Borlongan Cesar V, Hadman Martin, Sanberg Cyndy Davis, Sanberg Paul R
Department of Neurology, Medical College of Georgia, Augusta, GA 30912-3200, USA.
Stroke. 2004 Oct;35(10):2385-9. doi: 10.1161/01.STR.0000141680.49960.d7. Epub 2004 Sep 2.
To date, stem cell graft-mediated neuroprotection is equated with graft survival and secretion of neurotrophic factors in the brain. Here, we examined whether neuroprotection by systemically delivered human umbilical cord blood (HUCB) cells was dependent on their entry into the central nervous system in a rodent model of acute stroke.
Adult male Sprague-Dawley rats were subjected to right middle cerebral artery occlusion for 60 minutes. During the 1-hour occlusion, animals were randomly assigned to 1 of the following treatments: intravenous injection of HUCB (a subtherapeutic dose of 200,000 cells in 10 microL) with blood-brain barrier (BBB) permeabilizer (1.1 mol/L mannitol at 4 degrees C) or vehicle, intravenous vehicle alone, or intravenous mannitol alone. Behavioral tests, using elevated body swing test and passive avoidance test, were conducted at day 3 poststroke, and thereafter, animals were euthanized for: (1) immunohistochemical examination of HUCB, which were lentivirally labeled with green fluorescent protein; (2) cerebral infarction analysis using 2,3,5-triphenyl-tetrazolium chloride; and (3) enzyme-linked immunosorbent assay of trophic factors within the striatal region.
We did not detect intravenously administered low dose of HUCB cells in the brains of animals at day 3 after stroke even when cells were coinfused with a BBB permeabilizer (mannitol). However, HUCB-mannitol treatment significantly increased brain levels of neurotrophic factors, which correlated positively with reduced cerebral infarcts and improved behavioral functions.
Our data show that central nervous system availability of grafted cells is not a prerequisite for acute neuroprotection provided that therapeutic molecules secreted by these cells could cross the BBB.
迄今为止，干细胞移植介导的神经保护作用等同于移植细胞在脑内的存活及神经营养因子的分泌。在此，我们在急性脑卒中的啮齿动物模型中研究了全身递送的人脐带血（HUCB）细胞所提供的神经保护作用是否依赖于其进入中枢神经系统。
成年雄性Sprague-Dawley大鼠接受右侧大脑中动脉闭塞60分钟。在1小时的闭塞期间，将动物随机分配至以下治疗组之一：静脉注射HUCB（200,000个细胞，溶于10微升，为亚治疗剂量）并联合血脑屏障（BBB）通透剂（4℃下1.1摩尔/升甘露醇）或赋形剂，单独静脉注射赋形剂，或单独静脉注射甘露醇。在卒中后第3天进行行为测试，采用抬高身体摆动试验和被动回避试验，此后，对动物实施安乐死以进行：（1）对用绿色荧光蛋白进行慢病毒标记的HUCB进行免疫组织化学检查；（2）使用2,3,5-三苯基氯化四氮唑进行脑梗死分析；（3）对纹状体内区域的营养因子进行酶联免疫吸附测定。
即使细胞与BBB通透剂（甘露醇）共同输注，在卒中后第3天我们也未在动物脑内检测到静脉注射的低剂量HUCB细胞。然而，HUCB-甘露醇治疗显著提高了脑内神经营养因子水平，这与脑梗死面积减小及行为功能改善呈正相关。
我们的数据表明，移植细胞进入中枢神经系统并非急性神经保护的先决条件，前提是这些细胞分泌的治疗性分子能够穿过血脑屏障。
