Scarpini E, Kreider B Q, Lisak R P, Meola G, Velicogna M E, Baron P, Beretta S, Buscaglia M, Ross A H, Scarlato G
Department of Neurology, Dino Ferrari Center for Neuromuscular Diseases, University of Milan, Italy.
Brain Res. 1988 Feb 9;440(2):261-6. doi: 10.1016/0006-8993(88)90994-8.
We describe a technique for the preparation of highly purified populations of Schwann cells (SC) from human fetal nerves. Cultures were prepared by chemical and mechanical dissociation of human fetal sciatic nerves by modification of the method of Kreider et al. developed for newborn rat nerve. A time course analysis of some SC-associated markers at different times in vitro was performed employing immunofluorescence (IF) and immunoperoxidase (IP) to determine the percentage of SC in culture and to evaluate the maintenance of specific SC characteristics. We compared this method with that of Askanas et al. which produces enriched SC cultures by utilizing successive re-explantation of the original nerve explant. After 48 h, approximately 90% of the cells were bipolar and S-100+ and over the next two weeks about 70-80% of cells were SC by cytologic and immunocytologic criteria. At 35 days, 35% were SC, whereas less than or equal to 2.5% of 35-day-old multi-explant cultures were SC. The SC obtained by this method displayed the typical morphological and immunological characteristics: they expressed surface laminin and nerve growth factor receptors, whereas fibronectin, which is localized on fibroblast surface, was absent.
我们描述了一种从人胎儿神经中制备高纯度雪旺细胞（SC）群体的技术。通过对Kreider等人开发的用于新生大鼠神经的方法进行改进，采用化学和机械方法解离人胎儿坐骨神经来制备培养物。利用免疫荧光（IF）和免疫过氧化物酶（IP）对体外不同时间的一些与SC相关的标志物进行时间进程分析，以确定培养物中SC的百分比并评估特定SC特征的维持情况。我们将此方法与Askanas等人的方法进行了比较，后者通过对原始神经外植体进行连续再移植来产生富集的SC培养物。48小时后，约90%的细胞为双极且S-100阳性，在接下来的两周内，根据细胞学和免疫细胞学标准，约70-80%的细胞为SC。在35天时，35%为SC，而35天大的多次移植培养物中SC占比小于或等于2.5%。通过这种方法获得的SC表现出典型的形态和免疫学特征：它们表达表面层粘连蛋白和神经生长因子受体，而成纤维细胞表面定位的纤连蛋白则不存在。
