Mori Yukio, Koide Akihiro, Kobayashi Yoshinori, Morimura Keiichirou, Kaneko Masahiro, Fukushima Shoji
Laboratory of Radiochemistry, Gifu Pharmaceutical University, 6-1, Mitahora-higashi 5-chome, Gifu 502-8585, Japan.
Mutagenesis. 2002 May;17(3):251-6. doi: 10.1093/mutage/17.3.251.
In order to elucidate the mechanism underlying enhancement by ethanol of N-nitrosodiethylamine (DEN)- and N-nitrosomethylbenzylamine (NMBA)-induced esophageal tumorigenesis in rats, hepatic levels of cytochrome P-450 (CYP) enzymes, mutagenic activation of several N-nitrosamines and three kinds of UDP-glucuronyltransferase (UDPGT) activities were assayed in F344 rats. Immunoblot analyses of microsomal CYP proteins revealed induction of CYP2E1 (approximately 2-fold), but not CYP2B1/2, 1A1/2 or 3A2, by treatment with 10% ethanol in the drinking water for 2 weeks. In contrast, s.c. treatment with 0.5 mg/kg NMBA three times per week for 2 weeks produced no significant alterations in the levels of these CYP species. Ethanol treatment also elevated the mutagenic activities of N-nitrosodimethylamine (DMN), DEN and N-nitrosopyrrolidine (NPYR) in strain TA100 up to 2.1-, 1.6- and 2.3-fold above each control, respectively. However, this was not the cases for four N-nitrosamines, including NMBA, in strain TA100 and two heterocyclic amines and aflatoxin B(1) in strain TA98. In addition, ethanol did not affect UDPGT activities towards 4-nitrophenol, bilirubin and testosterone. Hepatic CYP species responsible for mutagenic activation of selected N-nitrosodialkylamines were confirmed by use of specific CYP inducers and inhibitors with the liver from F344 and Wistar rats, indicating that DMN, DEN and NMBA are selectively activated by CYP2E1, predominantly by CYP2E1 with a slight contribution by CYP2B2 and selectively by CYP2B1/2, respectively. These results demonstrate that ethanol exerts an enhancing effect on mutagenic activation by CYP2E1 of DMN, DEN and NPYR, but does not affect that of NMBA and the other carcinogens by CYP2B1/2, 1A1/2 and 3A2 and UDPGT1A1, 1A6 and 2B1 activities. Consequently, this suggests that enhancement by ethanol of DEN-induced esophageal carcinogenesis in F344 rats can be attributed to an increase in hepatic activation during the initiation phase, but that of NMBA-induced tumorigenesis is not attributable to metabolic activation and inactivation via glucuronidation in liver.
为了阐明乙醇增强大鼠中N-亚硝基二乙胺（DEN）和N-亚硝基甲基苄胺（NMBA）诱导的食管肿瘤发生的潜在机制，在F344大鼠中测定了细胞色素P-450（CYP）酶的肝脏水平、几种N-亚硝胺的诱变活化以及三种尿苷二磷酸葡萄糖醛酸转移酶（UDPGT）活性。微粒体CYP蛋白的免疫印迹分析显示，通过在饮用水中用10%乙醇处理2周，CYP2E1被诱导（约2倍），但CYP2B1/2、1A1/2或3A2未被诱导。相比之下，每周3次皮下注射0.5mg/kg NMBA，共2周，这些CYP种类的水平没有显著变化。乙醇处理还使TA100菌株中N-亚硝基二甲胺（DMN）、DEN和N-亚硝基吡咯烷（NPYR）的诱变活性分别比各自对照提高了2.1倍、1.6倍和2.3倍。然而，对于TA100菌株中的四种N-亚硝胺（包括NMBA）以及TA98菌株中的两种杂环胺和黄曲霉毒素B1，情况并非如此。此外，乙醇不影响UDPGT对4-硝基苯酚、胆红素和睾酮的活性。通过使用特定的CYP诱导剂和抑制剂以及来自F344和Wistar大鼠的肝脏，证实了负责选定N-亚硝基二烷基胺诱变活化的肝脏CYP种类，表明DMN、DEN和NMBA分别被CYP2E1选择性活化、主要被CYP2E1活化且有CYP2B2的轻微贡献以及被CYP2B1/2选择性活化。这些结果表明，乙醇对DMN、DEN和NPYR的CYP2E1诱变活化有增强作用，但不影响NMBA和其他致癌物通过CYP2B1/2、1A1/2和3A2以及UDPGT1A1、1A6和2B1活性的诱变活化。因此，这表明乙醇对F344大鼠中DEN诱导的食管癌发生的增强作用可归因于起始阶段肝脏活化的增加，但NMBA诱导的肿瘤发生的增强作用不归因于肝脏中的代谢活化和通过葡萄糖醛酸化的失活。
