Belayev L, Busto R, Zhao W, Ginsberg M D
Department of Neurology, University of Miami School of Medicine, FL 33101, USA.
Brain Res. 1996 Nov 11;739(1-2):88-96. doi: 10.1016/s0006-8993(96)00815-3.
A sensitive quantitative fluorescence method was used to explore the time course and regional pattern of blood-brain barrier (BBB) opening after transient middle cerebral artery occlusion (MCAo). Male Sprague-Dawley rats were anesthetized with halothane and subjected to 2 h of temporary MCAo by retrograde insertion of an intraluminal nylon suture, coated with poly-L-lysine, through the external carotid artery into the internal carotid artery and MCA. Damage to the BBB was judged by extravasation of Evans Blue (EB) dye, which was administered either 2, 3, 24 or 48 h after onset of MCAo. Fluorometric quantitation of EB was performed 1 or 2 h later in six brain regions. Cerebral infarction volumes were quantitated from histopathological material at 72 h. EB extravasation first became grossly visible in the ipsilateral caudoputamen and neocortex following 3 h of MCAo, was grossly unapparent at 24-26 h, and was maximal at 48-50 h. Fluorescence quantitation confirmed that BBB opening was absent at 2-3 h but present at all later times. In the hemisphere ipsilateral to MCAo, a 179% mean increase in extravasation of EB (compared to sham rats) was measured at 4 h, 407% at 5 h, 311% at 26 h and 264% at 50 h. (in each case, P < 0.05 vs. sham). The volume of infarcted tissue at 72 h in this model was 163.6 +/- 7.7 mm3. Our results indicate that an initial, acute disruption of the BBB occurs between 3 and 5 h following MCAo, and that a later, more widespread increase in regional BBB permeability is present at 48 h. Regional measurement of Evans Blue extravasation offers a precise means of quantitating BBB disruption in focal cerebral ischemia; this method will be of considerable utility in assessing the BBB-protective properties of pharmacological agents.
采用一种灵敏的定量荧光法，探究短暂性大脑中动脉闭塞（MCAo）后血脑屏障（BBB）开放的时间进程和区域模式。雄性Sprague-Dawley大鼠用氟烷麻醉，通过经颈外动脉逆行插入涂有聚-L-赖氨酸的腔内尼龙缝线至颈内动脉和大脑中动脉，使其经历2小时的暂时性MCAo。通过伊文思蓝（EB）染料外渗判断血脑屏障的损伤情况，EB染料在MCAo发作后2、3、24或48小时给药。1或2小时后，对六个脑区进行EB的荧光定量分析。72小时时，从组织病理学材料中定量测定脑梗死体积。MCAo 3小时后，同侧尾壳核和新皮质中EB外渗首先明显可见，24 - 26小时时明显不明显，48 - 50小时时达到最大。荧光定量分析证实，2 - 3小时时血脑屏障未开放，但之后所有时间均开放。在MCAo同侧半球，4小时时EB外渗平均增加179%（与假手术大鼠相比），5小时时增加407%，26小时时增加311%，50小时时增加264%。（每种情况，与假手术组相比，P < 0.05）。该模型中72小时时梗死组织体积为163.6±7.7立方毫米。我们的结果表明，MCAo后3至5小时会发生血脑屏障的初始急性破坏，48小时时会出现后期更广泛的区域血脑屏障通透性增加。伊文思蓝外渗的区域测量提供了一种精确的方法来定量局灶性脑缺血中血脑屏障的破坏；该方法在评估药物制剂的血脑屏障保护特性方面将具有相当大的实用性。
