Goldfarb R H, Quigley J P
Cancer Res. 1978 Dec;38(12):4601-9.
Cultures of Rous sarcoma virus-transformed chick embryo fibroblasts (RSVCEF) produce 50-fold more of the protease plasminogen activator (PA), than do normal chick embryo fibroblasts. Treatment of RSVCEF cultures with the tumor promoter phorbol myristate acetate (PMA) further enhances (8- to 12-fold) the level of PA activity. Increased levels of PA activity in RSVCEF are observed as early as 1 to 2 hr after PMA treatment. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis demonstrates that the PA produced by PMA-treated cultures has a molecular weight identical to that of the PA produced by untreated cultures. PA induction by PMA cannot be accomplished in cell-free extracts, but requires protein synthesis in intact cells. Under serum-free conditions, PMA-treated RSVCEF secrete high levels of PA for 4 to 6 days and undergo pronounced morphological alterations. Modified culture conditions and the use of PMA to induce PA has allowed for the accumulation of large amounts of RSVCEF culture fluid and the subsequent purification of the enzyme. The sensitivity of transformed CEF to PMA and the generation of enhanced proteolytic activity in the cellular microenvironment may provide a model system to examine the role of both PA in malignant transformation and PMA in tumor promotion.
劳氏肉瘤病毒转化的鸡胚成纤维细胞（RSVCEF）培养物产生的蛋白酶纤溶酶原激活剂（PA）比正常鸡胚成纤维细胞多50倍。用肿瘤促进剂佛波酯肉豆蔻酸酯（PMA）处理RSVCEF培养物可进一步提高（8至12倍）PA活性水平。早在PMA处理后1至2小时就观察到RSVCEF中PA活性水平升高。十二烷基硫酸钠-聚丙烯酰胺凝胶电泳表明，经PMA处理的培养物产生的PA分子量与未经处理的培养物产生的PA分子量相同。PMA诱导PA不能在无细胞提取物中完成，而是需要完整细胞中的蛋白质合成。在无血清条件下，经PMA处理的RSVCEF可在4至6天内分泌高水平的PA，并发生明显的形态改变。改良的培养条件以及使用PMA诱导PA使得能够积累大量的RSVCEF培养液并随后纯化该酶。转化的CEF对PMA的敏感性以及细胞微环境中增强的蛋白水解活性的产生可能提供一个模型系统，以研究PA在恶性转化中的作用以及PMA在肿瘤促进中的作用。
