Oxidative DNA damage induced by carcinogenic dinitropyrenes in the presence of p450 reductase
Oxidative DNA damage induced by carcinogenic dinitropyrenes in the presence of p450 reductase
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DOI：
10.1021/tx0497550
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发表时间：
2004-12-01
影响因子：
4.1
通讯作者：
Kawanishi, S
中科院分区：
文献类型：
--
作者：
Murata, M;Ohnishi, S;Kawanishi, S
硝基芘广泛存在于环境中，主要是由于柴油发动机的排放。二硝基芘（DNP），特别是1，8-DNP和1，6-DNP，是比其它硝基芘更强的诱变剂。1，8-DNP和1，6-DNP的致癌性强于1，3-二硝基芘（1，3-DNP）。认为代谢活化后加合物的形成在硝基芘致癌性的表达中起重要作用。然而，Djuric等人[（1993）Cancer Lett.]报道，在用1，6-DNP处理的大鼠中也发现了氧化DNA损伤以及加合物形成。我们使用P-32- 5 '-末端标记的DNA研究了在NAD（P）H-细胞色素P450还原酶存在下DNPs对DNA的氧化损伤。P450还原酶处理后，在NAD（P）H存在下，DNPs诱导Cu（II）介导的DNA损伤。1，8-DNP和1，6-DNP对DNA的损伤强度均大于1，3-DNP。我们还检测了合成的1-硝基-8-亚硝基芘（1，8-NNOP）和1-硝基-6-亚硝基芘（1，6-NNOP）分别作为1，8-DNP和1，6-DNP的代谢产物之一，发现1，8-NNOP和1，6-NNOP在NAD（P）H存在下诱导Cu（II）介导的DNA损伤，而未处理的DNP则没有。在P450还原酶处理的DNP和NNOP的两种情况下，过氧化氢酶和Cu（I）特异性螯合剂减弱了DNA损伤，表明H2 O2和Cu（I）参与。使用Clarke氧电极，通过NNOP与NAD（P）H和Cu（II）的反应测量氧消耗，发现NNOP被NAD（P）H非酶还原，并且Cu（II）的加入促进了氧化还原循环。因此，这些结果表明，DNPs通过硝基自由基阴离子酶促还原为NNOP，NNOP通过NAD（P）H进一步非酶促还原。随后，发生硝基自由基阴离子的自氧化和NNOP的还原形式，导致O-2（-）产生和DNA损伤。我们的结论是，氧化DNA损伤，除了DNA加合物的形成可能发挥重要作用的致癌DNPs通过其代谢产物。
Nitropyrenes are widespread in the environment due to mainly diesel engine emissions. Dinitropyrenes (DNPs), especially 1,8-dinitropyrene (1,8-DNP) and 1,6-dinitropyrene (1,6-DNP), are much more potent mutagens than other nitropyrenes. The carcinogenicity of 1,8-DNP and 1,6-DNP is stronger than 1,3-dinitropyrene (1,3-DNP). It is considered that adduct formation after metabolic activation plays an important role in the expression of carcinogenicity of nitropyrenes. However, Djuric et al. [(1993) Cancer Lett.] reported that oxidative DNA damage was also found as well as adduct formation in rats treated with 1,6-DNP. We investigated oxidative DNA damage by DNPs in the presence of NAD(P)H-cytochrome P450 reductase using P-32-5'-end-labeled DNA. After P450 reductase treatment, DNPs induced Cu(II)-mediated DNA damage in the presence of NAD(P)H. The intensity of DNA damage by 1,8-DNP or 1,6-DNP was stronger than 1,3-DNP. We also examined synthetic 1-nitro-8-nitrosopyrene (1,8-NNOP) and 1-nitro-6-nitrosopyrene (1,6-NNOP) as one of the metabolites of 1,8-DNP and 1,6-DNP, respectively, to find that 1,8-NNOP and 1,6-NNOP induced Cu(II)-mediated DNA damage in the presence of NAD(P)H but untreated DNPs did not. In both cases of P450 reductase-treated DNPs and NNOPs, catalase and a Cu(I) specific chelator attenuated DNA damage, indicating the involvement of H2O2 and Cu(I). Using a Clarke oxygen electrode, oxygen consumption by the reaction of NNOPs with NAD(P)H and Cu(II) was measured to find that NNOP was nonenzymatically reduced by NAD(P)H and that the addition of Cu(II) promoted the redox cycle. Therefore, these results suggest that DNPs are enzymatically reduced to NNOPs via nitro radical anion and that NNOPs are further reduced nonenzymatically by NAD(P)H. Subsequently, autoxidation of nitro radical anion and the reduced form of NNOP occurs, resulting in O-2(-) generation and DNA damage. We conclude that oxidative DNA damage in addition to DNA adduct formation may play important roles in the carcinogenesis of DNPs via their metabolites.