Marone M, Quiñones-Jenab V, Meiners S, Nowakowski R S, Ho S Y, Geller H M
Department of Pharmacology,UMDNJ-Robert Wood Johnson Medical Shool, Piscataway 08854, USA.
Dev Neurosci. 1995;17(5-6):311-23. doi: 10.1159/000111301.
Evidence from retroviral marking techniques and immortalized cell lines indicates that multipotential stem cells exist in many areas of the developing central nervous system. However, the factors that influence the commitment of these stem cells into distinct neuronal or glial lineages are not known. We have created an immortalized hypothalamic cell line derived from embryonic day 14 hypothalamic cells with a replication-defective retroviral construct containing a temperature-sensitive allele (tsA58) of the large T antigen of the simian virus 40. The clonality of this cell line, which we have named V1, was established by single cell cloning and by Southern blot analysis. V1 cells exhibit two different morphologies: the vast majority of cells are flat and stellate, and a smaller number are phase-bright round cells with processes. V1 cells express nestin and neural-cell adhesion molecule, typical of proliferating neuroepithelial cells. They also express glial fibrillary acidic protein and S100 as well as the low molecular weight neurofilament protein. In addition, the phase-bright, process-bearing V1 cells stain intensely for many typical neuronal proteins, such as low, medium and high molecular weight neurofilament proteins, tau protein, microtubule-associated protein-2, and neuron-specific enolase. The phase-bright cells also have condensed chromatin and display mitotic spindles, indicating that they are in mitosis. When V1 cells are transferred from the permissive temperature (33 degrees C) to the restrictive temperature (39 degrees C), there is a decrease in expression of NF-L and an increase in expression of NF-H and glial fibrillary acidic protein in the flat V1 cells. The enhanced expression of neuronal antigens in mitotically active V1 cells is novel and may represent a more general property of the differentiation process. We suggest that V1 cells arise from a mixed neural/glial neuroepithelial progenitor cell that expresses both neuronal- and glial-specific proteins in the developing hypothalamus.
来自逆转录病毒标记技术和永生化细胞系的证据表明，多能干细胞存在于发育中的中枢神经系统的许多区域。然而，影响这些干细胞分化为不同神经元或神经胶质谱系的因素尚不清楚。我们用含有猿猴病毒40大T抗原温度敏感等位基因（tsA58）的复制缺陷型逆转录病毒构建体，从胚胎第14天的下丘脑细胞中创建了一个永生化下丘脑细胞系。我们将这个细胞系命名为V1，其克隆性通过单细胞克隆和Southern印迹分析得以确定。V1细胞呈现出两种不同的形态：绝大多数细胞扁平呈星状，少数是有突起的亮相圆形细胞。V1细胞表达巢蛋白和神经细胞黏附分子，这是增殖性神经上皮细胞的典型特征。它们还表达胶质纤维酸性蛋白和S100以及低分子量神经丝蛋白。此外，有突起的亮相V1细胞对许多典型的神经元蛋白染色强烈，如低、中、高分子量神经丝蛋白、tau蛋白、微管相关蛋白-2和神经元特异性烯醇化酶。亮相细胞还具有浓缩的染色质并显示有丝分裂纺锤体，表明它们处于有丝分裂状态。当V1细胞从允许温度（33摄氏度）转移到限制温度（39摄氏度）时，扁平V1细胞中NF-L的表达下降，NF-H和胶质纤维酸性蛋白的表达增加。有丝分裂活跃的V1细胞中神经元抗原表达的增强是新颖的，可能代表了分化过程中更普遍的特性。我们认为V1细胞源自一种混合的神经/胶质神经上皮祖细胞，该祖细胞在发育中的下丘脑中同时表达神经元和胶质特异性蛋白。
