School of Environmental and Chemical Engineering, Shanghai University, Shanghai, People's Republic of China.
Mutat Res. 2012 Sep 18;747(2):240-5. doi: 10.1016/j.mrgentox.2012.06.001. Epub 2012 Jun 12.
1,3-Butadiene (BD) is a carcinogenic air pollutant. Its bioactivation produces four major metabolites, i.e., 3,4-epoxy-1-butene (EB), 3,4-epoxy-1,2-butanediol (EBD), 1,2,3,4-diepoxybutane (DEB), and 3-butene-1,2-diol (BDD). Studies have been mostly focused on DEB due to its strong mutagenicity/carcinogenicity. In contrast, studies of genotoxicity of EB, EBD, and BDD have been limited. In particular, genotoxicity of EBD and BDD using strand breaks as the endpoint has not been investigated. To obtain a more complete understanding of BD toxicity, in the present study, we used comet assay to investigate DNA damage induced by EB, EBD, and BDD in human hepatocyte L02 cells, with the aim to determine their relative potencies, the types of DNA damage, and the possible pathway to form strand breaks. Using alkaline comet assay (pH>13), it was observed that EB and EBD caused similar concentration-dependent increases in DNA migration from 50 to 1000μM. However, BDD induced a statistically significant increase only at 1000μM, and the increase itself was very small. EBD was as potent as EB at lower concentrations (≤200μM), and was slightly less potent than EB at higher concentrations. The results indicated that these metabolites could generate strand breaks in cells with the rank order of the potencies being EB>≈EBD≫BDD. All three compounds failed to cause statistically significant increases in DNA migration in pre-lysed cells, suggesting that they did not produce strand breaks through chemical pathways under our experimental conditions. By using comet assays at pH 11.9 and pH 9, it was demonstrated that EB and EBD generated both single-strand breaks (SSB) and alkali-labile sites, but BDD produced only SSB. To our knowledge, this is the first report to investigate EBD- and BDD-induced strand breaks in cells. The results implied that EBD could play an important role in toxicity of BD.
1,3-丁二烯（BD）是一种致癌的空气污染物。其生物活化产生四种主要代谢物，即 3,4-环氧-1-丁烯（EB）、3,4-环氧-1,2-丁二醇（EBD）、1,2,3,4-二环氧丁烷（DEB）和 3-丁烯-1,2-二醇（BDD）。由于其强烈的致突变性/致癌性，研究主要集中在 DEB 上。相比之下，对 EB、EBD 和 BDD 的遗传毒性研究有限。特别是，使用链断裂作为终点的 EBD 和 BDD 的遗传毒性尚未得到研究。为了更全面地了解 BD 的毒性，在本研究中，我们使用彗星试验研究了 EB、EBD 和 BDD 在人肝细胞 L02 细胞中诱导的 DNA 损伤，旨在确定它们的相对效力、DNA 损伤的类型以及形成链断裂的可能途径。使用碱性彗星试验（pH>13）观察到，EB 和 EBD 引起了类似的浓度依赖性 DNA 迁移增加，从 50 到 1000μM。然而，BDD 仅在 1000μM 时引起统计学上显著的增加，并且增加本身非常小。EBD 在较低浓度（≤200μM）时与 EB 一样有效，在较高浓度时略低于 EB。结果表明，这些代谢物可以在细胞中产生链断裂，效力顺序为 EB>≈EBD≫BDD。在预裂解细胞中，所有三种化合物都未能引起 DNA 迁移的统计学显著增加，这表明它们在我们的实验条件下没有通过化学途径产生链断裂。通过在 pH 11.9 和 pH 9 下使用彗星试验，证明 EB 和 EBD 均产生单链断裂（SSB）和碱不稳定部位，但 BDD 仅产生 SSB。据我们所知，这是首次研究细胞中 EBD 和 BDD 诱导的链断裂。结果表明，EBD 在 BD 的毒性中可能发挥重要作用。
