量子点标记神经生长因子纳米化颗粒修饰羊膜间充质干细胞移植治疗大鼠实验性脑损伤  

Transplanting amniotic-derived mesenchymal stem cells modified with quantum dot-tagged nerve growth factor nanoparticles in the treatment of experimental brain injury in rats

作　　者：王公平[1] 杨波[1] 关方霞[2] 杜英[3] 常克亮[1] 宋来君[1] 胡祥 曾光伟[5] 

机构地区：[1]郑州大学第一附属医院神经外科 [2]郑州大学生物工程系,河南省郑州市450001 [3]郑州大学基础医学院微生物与免疫学教研室,河南省郑州市450052 [4]深圳市北科细胞工程研究所,广东省深圳市518000 [5]河南科技大学第一附属医院肿瘤科,河南省洛阳市471003

出　　处：《中国组织工程研究与临床康复》2008年第10期1806-1810,共5页Journal of Clinical Rehabilitative Tissue Engineering Research

基　　金：河南省医学科技创新人才工程项目(2005018);郑州大学优秀博士生培育基金(2006008)~~

摘　　要：目的：量子点是由Ⅱ-Ⅵ族元素或Ⅲ-V族元素组成的纳米颗粒,因三维上受到量子限制而表现出独特的光学特征,其标记的神经生长因子纳米化颗粒由于是纳米级,易通过细胞生物膜。实验观察负载此纳米颗粒的羊膜间充质干细胞移植治疗大鼠脑损伤的效果。方法：实验于2007-01/10在郑州大学完成。①材料：SPF级健康成年Wistar大鼠40只,按体质量编号分为4组：神经生长因子细胞组、常规细胞组、损伤模型组、培养基对照组,10只/组,实验过程中对动物的处置符合动物伦理学标准。羊膜间充质干细胞来自健康产妇胎盘羊膜,产妇对实验知情同意,实验经医院医学伦理委员会批准。量子点神经生长因子纳米粒由兰州大学功能有机分子化学国家重点实验室协助构建。②实验方法：向羊膜间充质干细胞中加入含10%胎牛血清、20μg/L碱性成纤维生长因子的DMEM/F12培养基,置于37℃、体积分数为0.05的CO2饱和湿度培养箱中培养,待细胞达80%～90%融合时胰酶消化传代。取传至第3代细胞,分别加入终浓度为20,40,60μg/L的量子点标记神经生长因子纳米粒进行修饰处理72h,并设立空白对照和空载量子点对照。4组大鼠均采用Feeney自由落体法建立脑损伤模型,神经生长因子细胞组于大鼠损伤部位注入经40mg/L量子点标记神经生长因子纳米粒修饰的羊膜间充质干细胞悬液10μL（约4.0×10^8个细胞）,常规细胞组注入等量单纯量子点标记的羊膜间充质干细胞悬液,培养基对照组注入等量DMEM/F12细胞培养基,损伤模型组不给予移植操作。③实验评估：经量子点标记的神经生长因子纳米粒修饰后,MTT法检测细胞活性,荧光显微镜观察量子点在细胞内的分布,免疫细胞化学鉴定细胞分化。细胞移植后对各组大鼠运动及感觉功能进行评分,免疫组织化学染色及荧光显微镜观察量子点荧光化羊膜间充质干细胞�AIM： Quantum dot, composed of Ⅱ-Ⅳ family elements or Ⅲ-Ⅳ family elements, is one kind of important nanoparticles which bears unique optical properties confined by tridimensional restriction of quantum. As the quantum dot-tagged nerve growth factor （NGF） nanoparticles belong to nanometer grade, it is easy to pass through the biomembrane of cells. This experiment aims to explore the effect of treating brain injured rats by transplanting anmiotic-derived mesenchymal stem cells （AD-MSCs） carried with such nanoparticles. METHODS： This experiment was undertaken in Zhengzhou University from January to October in 2007.①Materials： Forty SPF healthy Wistar rats were evenly divided into four groups according to their body weight： NGF nanoparticle cell group, normal cell group, brain injury model group and control group of nutrient medium. Rats in the experiment were disposed according to the animal ethics. Human AD-MSCs were extracted from the placenta amniosis of healthy parturient by professor Yang Bo working in the First Affiliated Hospital of Zhengzhou University. All the parturients signed informed consents. Moreover, the experiment was approved by the Medical Ethical Committee of the hospital. Quantum dot-tagged NGF nanoparticles were constructed with the assistance of the State Key Laboratory of Applied Organic Chemistry of Lanzhou University.②Empirical methods： AD-MSCs were added to nutritive medium DMEM/FI2 which contained 10% calf serum and 20 ta g/L basic fibroblast growth factor. Cells were then cultivated in an incubator of saturated humidity at 37℃, with 0.05 volume fraction of CO2 nutritive medium. When 80%-90% cells were mixed, trypsinization and was underwent and cells were subcultured. The cells in the third generation was added with quantum dot-tagged NGF nanoparticles at different final concentrations of 20, 40, 60 μg/L respectively, then modified 72 hours. Blank controls and empty quantum dot controls were set up. Brain injured models were constructed to all the rats in

关 键 词：羊膜间充质干细胞 神经生长因子 纳米 量子点标记 脑损伤 

分 类 号：R318[医药卫生—生物医学工程]