【作者】 林志钦；

【导师】 王玮；

【作者基本信息】 福建医科大学 ， 人体解剖与组织胚胎学， 2005， 硕士

【摘要】 目的1. 从新生大鼠海马中分离培养并鉴定神经干细胞,为神经干细胞的体外移植作 准备。2. 制作血管性痴呆大鼠动物模型,为研究血管性痴呆的发病机理和防治建立基 础。3. 探讨神经干细胞在血管性痴呆大鼠侧脑室的存活、迁移和分化,从而为神经 干细胞的体内存活、迁移和分化机理研究和临床应用提供实验依据。方法1. 利用无血清培养和单细胞克隆技术从新生大鼠海马分离具有单细胞克隆能力的细胞群并连续传代培养,获得大量细胞,采用免疫荧光细胞化学技术检测巢蛋白(Nestin)和分化后特异性成熟神经细胞抗原表达。2. 采用永久性结扎大鼠双侧颈总动脉法(2-VO)建立血管性痴呆大鼠动物模型,结扎后30 天,用Morris 水迷宫定位航行试验和空间探索试验来检测大鼠的学习记忆能力变化,并在光镜下观察大鼠海马组织结构的变化。3. 采用脑立体定位技术将5-溴-2’-脱氧尿苷( BrdU)标记的神经干细胞移植入血管性痴呆大鼠侧脑室内,饲养4 周后取脑,应用免疫荧光化学技术检测移植后细胞的存活、迁移和分化。结果1. 从新生大鼠海马分离得到的细胞经无血清培养后,具有连续传代能力,能表 达巢蛋白Nestin,经诱导分化后能表达神经元、神经胶质细胞的特异性抗原, 说明大鼠海马内存在神经干细胞。更多还原

【Abstract】 Objective1. Establish vascular dementia rat model.2. To isolated neural stem cells from hippocampus of neonatal rats and transplant into the lateral ventricle of adult vascular dementia rats.3. To investigate the survival, migration and differentiation of neural stem cells whichtransplanted into the lateral ventricle of vascular dementia rats.MethodsThe NSCs which had ability to form clones were isolated from hippocampus ofneonatal rats by using serum-free cell culture and single cell clone test. Clones werepassaged continuously in order to acquire a large number of clones, andimmunofluorescence was used to detect Nestin antigen of the clone and specificantigen of mature neural cells after the clone was induced to differentiate. Thevascular dementia rat model was reproduced by permanent occlusion of bilateralcommon carotid arteries of rats. 30 days after permanent occlusion the arteries,abilities of learning and memory of rats were tested by using the Morris water maze.The histomorphology of hippocampus was observed and analyzed by microscopy. Thecultured neural stem cells were labeled by BrdU and transplanted into lateral ventricleof adult vascular dementia rats. Immunofluorescence was used to detect survival,migration and differentiation of neural stem cells.Results Cells were isolated from hippocampus of neonatal rats express Nestinantigen and can differentiate to neuron, astrocytes. The average escape latency ofmodel is longer than that in control in place navigation test. For spatial probe test, itwas seen that the lingering time in platform area of model rat is shorter than that ofcontrol. Loss of cells was seen in CA1 subfield of hippocampus 30 days afterocclusion of bilateral common carotid arteries on rats. Neural stem cells weretransplanted into lateral ventricle of vascular dementia rats. After four weeks,immunofluorescence was used to detect survival, migration and differentiation ofneural stem cells. The results show neural stem cells can survive, migrate to cortexand hippocampus, and differentiate to neuron and astroglialcells.ConculsionThe kind of rat model reproduced by permanent occlusion of bilateral commoncarotid arteries of rats can result vascular dementia. Cells which express nestinantigen are multipotent cells and have ability to self-renew, and we believe there haveneural stem cells in hippocampus. The neural stem cells which transplanted intolateral ventricle of vascular dementia rats can survive, migrate and differentiate.更多还原