摘要
目的：目前对于体外诱导分化的多巴胺能神经元的生物学功能、体内存活状况,以及与胚胎中脑多巴胺能神经元移植治疗帕金森病大鼠的疗效比较方面尚缺乏深入的研究。观察神经干细胞诱导分化的多巴胺能神经元移植治疗帕金森病大鼠的作用,并与胚胎中脑多巴胺能神经元组织移植治疗帕金森病大鼠的疗效进行比较。方法：实验于2006-06/2007-09在中山大学附属第一医院完成。①实验材料：健康成年雄性SD大鼠及孕14-15d的SD大鼠由中山大学动物实验中心提供,实验过程中对动物处置符合动物伦理学标准。②实验方法：在含表皮生长因子及碱性成纤维细胞生长因子的无血清培养液中培养胚胎大鼠中脑神经干细胞。经传代扩增后,在含白细胞介素1a、白细胞介素11、白血病抑制因子、胶质细胞源性神经营养因子的诱导分化液中向多巴胺能神经元分化,并进行免疫细胞化学鉴定和流式细胞仪检测。参考Sauer和Lee等方法制备帕金森病大鼠模型,并将造模成功大鼠随机分组,每组9只：分化细胞组向每一坐标点注入1× 10^11 L^-1神经干细胞诱导分化的多巴胺能神经元;胚胎中脑组注入1× 10^11 L^-1胚胎中脑多巴胺能神经元;手术对照组注入DMEM/F12细胞培养液。③实验评估：移植术后10、20、40、60d诱发大鼠不对称旋转行为以观察治疗效果,并对移植区进行酪氨酸羟化酶免疫组织化学检测以了解移植细胞体内存活的情况。结果：①大鼠神经干细胞球在诱导分化液中呈贴壁生长,球内细胞从球体中央逐渐向四周分化扩展出形态各异的细胞。免疫细胞化学染色显示,分化细胞中含有酪氨酸羟化酶染色阳性细胞。流式细胞仪检测诱导分化6d的细胞中酪氨酸羟化酶染色阳性细胞的比率为（16.7±2.8）%。②移植后20d分化细胞组大鼠的不对称旋转行为开始明显下降（P〈0.05）。移植后40d�
AIM： Biological function and in vivo survival of differentiated dopaminergic neurons （DNs）, and comparison to the transplantation of embryonic mesencephalic DNs for treating Parkinson＇s disease rats are seldom studied. This study investigated the effect of transplantation of dopaminergic neurons differentiated from neural stem cells （NSCs） into Parkinson＇s disease rats, and compared to the transplantation of embryonic mesencephalic DNs.
METHODS： Experiments were performed at the First Affiliated Hospital of Sun Yat-sen University between June 2006 and September 2007. （1）Healthy adult male SD rats and pregnant 14-15 day SD rats were provided by Animal Experiment Center of Sun Yat-sen University. Animal intervention met the animal ethical staodards.（2）NSCs derived from embryonic rats were cultivated in serum-free culture solution containing epidermal growth factor and basic fibroblast growth factor. The cells were passaged and differentiated to DNs in differentiate solution containing interleukin （IL） 1a, IL11, leukemia inhibitory factor （LIF） and glial cell line-derived neurotrophic factor （GDNF）. The differentiated cells were detected by immunocytochemistry and flow cytometry. Rat models of Parkinson＇s disease were established according to the methods of Sauer and Lee et al. Successful models were randomly divided into 3 groups, 9 in each group. Differentiated DNs （1 × 10^11 L^-1） were injected into rats of differentiated DNs group. Embryonic mesencephalic DNs （× 10^11 L^-1） were injected into rats of embryonic mesencephalic DNs group. DMEM/F12 medium was injected into rats of control group. （3）The rotational asymmetry of Parkinson＇s disease rats was observed 10, 20, 40 and 60 days after the surgery to assess the effect. In transplanted areas, Tyrosine Hydroxylase （TH） was detected with immunocytochemistry to assess the survival of transplanted cells in vivo.
RESULTS： （1）In differentiate solution, the rat neural stem cell globes became attaching
出处
《中国组织工程研究与临床康复》
CAS
CSCD
北大核心
2008年第16期3019-3023,共5页
Journal of Clinical Rehabilitative Tissue Engineering Research
基金
国家自然科学基金资助(30300115)~~
