多巴胺能神经元的体外培养及其纯度鉴定

CELL CULTURE IN VITRO OF DOPAMINERGIC NEURONS AND PURITY IDENTIFICATION

目的通过建立体外培养的多巴胺能神经元模型,应用于帕金森病(PD)的病因和发病机制等基础研究。方法用孕14d的胎鼠中脑嘴侧被盖部组织进行分散细胞原代培养,并用酪氨酸羟化酶(TH)免疫组化染色法鉴定其纯度。结果本实验可观察细胞历时20～30d的生长过程,并观察到细胞生长的第7～11d是细胞生长最旺盛的时期,最适宜加入处理因素观察细胞变化。培养10d的神经细胞进行TH免疫组化染色,可见胞浆被染成棕黄色的细胞遍布视野,阳性率达90%。结论体外培养的神经细胞可保持结构和功能上的某些特点,提供了体内生长过程在体外重现的可能,可广泛应用于PD病因和发病机制的研究。

? Objective To study the etiology and pathogenesis of Parkinson’s disease by establishing a model of the primary dissociated cell culture in vitro of dopaminergic neurons. Methods The rostral mesencephalic tegmentum of ratembryos on gestational day 14was used for the primary dissociated cell culture, and the purity was detected by using TH immunohistochemical staining. Results We successfully established a model in vitro of dopaminergic neurons and observed the growth and maturation of the cells in a period of one month. The cell growth was the most vigorous on 7d～11d: most cells were pyramidal or polypolar in shape, highly refractile and stereognostic, interconnected by a rich fiber network. Immunohistochemically, tyrosine hydroxylase (TH) positive neurons and fine dendrites were identified clearly. The stained neurons comprised about 90% of all the cells. The cultured cells were useful experiment model for carrying out in vitro studies. Conclusion Neuronal cells in culture can keep the characteristics of cell structure and function and thus provide the possibility of dupticating in vitro the growth process in vivo. The model can be widely used in the studyof the etiology and pathogenesis ofParkinson’s disease.〔