造血干细胞与神经生长因子联合移植治疗脑梗死  

Hematopoietic stem cell transplantation combined with nerve growth factor for treatment of cerebral infarction

作　　者：李永秋[1] 马洪颖[1] 刘长杰[2] 周红霞 张冬森[1] 

机构地区：[1]河北医科大学唐山临床学院神经内科,河北省唐山市063000 [2]河北省玉田县医院神经内科,河北省唐山市064100 [3]河北省遵化市第二医院内科,河北省遵化市064200

出　　处：《中国组织工程研究与临床康复》2009年第32期6231-6235,共5页Journal of Clinical Rehabilitative Tissue Engineering Research

基　　金：2009年河北省科技支撑计划课题(09276101D-10)~~

摘　　要：背景:目前采用外周血造血干细胞移植治疗脑缺血方面的研究还比较少,缺乏大量、系统的实验数据。神经生长因子可调控神经元再生所需的微环境,对神经细胞的存活、增殖起重要作用。目的:观察造血干细胞移植后,神经生长因子对其在脑梗死大鼠脑内的存活、分化及神经功能的影响。设计:随机对照动物实验。材料:清洁级成年雄性SD大鼠120只,随机分为3组:单纯细胞移植组、细胞移植+神经生长因子组、模型对照组,40只/组。神经生长因子为北大之路药业有限公司产品。方法:大鼠颈部皮下注射重组人粒细胞集落刺激因子后,密度梯度离心法分离培养造血干细胞,移植前行Brdu标记。3组大鼠根据改良Longa线栓法建立大脑中动脉闭塞模型,造模后7d,大鼠麻醉后暴露前囟,选取注射位点:前囟后方1.2mm,中线向右旁开3mm,硬膜下4mm。单纯细胞移植组缓慢注入1×106个造血干细胞,细胞移植+神经生长因子组注入1×106个造血干细胞+200ng神经生长因子,模型对照组给予等量磷酸盐缓冲液。主要观察指标:采用神经功能缺损评分检测神经功能的改善情况,免疫组化染色观察梗死半球Brdu/CD34、Brdu/Nestin、Brdu/NSE、Brdu/vWF双阳性细胞的分布,TCC染色观察梗死体积的变化。结果:与移植前比较,移植后1,2周仅细胞移植+神经生长因子组神经功能缺损评分均明显下降(P<0.05),移植后3,4周各组神经功能缺损评分均明显下降(P<0.05或0.01),且细胞移植+神经生长因子组下降幅度尤为显著(P<0.01)。移植后1周,细胞移植+神经生长因子组Brdu/CD34、Brdu/Nestin、Brdu/NSE、Brdu/vWF双阳性细胞数明显多于单纯细胞移植组(P<0.01);第4周两组未见Brdu/CD34双阳性细胞,Brdu/Nestin较1周时减少,Brdu/NSE、Brdu/vWF双阳性细胞均明显增多(P<0.05),且细胞移植+神经生长因子组明显多于单纯细胞移植组(P<0.01);模型对照组始终未见Brdu双阳性细BACKGROUND： Studies concerning peripheral blood hematopoietic stem cell （HSC） transplantation for cerebral ischemia are few, and there are lack of abundant professional experimental data. Nerve growth factor （NGF） can regulate microenvironment of neuron regeneration, and play important roles in survival and proliferation of neural cells. OBJECTIVE： To explore the influence of NGF on the survival, migration and differentiation of HSCs following transplantation in rats with cerebral infarction. DESIGN： Randomized controlled animal study. MATERIALS： A total of 120 clean adult male Sprague Dawley rats were equally and randomly divided into 3 groups, cell transplantation, cell transplantation ＋ NGF, and model control groups. NGF purchased from Beida Zhilu Pharmacological Co., Ltd., was used in this study. METHODS： Following subcutaneous injection of recombinant human granulocyte colony-stimulating factor in the rat neck, HSCs were isolated and cultured by the density gradient centrifugation, and labeled by Brdu prior to transplantation. Rat models of middle cerebral artery occlusion were established by the modified Longa in three groups. Seven days later, rats were anesthetized and the anterior fontanelle was exposed to select injection sites： 1.2 mm posterior to the anterior fontanelie, 3 mm right of the median line, and 4 mm below the dura mater. HSCs （1×10^6） were slowly infused into the rats in the cell transplantation group. HSCs （1×10^6） ＋ NGF （200 ng） were infused into rats of the cell transplantation ＋ NGF group. An equal volume of phosphate buffer saline was given in rats of the model control group. MAIN OUTCOME MEASURES： Improvement of neural function was assessed by neurological deficit scores. Distribution of Brdu/CD34, Brdu/Nestin, Brdu/NSE, Brdu/vWF double positive cells was observed in infarct hemisphere by immunohistochemical staining. Infarction volume was measured by TTC staining. RESULTS： Neurological deficit scores were significantly reduced in the cell t

关 键 词：造血干细胞 神经生长因子 移植 脑梗死 

分 类 号：R394.2[医药卫生—医学遗传学]