肢体缺血后适应对小鼠脑缺血再灌注损伤的影响及机制  

Effect of limb ischemic postconditioning on brain focal ischemia and reperfusion injury in mice

作　　者：赵瑾[1,2] 李杨[1,3] 黄磊[1] 邱林[1] 田单[1] 潘慧[1] 祝世功[1] 

机构地区：[1]北京大学医学部生理学及病理生理学系,北京100191 [2]蚌埠医学院病理生理学教研室,安徽蚌埠233030 [3]西藏民族学院生理学教研室,陕西咸阳712082

出　　处：《中国病理生理杂志》2010年第6期1096-1101,共6页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.30770804;No.30871034);安徽省教育厅资助项目(No.KJ2010B122);国家基础科学人才培养基金资助项目(No.J0630853/J0108)

摘　　要：目的:建立小鼠脑缺血后进行短暂肢体缺血提高脑缺血耐受模型,确定肢体缺血后适应对脑缺血时程的影响及热休克蛋白70(HSP70)的作用,探讨肢体缺血后适应(LIPostC)对脑缺血/再灌注损伤抑制作用和机制。方法:复制小鼠大脑中动脉闭塞模型(MCAO),第1批实验将小鼠分为9组:假手术组、脑缺血/再灌注组(缺血时间分别0.5 h、1 h、1.5 h、2 h组),脑缺血/再灌注+短暂肢体缺血(LIPostC)组(0.5 h+LIPostC、1 h+LIPostC、1.5 h+LIPostC、2 h+LIPostC组)。分别观察小鼠运动行为变化;TTC染色测量脑梗死体积;HE染色观察脑组织损伤程度;TUNEL法检测神经元凋亡程度。第2批实验将小鼠随机分为4组:假手术组、脑缺血/再灌注组、MCAO+LIPostC组和MCAO+LIPostC+quercetin组(缺血时间为2 h)。术后24 h用Western blotting法检测脑皮质中HSP70蛋白表达和神经功能评分。结果:脑缺血时程影响小鼠运动行为和脑损伤程度,随脑缺血时间延长,小鼠的脑再灌注损伤程度加重,其行为缺陷和脑病理变化明显;缺血2 h组脑损伤程度比缺血1.5 h组和缺血1 h组严重(P<0.05)。脑缺血后不同时间施加LIPostC显示不同程度的神经保护作用。LIPostC各组与相对应的I/R组比较,其脑再灌注损伤程度呈现不同程度减轻,行为学评分降低、脑梗死体积减小,脑皮质损伤减轻,TUNEL阳性凋亡细胞数目减少。脑缺血2 h再灌注损伤较重,但LIPostC仍具有明显的脑保护作用。以2 h脑缺血小鼠为模型进行机制研究,结果表明,LIPostC可提高缺血脑组织HSP70蛋白表达,改善神经功能,HSP70抑制剂quercetin可削弱LIPostC的这种脑保护作用。结论:LIPostC可抑制MCAO小鼠的脑缺血再灌注损伤,促进缺血脑区HSP70表达和改善神经功能。HSP70在LIPostC提高MCAO小鼠的脑缺血耐受机制中发挥重要作用。AIM： To establish the mouse model in which the limbic ischemic postconditionning （LIPostC） enhances the tolerance against brain ischemia, and to investigate the effects of LIPostC on the ischemic extent and roles of heat shock protein 70 （HSP70） in ischemia and reperfusion injury. METHODS： The male Kunming mice were used in the study. The brain ischemia reperfusion （I/R） model was made by middle cerebral artery occlusion （MCAO）. In the first test, the male mice were randomly divided into 9 groups （n = 10） ： sham group, ischemia/reperfusion （I/R） groups （with ischemia for 0. 5 h, 1 h, 1.5 h and 2 h） and LIPostC ＋ I/R groups （0.5 h ＋ LIPostC, 1 h ＋ LIPostC, 1.5 h ＋LIPostC,2 h ＋ LIPostC）. The reperfusion was performed after LIPostC for 24 h. After the neurologie deficit scores were evaluated, the brains were taken out to measure the infarct volume with TTC staining and to observe the pathological changes of cerebral cortex with HE staining. The neuronal apoptosis was determined by TUNEL. In the second test, the male mice were randomized into 4 groups （ n = 6） ： sham group, I/R group, LIPostC ＋ I/R group and LIPostC ＋ I/R ＋ quercetin group （2 h ischemia）. The neurological deficit scores were evaluated at 24 h after operation. The expression of HSP70 was determined by Western blotting. RESULTS： The duration of brain ischemia was related to the motor behavior and degree of brain injury. The longer the ischemic duration of the brain was performed, the more severe the pathological and behavioral changes were observed. The brain injury in 2 h MCAO mice was more severe than that in 1 h and 1.5 h MCAO mice （P 〈 0. 05 ）. Compared to I/R group, each LIPostC group showed lower neurological score, less infarct volume and TUNEL positive neuron. The expression of HSP70 protein was increased and neurological functions were improved significantly in the mice with LIPostC. However, the neuroprotective role of LIPostC was attenuated by treating with quercetin, an

关 键 词：再灌注损伤 脑缺血耐受 肢体缺血后适应 热休克蛋白质70 小鼠 

分 类 号：R363[医药卫生—病理学]