Analysis of the mechanism of chemical carcinogenesis and short-term screening of tumor-promoters using clonogenic a ssay

用N-丁基-N-（4-羟基丁基）亚硝胺（BBN）处理大鼠，观察其膀胱细胞在软琼脂中乳头瘤形成前的菌落形成，菌落形成的数量与摄入BBN的量有关。结果表明，糖精钠和几种已知的大鼠膀胱癌启动子氨基酸，在亚致癌剂量的BBN处理后，诱导大鼠膀胱细胞集落形成，提示促瘤活性与集落诱导活性之间可能存在相关性。在两阶段小鼠皮肤癌变实验中评估抗肿瘤启动子对肿瘤集落形成的影响。经维甲酸处理后，单次施用7,12-二甲基苯[a]蒽（DMBA）后，12-0-十四烷酚-13-乙酸酯（TPA）诱导的小鼠表皮细胞软琼脂集落形成受到抑制。膳食中醋酸视黄酯的影响。观察了去甲二氢愈创木酸（NDGA）和盐酸醌对大鼠膀胱细胞软琼脂集落形成的影响。经亚致癌剂量BBN处理后，糖精钠诱导的膀胱细胞软琼脂集落形成被视黄醇醋酸酯或NDGA抑制。提示两种药物可能具有抗肿瘤促进作用。因此，将克隆测定技术应用于肿瘤发生实验，有助于分析肿瘤发生的早期事件，并为发现新的肿瘤启动子和抗肿瘤启动子提供一种短期筛选方法。

Colony formation in soft agar was observed with bladder cells prior to papilloma development in rats treated with N- Butyl-N-(4-hydroxybutyl)nitrosamine(BBN), and the number of colonies formed was dependent on the quantity of BBN consumed. It was then demonstrated that sodium saccharin and several amino acids, which are known to act as rat bladder cancer promoters, induce colony formation of rat bladder cells after treatment with subcarcinogenic dose of BBN, suggesting that there may be a correlation between tumor-promoting activity and colony inducing activity.Effect of anti-tumor-promoter on colony formation was then assessed in two-stage mouse skin carcinogenesis experiment. Soft agar colony formation of mouse epidermal cells induced by 12-0- tetradecanoylphorbol-13-acetate(TPA) after single application of 7,12-dimethylbenz[a]anthracene(DMBA) was inhibited by treatment of skin with retinoic acid. Effect of dietary retinyl acetate. nordihydroguaiaretic acid(NDGA) and quinacrine hydrochloride on soft agar colony formation of rat bladder cells was also eaaluated. Soft agar colony formation of bladder cells induced by sodium saccharin after treatment with subcarcinogenic dose of BBN was inhibited by the administration of retinyl acetate or NDGA. suggesting the possible anti-tumor-promoting activity of the two agents.Thus, an application of clonogenic assay to carcinogenesis experiment could be useful to analyse early events in carcinogenesis and provide a short-term screening method for detecting new tumor-promoters and anti-tumor-promoters.