Platika D, Boulos M H, Baizer L, Fishman M C
Proc Natl Acad Sci U S A. 1985 May;82(10):3499-503. doi: 10.1073/pnas.82.10.3499.
In an attempt to immortalize the gene products of single neurons, somatic cell hybrids were produced by fusion of embryonic rat dorsal root ganglion (DRG) neurons with mouse neuroblastoma cells. Embryonic day 13 rat DRGs were fused with mouse neuroblastoma cells deficient in hypoxanthine phosphoribosyltransferase (HPRT; IMP:pyrophosphate phosphoribosyltransferase, EC 2.4.2.8). The hybrid cells were selected in medium with 100 microM hypoxanthine/1 microM aminopterin/12 microM thymidine to eliminate the neuroblastoma cells and with cis-hydroxyproline to retard fibroblast growth. Of the 17 lines derived, 4 manifested neuronal properties and were cloned. These lines retain both rat and mouse chromosomes and synthesize characteristic rat and mouse isoenzymes. Neuronal gangliosides, action potentials, and extensive neurite-like processes are exhibited by these hybrid cells, properties characteristic of DRG neurons but not of the neuroblastoma parent. Each line manifests a unique combination of action-potential properties and cell-surface markers, suggesting the selective expression of subsets of DRG neuronal genes. All of these neuronal properties are expressed constitutively, without the need for chemical induction or mitotic inhibition, and stably, without diminution after at least 5 months in culture. These lines may prove useful in the identification and isolation of gene products that characterize individual or small subsets of DRG neurons.
为了使单个神经元的基因产物永生化，通过将胚胎大鼠背根神经节（DRG）神经元与小鼠神经母细胞瘤细胞融合产生了体细胞杂种。将胚胎第13天的大鼠DRG与缺乏次黄嘌呤磷酸核糖基转移酶（HPRT；IMP：焦磷酸磷酸核糖基转移酶，EC 2.4.2.8）的小鼠神经母细胞瘤细胞融合。杂种细胞在含有100 microM次黄嘌呤/1 microM氨基蝶呤/12 microM胸苷的培养基中进行筛选，以消除神经母细胞瘤细胞，并加入顺式羟脯氨酸以抑制成纤维细胞生长。在衍生出的17个细胞系中，有4个表现出神经元特性并进行了克隆。这些细胞系保留了大鼠和小鼠的染色体，并合成特征性的大鼠和小鼠同工酶。这些杂种细胞表现出神经元神经节苷脂、动作电位和广泛的神经突样突起，这些是DRG神经元而非神经母细胞瘤亲本所特有的特性。每个细胞系都表现出动作电位特性和细胞表面标志物的独特组合，表明DRG神经元基因子集的选择性表达。所有这些神经元特性都是组成性表达的，无需化学诱导或有丝分裂抑制，并且是稳定表达的，在培养至少5个月后不会减弱。这些细胞系可能有助于鉴定和分离表征DRG神经元个体或小亚群的基因产物。
