Mutation Research-genetic Toxicology and Environmental Mutagenesis, 2002
Benzo[a]pyrene (B[a]P) is the most thoroughly studied polycyclic aromatic hydrocarbon (PAH). Many... more Benzo[a]pyrene (B[a]P) is the most thoroughly studied polycyclic aromatic hydrocarbon (PAH). Many mechanisms have been suggested to explain its carcinogenic activity, yet many questions still remain. K-region dihydrodiols of PAHs are metabolic intermediates depending on the specific cytochrome P450 and had been thought to be detoxification products. However, K-region dihydrodiols of several PAHs have recently been shown to morphologically transform mouse embryo C3H10T1/2CL8 cells (C3H10T1/2 cells). Because K-region dihydrodiols are not metabolically formed from PAHs by C3H10T1/2 cells, these cells provide a useful tool to independently study the mechanisms of action of PAHs and their K-region dihydrodiols. Here, we compare the morphological cell transforming, DNA damaging, and DNA adducting activities of the K-region dihydrodiol of B[a]P, trans-B[a]P-4,5-diol with B[a]P. Both trans-B[a]P-4,5-diol and B[a]P morphologically transformed C3H10T1/2 cells by producing both Types II and III transformed foci. The morphological cell transforming and cytotoxicity dose response curves for trans-B[a]P-4,5-diol and B[a]P were indistinguishable. Since morphological cell transformation is strongly associated with mutation and/or larger scale DNA damage in C3H10T1/2 cells, the identification of DNA damage induced in these cells by trans-B[a]P-4,5-diol was sought. Both trans-B[a]P-4,5-diol and B[a]P exhibited significant DNA damaging activity without significant concurrent cytotoxicity using the comet assay, but with different dose responses and comet tail distributions. DNA adduct patterns from C3H10T1/2 cells were examined after trans-B[a]P-4,5-diol or B[a]P treatment using 32 P-postlabeling techniques and improved TLC elution systems designed to separate polar DNA adducts. While B[a]P treatment produced one major DNA adduct identified as anti-trans-B[a]P-7,8-diol-9,10-epoxide-deoxyguanosine, no stable covalent DNA adducts were detected in the DNA of trans-B[a]P-4,5-diol-treated cells. In summary, this study provides Abbreviations: ara-C, cytosine arabinoside; B[a]P, benzo[a]pyrene; BNF, -naphthoflavone; C3H10T1/2, C3H10T1/2CL8; trans-B[a]P-4,5-diol, (±) trans-4,5-dihydro-4,5-dihydroxyB[a]P; BPDE, anti-trans-B[a]P-7,8-diol-9,10-epoxide; MMS, methyl methanesulfonate; PAH, polycyclic aromatic hydrocarbon; ROS, reactive oxygen species ଝ The research described in this article has been reviewed by the National Health and Environmental Effects Research Laboratory, U.S. Environmental Protection Agency and approved for publication. Approval does not signify that the contents necessarily reflect the views of the Agency nor does mention of trade names or commercial products constitute endorsement or recommendation for use. (S. Nesnow). 1383-5718/02/$ -see front matter. Published by Elsevier Science B.V. PII: S 1 3 8 3 -5 7 1 8 ( 0 2 ) 0 0 2 1 8 -8
Mutation Research-genetic Toxicology and Environmental Mutagenesis, 2002
Benzo[a]pyrene (B[a]P) is the most thoroughly studied polycyclic aromatic hydrocarbon (PAH). Many... more Benzo[a]pyrene (B[a]P) is the most thoroughly studied polycyclic aromatic hydrocarbon (PAH). Many mechanisms have been suggested to explain its carcinogenic activity, yet many questions still remain. K-region dihydrodiols of PAHs are metabolic intermediates depending on the specific cytochrome P450 and had been thought to be detoxification products. However, K-region dihydrodiols of several PAHs have recently been shown to morphologically transform mouse embryo C3H10T1/2CL8 cells (C3H10T1/2 cells). Because K-region dihydrodiols are not metabolically formed from PAHs by C3H10T1/2 cells, these cells provide a useful tool to independently study the mechanisms of action of PAHs and their K-region dihydrodiols. Here, we compare the morphological cell transforming, DNA damaging, and DNA adducting activities of the K-region dihydrodiol of B[a]P, trans-B[a]P-4,5-diol with B[a]P. Both trans-B[a]P-4,5-diol and B[a]P morphologically transformed C3H10T1/2 cells by producing both Types II and III transformed foci. The morphological cell transforming and cytotoxicity dose response curves for trans-B[a]P-4,5-diol and B[a]P were indistinguishable. Since morphological cell transformation is strongly associated with mutation and/or larger scale DNA damage in C3H10T1/2 cells, the identification of DNA damage induced in these cells by trans-B[a]P-4,5-diol was sought. Both trans-B[a]P-4,5-diol and B[a]P exhibited significant DNA damaging activity without significant concurrent cytotoxicity using the comet assay, but with different dose responses and comet tail distributions. DNA adduct patterns from C3H10T1/2 cells were examined after trans-B[a]P-4,5-diol or B[a]P treatment using 32 P-postlabeling techniques and improved TLC elution systems designed to separate polar DNA adducts. While B[a]P treatment produced one major DNA adduct identified as anti-trans-B[a]P-7,8-diol-9,10-epoxide-deoxyguanosine, no stable covalent DNA adducts were detected in the DNA of trans-B[a]P-4,5-diol-treated cells. In summary, this study provides Abbreviations: ara-C, cytosine arabinoside; B[a]P, benzo[a]pyrene; BNF, -naphthoflavone; C3H10T1/2, C3H10T1/2CL8; trans-B[a]P-4,5-diol, (±) trans-4,5-dihydro-4,5-dihydroxyB[a]P; BPDE, anti-trans-B[a]P-7,8-diol-9,10-epoxide; MMS, methyl methanesulfonate; PAH, polycyclic aromatic hydrocarbon; ROS, reactive oxygen species ଝ The research described in this article has been reviewed by the National Health and Environmental Effects Research Laboratory, U.S. Environmental Protection Agency and approved for publication. Approval does not signify that the contents necessarily reflect the views of the Agency nor does mention of trade names or commercial products constitute endorsement or recommendation for use. (S. Nesnow). 1383-5718/02/$ -see front matter. Published by Elsevier Science B.V. PII: S 1 3 8 3 -5 7 1 8 ( 0 2 ) 0 0 2 1 8 -8
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