately from C57BL/6 （ Group B, n = 3 ） and C57BL/6 Prf-/- （ Group C, n = 3 ） mice, then were activated, supernatant of activated CD8 ~ T lymphocytes and perforin antibodies （ Group D, n = 3 ） was added to the hippocampal neurons in vitro,group A（control group,n = 3 ）did not add any supernatant fluid, neuron viability was assessed in different groups. Results （ 1 ） 24 h after reperfusion, neurobehavioral scores （ Longa score） in WT-LEV group was less than WT group （ 1.82 ± 0.74 vs. 2. 81± 1.03, P 〈 0.05 ） ; brain infarction volume percentage in WT-LEV group was less than WT group [ （ 11.29 ±2. 30） % vs. （25.21 ± 5.31）% ,P 〈0. 05 ];brain cells apoptosis ratio in WT-LEV group was less than WT group [ （49. 05 ± 15.49）% vs. （77.73 ± 12. 77）% ,P 〈0. 05]. （2） Ischemic hippocampal neurons were cultured for 24,36, 48 h in vitro, relative number of positive cells marked by MTT in group B were significantly less than group A （2g h ：76. 93 ± g. 48 vs. 94. 57 ± 4. 17, P 〈 0. 05 ; 36 h ： 63.10 ± 12. 80 vs. 94. 57 ± 4. 45, P 〈 0. 05 ; 48 h ： 41.33 ± 9. 76 vs. 93.07± 4. 54, P 〈 0. 05 ）. Relative number of

关 键 词：再灌注损伤 神经保护药 左乙拉西坦 SV2A配体 

分 类 号：R743.3[医药卫生—神经病学与精神病学]