Studies on Expression-Mechanism (s) of Malignant Phenotypes Using Growth-Factor Gene-Transfer Methods

小鼠髓性白血病细胞系NFS-60的生长和发育依赖于几种造血生长因子，如白细胞介素-3（IL-3）、GM-CSF、G-CIF和促红细胞生成素。我们建立了一个只对IL-3有反应的亚克隆（1-7），并将鼠IL-3基因导入其中。从转染的亲本1-7亚克隆建立了10个具有自主增殖的IL-3非依赖性细胞系。从IL-3非依赖性细胞系分泌到条件培养基中的IL-3活性在小于10至280 U/mL之间变化。主要研究了两种代表性的转染子，产生最高IL-3活性的克隆1-7/7（#7）和产生最低活性的克隆I-7/5（#5）。只有一个拷贝的IL-3基因被引入到每个细胞系中，但是。它的反描述的层次是不同的，e. #7细胞表达的IL-3基因比#5细胞多约10倍。两种细胞系的细胞外IL-3活性差异显著，而细胞内IL-3活性差异不大。存在外源IL-3时#5细胞的生长曲线与不存在IL-3时观察到的生长曲线相似。抗IL-3抗血清不抑制转染细胞的增殖。这些结果表明，IL-3分泌外转染细胞可能不是强制性的自主生长，细胞内的IL-3可能在自分泌生长中起重要作用。同时，IL-3基因转染的细胞表现出显着的变化，膜鞘糖脂（GSL），特别是在酸性sialoGSL（神经节苷脂）。IL-3基因转染特征性地诱导了神经节苷脂GD 3的新出现，已知其与恶性转化密切相关。此外，在转染子中观察到中性GSL的相当大的变化，即。e.总量和某些成分显著增加。

The growth and development of a murine myeloid leukemia cell line, NFS-60, is dependent upon several hemopoietic growth factors such as interleukin-3 (IL-3), GM-CSF, G-CIF, and erythropoietin. We established a subclone (1-7) which was exclusively responsive to IL-3 and introduced into it a murine IL-3 gene. Ten IL-3-independent cell lines with autonomous proliferation were established from the transfected parental 1-7 subclone. The IL-3 activity, secreted from the IL3-independent cell lines into conditioned media, varied from less than 10 to 280 U/mL. Two representative transfectants, clone 1-7/7 (#7) producing the highest IL-3 activity and clone I-7/5 (#5) producing the lowest activity, were mainly investigated. Only a single copy of the IL-3 gene was introduced into each cell line, but. the level of its transdescription was different, i. e. the #7 cells expressed the IL-3 gene approximately ten times more than the #5 cells. A remarkable difference in extracellular IL-3 activity was found between the two cell lines while the intracellular IL-3 activity was not so different. The growth curve of the #5 cells in the presence of exogenous IL-3 was similar to that seen in the absence of IL-3. Anti-IL-3 antiserum did not inhibit the proliferation of the transfected cells. These results suggested that IL-3 secreted outside the transfected cells might not be obligatory for autonomous growth and that intracellular IL-3 might play an important role in autocrine growth. Simultaneously, the IL-3 gene-transfected cells exhibited remarkable changes in membrane glycosphingolipids (GSLs), especially in acidic sialoGSLs (gangliosides). The IL-3 gene transfection characteristically induced a new appearance of ganglioside GD3, which is known to be intimately related to malignant transformation. In addition, considerable changes of neutral GSLs were observed in the transfectants, i. e. the total amount and some constituents significantly increased.