The effect of cadmium on benzo[a]pyrene-induced DNA damage and repair in Sprague-Dawley rats

Peng, Cheng. (2002). The effect of cadmium on benzo[a]pyrene-induced DNA damage and repair in Sprague-Dawley rats PhD Thesis, School of Molecular and Microbial Sciences, The University of Queensland.

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Author Peng, Cheng.
Thesis Title The effect of cadmium on benzo[a]pyrene-induced DNA damage and repair in Sprague-Dawley rats
School, Centre or Institute School of Molecular and Microbial Sciences
Institution The University of Queensland
Publication date 2002
Thesis type PhD Thesis
Supervisor Arungundrum S. Prakash
Charles T. Dameron,
Total pages 158
Collection year 2002
Language eng
Subjects L
259902 Environmental Chemistry (incl. Atmospheric Chemistry)
770304 Physical and chemical conditions
Formatted abstract
Benzo[a]pyrene (BP) and cadmium (Cd) are environmental pollutants and constituents of tobacco. Both have been classified in the same group as A2 carcinogen by lARC (1993) and associated with smoking-related lung cancer. BPinduced carcinogenesis is mediated through BP diol epoxide (BPDE), the reactive metabolite of BP formed by cytochrome P450 oxidation. The reaction of BPDE with DNA leading to the formation of DNA adducts is the first and key step in initiating its carcinogenesis. Cd, on the other hand, is believed to express its carcinogenic potential via oxidative damage to DNA. It may also act as a co-carcinogen through inhibition of DNA repair. In many cases, BP and Cd co-exist in the human environment including food and tobacco. To study the potential synergism between BP and Cd in carcinogenesis, we investigated the role of the Cd in BP-induced DNA damage in vitro and in vivo.

In cells BP is metabolised by P450 enzymes to BPDE with syn and anti diastereoisomeric forms that show differences in the DNA alkylating properties. Using modified sequencing and 32P-postlabeling techniques we investigated the effect of Cd on the alkylating properties of the syn and anti BPDE diastereomers in vitro. Preincubation studies showed that both diastereomers of BPDE are hydrolysed completely in triethanolamine buffer in less than 2 minutes. The depurination kinetics showed that a fi-action of the N7 alkylated guanine depurinated rapidly; however a significant amount of N2 guanine alkylation remained stable to spontaneous depurination over a 3-hour period. Similar results were obtained for the hydrolysis and alkylation rates of syn isomer but it required nearly 500 times more concentration to induce similar levels of N7 guanine alkylation. Cd strongly inhibited the N7 guanine alkylation of both isomers, but the minor groove alkylation was not affected as demonstrated by 32P-postlabelling assay, which confirmed the presence of heat-and cadmium-stable minor groove adducts in BPDE-treated calf thymus DNA.

To study the effect of Cd on DNA adduct formation induced by BP in vivo, we dosed Sprague-Dawley (S-D) rats with 100 μg of BP per mammary gland (3 pairs) with or without 20 μg per gland of CdCl2 in DMSO and measured DNA adduct levels in the mammary gland and liver over at 2, 5, 14 and 28 days post-treatment using 32postlabeling assay. Results showed that in mammary glands of rats dosed with BP alone the DNA adduct levels peaked between 5 and 14 days in the mammary gland and liver. In comparison, simultaneous administration of BP and Cd led to an overall reduction in adduct formation and a reduced rate of adduct repair. The inhibition of adduct repair was much stronger when CdCl2 was administrated 5 days after initial BP treatment. DNA adducts in liver showed a similar pattern but overall levels were about 10 % lower than in mammary gland. We also measured total P450 content and P4501A1 activities in the liver and found that BP-induced activation of P4501A1 was strongly inhibited by Cd. Based on these and other results we present a tentative mechanism of the combined effect of Cd on BP-induced carcinogenesis.
Keyword DNA.

Document type: Thesis
Collection: UQ Theses (RHD) - UQ staff and students only
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Created: Fri, 24 Aug 2007, 17:52:15 EST