Oikawa S, Nishino K, Oikawa S, Inoue S, Mizutani T, Kawanishi S
Department of Hygiene, Mie University School of Medicine, Japan.
Biochem Pharmacol. 1998 Aug 1;56(3):361-70. doi: 10.1016/s0006-2952(98)00037-9.
DNA damage by metabolites of a food additive, butylated hydroxytoluene (BHT), was investigated as a potential mechanism of carcinogenicity. The mechanism of DNA damage by 2,6-di-tert-butyl-p-benzoquinone (BHT-quinone), 2,6-di-tert-butyl-4-hydroperoxyl-4-methyl-2,5-cyclohexadienone (BHT-OOH), and 3,5-di-tert-butyl-4-hydroxybenzaldehyde (BHT-CHO) in the presence of metal ions was investigated by using 32P-labeled DNA fragments obtained from the c-Ha-ras-1 proto-oncogene and the p53 tumor suppressor gene. BHT-OOH caused DNA damage in the presence of Cu(II), whereas BHT-quinone and BHT-CHO did not. However, BHT-quinone did induce DNA damage in the presence of NADH and Cu(II). Bathocuproine inhibited Cu(II)-mediated DNA damage, indicating the participation of Cu(I) in the process. Catalase also inhibited DNA damage induced by BHT-quinone, but not that induced by BHT-OOH. The DNA cleavage pattern observed with BHT-quinone plus NADH was different from that seen with BHT-OOH. With BHT-quinone plus NADH, piperidine-labile sites could be generated at nucleotides other than adenine residue. BHT-OOH caused cleavage specifically at guanine residues. Pulsed field gel electrophoresis showed that BHT-OOH and BHT-quinone induced DNA strand breaks in cultured cells, whereas BHT-CHO did not. Both BHT-quinone and BHT-OOH induced internucleosomal DNA fragmentation, which is the characteristic of apoptosis. Furthermore, flow cytometry analysis revealed an increase of peroxides in cultured cells treated with BHT-OOH or BHT-quinone. These results suggest that BHT-OOH participates in oxidative DNA damage directly, whereas BHT-quinone causes DNA damage through H2O2 generation, which leads to internucleosomal DNA fragmentation.
人们研究了食品添加剂丁基羟基甲苯（BHT）的代谢产物对DNA的损伤，将其作为一种潜在的致癌机制。通过使用从c-Ha-ras-1原癌基因和p53肿瘤抑制基因获得的32P标记的DNA片段，研究了在金属离子存在下2,6-二叔丁基对苯醌（BHT-醌）、2,6-二叔丁基-4-氢过氧基-4-甲基-2,5-环己二烯酮（BHT-OOH）和3,5-二叔丁基-4-羟基苯甲醛（BHT-CHO）对DNA的损伤机制。BHT-OOH在Cu(II)存在下会导致DNA损伤，而BHT-醌和BHT-CHO则不会。然而，BHT-醌在NADH和Cu(II)存在下确实会诱导DNA损伤。邻二氮菲抑制了Cu(II)介导的DNA损伤，表明Cu(I)参与了这一过程。过氧化氢酶也抑制了BHT-醌诱导的DNA损伤，但没有抑制BHT-OOH诱导的损伤。BHT-醌加NADH观察到的DNA切割模式与BHT-OOH不同。对于BHT-醌加NADH，除腺嘌呤残基外的其他核苷酸处可产生哌啶不稳定位点。BHT-OOH特异性地在鸟嘌呤残基处导致切割。脉冲场凝胶电泳显示，BHT-OOH和BHT-醌在培养细胞中诱导DNA链断裂，而BHT-CHO则不会。BHT-醌和BHT-OOH均诱导核小体间DNA片段化，这是细胞凋亡的特征。此外，流式细胞术分析显示，用BHT-OOH或BHT-醌处理的培养细胞中过氧化物增加。这些结果表明，BHT-OOH直接参与氧化性DNA损伤，而BHT-醌通过产生H2O2导致DNA损伤，进而导致核小体间DNA片段化。
