Mauthe R J, Cook V M, Coffing S L, Baird W M
Department of Medicinal Chemistry, School of Pharmacy and Pharmacal Sciences, Purdue University, West Lafayette, IN 47907.
Carcinogenesis. 1995 Jan;16(1):133-7. doi: 10.1093/carcin/16.1.133.
Carcinogenic polycyclic aromatic hydrocarbons induce DNA damage through direct covalent interactions with nucleotides of the DNA in cells in which they are activated to 'ultimate carcinogenic metabolites'. To determine whether they also induce oxidative damage to DNA under the same circumstances, early passage Syrian hamster embryo and human mammary carcinoma cell line MCF-7 cultures were treated for 24 h with 0-5 micrograms/ml benzo[a]pyrene (BaP) or for 1 h with 0-100 microM methylene blue (as a positive control for oxidative damage). The cells were then exposed to fluorescent light for 1 or 4 h or retained in darkness. After cell harvest, DNA isolation and enzymatic digestion of the DNA to deoxyribonucleosides, the amounts of 8-hydroxy-2'deoxyguanosine (8-OH-dGuo) and unmodified deoxyguanosine present were determined by reverse-phase HPLC with electrochemical and UV detection respectively. Cultures treated with methylene blue for 1 h followed by light exposure for 1 h contained 5-fold (10 microM) and 8- to 28-fold (100 microM) higher 8-OH-dGuo levels than cells treated with methylene blue not exposed to light or untreated cells with methylene blue not exposed to light or untreated cells exposed to light. There was no significant change in 8-OH-dGuo levels in cultures treated with 1-5 micrograms/ml BaP for 24 h in the absence of light. However, both the human and hamster cell cultures treated with BaP and then exposed to fluorescent light for 4 h contained 3-fold (1 micrograms/ml) and 8- to 10-fold (5 micrograms/ml) higher 8-OH-dGuo levels than those not exposed to light or not treated with BaP. These results indicate that BaP treatment does not cause 8-OH-dGuo formation in DNA of cells maintained in darkness. Exposure of BaP-treated cells to fluorescent light causes formation of significant amounts of oxidative DNA damage as measured by 8-OH-dGuo formation. These findings suggest that oxidative damage of DNA could be involved in tumor induction by BaP in tissues, such as skin, in which exposure to BaP can occur in the presence of light.
致癌性多环芳烃通过与细胞中被激活为“最终致癌代谢物”的DNA核苷酸直接共价相互作用来诱导DNA损伤。为了确定它们在相同情况下是否也会诱导DNA的氧化损伤，用0 - 5微克/毫升苯并[a]芘（BaP）对传代早期的叙利亚仓鼠胚胎和人乳腺癌细胞系MCF - 7培养物处理24小时，或用0 - 100微摩尔/升亚甲蓝（作为氧化损伤的阳性对照）处理1小时。然后将细胞暴露于荧光下1或4小时，或置于黑暗中。细胞收获后，进行DNA分离并将DNA酶解为脱氧核糖核苷，分别通过电化学和紫外检测的反相高效液相色谱法测定8 - 羟基 - 2'-脱氧鸟苷（8 - OH - dGuo）和未修饰的脱氧鸟苷的含量。用亚甲蓝处理1小时后再暴露于光下1小时的培养物中，8 - OH - dGuo水平比未暴露于光的亚甲蓝处理细胞或未处理细胞（无论是否暴露于光）高5倍（10微摩尔）和8至28倍（100微摩尔）。在无光条件下，用1 - 5微克/毫升BaP处理24小时的培养物中8 - OH - dGuo水平没有显著变化。然而，用BaP处理后再暴露于荧光下4小时的人及仓鼠细胞培养物中，8 - OH - dGuo水平比未暴露于光或未用BaP处理的细胞高3倍（1微克/毫升）和8至10倍（5微克/毫升）。这些结果表明，在黑暗中培养的细胞中，BaP处理不会导致DNA中8 - OH - dGuo的形成。用荧光照射经BaP处理的细胞会导致形成大量的氧化性DNA损伤，这通过8 - OH - dGuo的形成来衡量。这些发现表明，DNA的氧化损伤可能参与BaP在诸如皮肤等组织中的肿瘤诱导过程，在这些组织中，BaP的暴露可能发生在有光的情况下。
