Autocrine differentiation of PC12 cells mediated by retroviral vectors.
Autocrine differentiation of PC12 cells mediated by retroviral vectors.
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逆转录病毒载体介导的 PC12 细胞的自分泌分化。
DOI：
10.1159/000111833
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发表时间：
1990
影响因子：
2.9
通讯作者：
Breakefield,XO
中科院分区：
文献类型：
--
作者：
Short,MP;Rosenberg,MB;Ezzedine,ZD;Gage,FH;Friedmann,T;Breakefield,XO
使用含有 (3-半乳糖苷酶 (lac Z) 的细菌基因或小鼠 β-神经生长因子 (NGF) 的 cDNA 和新霉素抗性细菌基因的复制缺陷型逆转录病毒载体对大鼠嗜铬细胞瘤 PC12 细胞进行基因修饰。使用 lac Z 载体，获得了 PC12 细胞的克隆系，其中几乎 100% 的细胞稳定表达该组织化学标记。结果显示，表达β-半乳糖苷酶的 PC12 细胞或衍生的亚克隆 PC12-BAG 与 NGF 载体导致自分泌分化，这种分化在感染后数小时内发生，并在培养物中维持数周。表达神经突生长的细胞百分比高于用 NGF 载体感染的外源 NGF 细胞处理的 PC12 细胞。使用双位点酶免疫测定和对使用这些载体进行基因修饰的“原始”PC12 细胞进行生物测定，可将这种营养因子释放到培养基中，从而提供了一种方法：移植到动物体内后跟踪细胞的命运；测试移植的自分泌分化的 PC 12 细胞体内 NGF 和儿茶酚胺的递送；以及研究 NGF 对反应性细胞的长期作用，而不添加外源性 NGF。
Rat pheochromocytoma PC12 cells have been modified genetically by the use of replication-defective retroviral vectors containing either the bacterial gene for (3-galactosidase(lac Z)or cDNAs for mouse β-nerve growth factor (NGF) and the bacterial gene for neomycin resistance. Using thelac Zvector, clonal lines of PC12 cells were obtained in which almost 100% of cells stably expressed this histochemical marker. Infection of PC12 cells or the derived subclone PC12-BAG, which expresses β-galactosidase, with the NGF vectors resulted in autocrine differentiation as assessed by extensive neurite formation, which occurred within hours after infection and was maintained for weeks in culture. Neurite formation could be partially blocked by antibodies to NGF. The percentage of cells expressing neurite outgrowth was greater than that of PC12 cells treated with exogenous NGF. PC12 cells infected with the NGF vectors were shown to release this trophic factor into the medium using a two-site enzyme immunoassay and a bioassay on ''naive'' PC12 cells. PC12 cells genetically modified using these vectors provide a means to: follow the fate of the cells after transplantation into animals; test for delivery in vivo of NGF and catecholamines by grafted, autocrine-differentiated PC 12 cells; and study the long-term actions of NGF on responsive cells without adding exogenous NGF.