缺血预处理诱导脑缺血耐受时间依赖性的实验研究

Time-dependent contribution of cerebral ischemic tolerance induced by ischemic preconditioning in rats

目的探讨缺血预处理诱导缺血耐受的时间依赖性及可能的机制。方法健康SD大鼠110只，采用随机数字表法分为假手术（SS）＋大脑中动脉阻塞（MCAO）组（简称SS组）和缺血预处理（IP）＋MCA0组（简称1P组），每组50只，另10只备用。将两组大鼠再随机分为5个亚组（n=10），IP组给予预缺血10min后分别于6h、24h、3d、7d、14d后给予大脑中动脉完全阻塞22h，再灌注2h；SS组未进行缺血预处理，而单纯暴露颈总动脉处的解剖结构10min，余步骤同IP组。采用Swanson间接法和Nick＆Spot图像采集分析系统计算各组大鼠梗死体积，光镜下观察各组脑组织病理变化，采用免疫组化方法检测大鼠脑组织胶质纤维酸性蛋白（GFAP）、脑源性神经营养因子（BDNF）表达。结果给予10rain的大脑中动脉短暂的缺血预处理能够明显减少24h、3d、7d、14d后再次缺血后的梗死体积（P〈0．05），而6h时间点脑梗死体积与SS组比较无统计学差异（P〉0．05）。与SS组比较，IP组各时间点GFAP和BNDF阳性表达增多（P〈0．05），且间隔3d、7d组增多明显（P〈0．05），14d后开始逐渐降低。结论IP不但能引起神经系统损害，而且能够诱导缺血耐受的产生；IP诱导的缺血耐受对IP后3～7d发生的再缺血损害保护作用最强。BDNF表达基本与缺血耐受的保护作用时间规律一致，提示BDNF可能是脑缺血耐受的重要机制之一。

Objective To investigate the ischemic tolerance induced by ischemic preconditioning （IP） in difference time intervals in rats. Methods IP was induced by intraluminal filaments in middle cerebral artery for 10 min. Middle cerebral artery occlusion （MCAO） was induced by intraluminal filaments on the different time points of 6 h, 24 h, 3 d, 7 d and 14 d post IP and continued for 2 hours. 110 SD rats were randomly divided into 2 groups： The SS＋MCAO group only received 2 h MCAO followed by 22 h reperfusion with pretreatment of sham surgery, focal IP group （IP＋MCAO） received 2 h MCAO followed by 22 h reperfusion with pretreatment of IP. At the end point of 22 h perfusion, the infarct volume, histopathological changes, the expressions of glial fibrillary acidic protein （GFAP） and brain-derived neurotrophic factor （BDNF） by using immunohistochemistry method were evaluated in each group. Results The infarct volume in IP＋MCAO group （24 h, 3 d, 7 d, 14 d） were significantly less than that in SSq MCAO group （P〈0.05）, and within 6 h the infarction volume was comparable between these two groups. The brain pathological changes were more pronounced in IP＋ MCAO groui3 than in the SS＋MCAO group. The numbers of GFAP and BDNF positive cells in the brain in the 3 d/7 d subgroups of IP＋MCAO were significantly more than those in the SS＋ MCAO group （P〈0.05）. The GFAP expression in the SS ＋ MCAO group was few, increased in each group of IP ＋ MCAO, and the increase was more in the 3 d and 7 d subgroup of IP ＋ MCAO （P〈0.05）. BNDF expression was detectable at 6 h after ischemia, became maximum 3 d and 7 d, and began to decrease 14 d post IP. Conclusions IP not only caused damages to the nervous system, but also produced ischemic tolerance; ischemic tolerance occurred in 24 h, 3 d, 7 d, 14 d post IP, and showed the strongest protective effects in brain in 3 7 d. The BDNF expression and the protective effects of ischemie tolerance were in the similar time courses, suggesting that BDNF may be one of the important mechanisms for ischemic tolerance.