Imaoka S, Yoneda Y, Matsuda T, Degawa M, Fukushima S, Funae Y
Laboratory of Chemistry, Osaka City University Medical School, Osaka, Japan.
Biochem Pharmacol. 1997 Sep 15;54(6):677-83. doi: 10.1016/s0006-2952(97)00216-5.
We investigated the mutagenic activation of 2-naphthylamine (2-NA), 3,2'-dimethyl-4-aminobiphenyl (DMAB), and 3,3'-dichlorobenzidine (DCB), bladder carcinogens, by renal and bladder microsomes and by purified P450s using the umu gene expression system, which detects DNA damage. Mouse renal microsomes had high mutagenic activation toward DCB and low activity toward 2-NA. Purified mouse Cyp4b1 also had high mutagenic activity toward DCB. Anti-Cyp4b1 antibody efficiently inhibited DCB bioactivation by mouse renal microsomes with a high Cyp4b1 content. Lauric acid, a substrate of Cyp4b1, efficiently inhibited DCB bioactivation by renal and bladder microsomes of the mouse and by purified Cyp4b1. To assess the contribution of CYP4B1 to bladder carcinoma, further investigation was done with the umu test and an immunochemical study. Ten forms of purified rat P450s including rat CYP4B1 were used in the umu test for 2-NA, DMAB, and DCB. CYP4B1 had the highest activity toward DMAB and DCB. Other P450s had activities of less than 20% that of CYP4B1. CYP4B1 also activated 2-NA, but its activity was about 10% of that toward DMAB or DCB. Rat bladder epithelium was stained specifically with anti-Cyp4b1 antibody, indicating the presence of CYP4B1 in the rat bladder mucosa. Also, CYP4B1 mRNA was detected by northern blotting and reverse transcription-polymerase chain reaction (RT-PCR). These findings suggested that CYP4B1 could contribute to the initiation of carcinogenesis in rat and mouse bladder by activation of aromatic amines.
我们使用检测DNA损伤的umu基因表达系统，研究了肾微粒体和膀胱微粒体以及纯化的细胞色素P450（P450s）对膀胱致癌物2-萘胺（2-NA）、3,2'-二甲基-4-氨基联苯（DMAB）和3,3'-二氯联苯胺（DCB）的诱变激活作用。小鼠肾微粒体对DCB具有高诱变激活活性，而对2-NA的活性较低。纯化的小鼠Cyp4b1对DCB也具有高诱变活性。抗Cyp4b1抗体能有效抑制Cyp4b1含量高的小鼠肾微粒体对DCB的生物激活作用。月桂酸作为Cyp4b1的底物，能有效抑制小鼠肾和膀胱微粒体以及纯化的Cyp4b1对DCB的生物激活作用。为了评估CYP4B1对膀胱癌的作用，我们通过umu试验和免疫化学研究进行了进一步调查。包括大鼠CYP4B1在内的10种纯化的大鼠P450s用于对2-NA、DMAB和DCB的umu试验。CYP4B1对DMAB和DCB的活性最高。其他P450s的活性不到CYP4B1的20%。CYP4B1也能激活2-NA，但其活性约为对DMAB或DCB活性的10%。大鼠膀胱上皮细胞被抗Cyp4b1抗体特异性染色，表明大鼠膀胱黏膜中存在CYP4B1。此外，通过Northern印迹法和逆转录-聚合酶链反应（RT-PCR）检测到了CYP4B1 mRNA。这些发现表明，CYP4B1可能通过激活芳香胺在大鼠和小鼠膀胱致癌过程的起始阶段发挥作用。
