Sulforaphane and quercetin modulate PhIP-DNA adduct formation in human HepG2 cells and hepatocytes
Sulforaphane and quercetin modulate PhIP-DNA adduct formation in human HepG2 cells and hepatocytes
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DOI：
10.1093/carcin/bgg157
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发表时间：
2003-12-01
期刊：
影响因子：
4.7
通讯作者：
Bao, YP
中科院分区：
文献类型：
--
作者：
Bacon, JR;Williamson, G;Bao, YP
用加速器质谱法（AMS）检测了C-14标记的2-氨基-1-甲基-6-苯基咪唑[4，5-B]吡啶（PhIP）在人HepG 2细胞和人肝细胞中DNA加合物的形成。PhIP在100 pM和20 μ M之间以线性剂量依赖性方式产生DNA加合物。与膳食异硫氰酸酯，萝卜硫素（SFN，1-10 μ M），或类黄酮，槲皮素（5-20 μ M）的共同治疗，显着降低PhIP-DNA加合物的水平，在剂量依赖性的方式。保护程度取决于PhIP浓度，即在100 pM PhIP暴露后，SFN或槲皮素将加合物水平降低至检测限以下（0.15阿莫尔PhIP/μ g DNA），但在更高的PhIP暴露（10 nM和1 μ M）下，保护分别为60%和10%。使用实时RT-PCR和酶活性测定研究了I相，II相和DNA修复酶在这种保护中对PhIP-DNA加合物形成的参与。在完整的HepG 2细胞中，槲皮素抑制细胞色素P450（P450）1A 2，负责PhIP生物活化的主要I相酶。相反，SFN诱导II相解毒酶，UDP-葡萄糖醛酸转移酶1A 1和谷胱甘肽S-转移酶A1 mRNA的表达。SFN和槲皮素对DNA修复没有影响，无论是在PhIP-DNA加合物的水平方面，当细胞在致癌物暴露后用植物化学物质处理时，也没有调节两种DNA修复酶，脱嘌呤核酸内切酶和DNA聚合酶β的mRNA表达。这项研究表明，膳食异硫氰酸酯和类黄酮调节I相和II相酶的表达，从而提高了人类HepG 2细胞中膳食致癌物PhIP的解毒率，但不影响PhIP-DNA加合物修复率。人肝细胞中PhIP-DNA加合物的形成也与PhIP浓度呈剂量依赖性，10 nM PhIP处理后SFN或槲皮素的保护水平高达60%，但显示出较大的个体间差异，在某些个体中未观察到保护作用。
The formation of DNA adducts in human HepG2 cells and human hepatocytes exposed to C-14-labelled 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) was examined using Accelerator Mass Spectrometry (AMS). PhIP generated DNA adducts in a linear dose-dependent manner between 100 pM and 20 muM. Co-treatment with the dietary isothiocyanate, sulforaphane (SFN, 1-10 muM), or the flavonoid, quercetin (5-20 muM), significantly reduced the level of PhIP-DNA adducts in a dose-dependent manner. The degree of protection was dependent on PhIP concentration, i.e. after 100 pM PhIP exposure, SFN or quercetin reduced adduct levels to below the limit of detection (0.15 amol PhIP/mug DNA) but at higher PhIP exposure (10 nM and 1 muM), the protection was 60 and 10%, respectively. The involvement of phase I, phase II and DNA repair enzymes in this protection against PhIP-DNA adduct formation was investigated using real-time RT-PCR and enzyme activity assays. In intact HepG2 cells, quercetin inhibited cytochrome P450 (CYP)1A2, the main phase I enzyme responsible for PhIP bioactivation. In contrast, SFN induced phase II detoxification enzymes, UDP-glucuronosyltransferase 1A1 and glutathione S-transferase A1 mRNA expression. SFN and quercetin showed no effect on DNA repair, neither in terms of the level of PhIP-DNA adducts, when cells were treated with phytochemicals after the carcinogen exposure, nor the regulation of mRNA expression of two DNA repair enzymes, apurinic endonuclease and DNA polymerase beta. This study indicates that dietary isothiocyanates and flavonoids modulate phase I and phase II enzyme expression, hence increasing the rate of detoxification of the dietary carcinogen PhIP in human HepG2 cells but do not affect the rate of PhIP-DNA adduct repair. The formation of PhIP-DNA adducts in human hepatocytes was also dose-dependent with PhIP-concentration and the levels of protection by SFN or quercetin were up to 60% after 10 nM PhIP treatment, but showed large inter-individual variation with no observed protection in some individuals.