Milo G E, Oldham J W, Noyes I, Lehman T A, Kumari L, West R W, Kadlubar F F
Department of Physiological Chemistry, Ohio State University, Columbus, Ohio.
J Toxicol Environ Health. 1988;24(3):413-21. doi: 10.1080/15287398809531171.
Previous attempts to transform human foreskin fibroblasts in vitro with N-methylnitrosourea (MNU) or N-ethylnitrosourea (ENU) have been unsuccessful, and concurrent treatment with cocarcinogens or tumor promotors and either MNU or ENU have also failed to produce a neoplastic response. The present study was undertaken to test the effect of sodium saccharin on MNU- or ENU-induced cell transformation. Saccharin alone was not effective in inducing the growth of colonies in soft agar (anchorage-independent growth). However, concurrent treatment with saccharin (50 micrograms/ml, nontoxic dose) and MNU or ENU (29 micrograms/ml or 44 micrograms/ml, respectively) was effective in inducing transformation (greater than 300 colonies/10(5) cells), but only when the cells were treated with saccharin after being released from a G1 block (amino acid deprivation) and followed by MNU or ENU treatment in early S phase. In contrast to results obtained with other chemical carcinogens, transformation frequencies induced by saccharin and MNU or ENU were only slightly decreased in the absence of insulin, which is normally required for growth in this system. Saccharin-MNU- or saccharin-ENU-treated cells that exhibited growth in soft agar also exhibited cellular invasiveness in 9-d-old embryonic chick skin in vitro. In addition, these cells reacted with a monoclonal antibody prepared against a molecular weight 115,000 sarcoma-cell surface-associated glycoprotein and also developed tumors in nude mice. These data demonstrate the cell-cycle-dependent cocarcinogenic potential of saccharin and MNU or ENU in cultured human skin fibroblasts.
以往尝试用N-甲基亚硝基脲（MNU）或N-乙基亚硝基脲（ENU）在体外转化人包皮成纤维细胞均未成功，同时使用促癌剂或肿瘤促进剂与MNU或ENU联合处理也未能产生肿瘤反应。本研究旨在测试糖精钠对MNU或ENU诱导的细胞转化的影响。单独使用糖精在软琼脂中诱导集落生长（不依赖贴壁生长）无效。然而，糖精（50微克/毫升，无毒剂量）与MNU或ENU（分别为29微克/毫升或44微克/毫升）联合处理在诱导转化方面是有效的（大于300个集落/10⁵个细胞），但前提是细胞在从G1期阻滞（氨基酸剥夺）中释放后用糖精处理，随后在S期早期用MNU或ENU处理。与其他化学致癌物的结果相反，在缺乏胰岛素的情况下，糖精与MNU或ENU诱导的转化频率仅略有下降，而在该系统中生长通常需要胰岛素。在软琼脂中生长的经糖精-MNU或糖精-ENU处理的细胞在体外9日龄鸡胚皮肤中也表现出细胞侵袭性。此外，这些细胞与针对分子量为115,000的肉瘤细胞表面相关糖蛋白制备的单克隆抗体发生反应，并且在裸鼠中也形成了肿瘤。这些数据证明了糖精与MNU或ENU在培养的人皮肤成纤维细胞中具有细胞周期依赖性的促癌潜力。
