Mutagenesis and antimutagenesis in Big Blue ® lacI transgenic rats




Yang, Haiyan

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The initiation of the cancer process is associated with mutations. Analysis of environmental exposure to chemical or physical agents causing these genetic alterations is of great importance in order to develop strategies for avoiding or reducing cancer risk in humans. The causality between mutagenesis and carcinogenesis also prompts the concept that the modifying effect on mutagenesis by a compound would be predictive of the cancer preventive potential of that compound. The Big Blue© transgenic assay, using the E. coli lacI gene as the mutational target provides an opportunity to evaluate mutagenesis and its modulation m vivo. This model system was used to study the tissue-specific effect of the potential chemopreventive agent conjugated linoleic acid (CLA), on the mutagenicity of the suspected human carcinogen, 2-amino-1 -methyl-6- phenylimidazo[4,5-b]pyridine (PhIP). PhIP and CLA were selected for study since both compounds are consumed by humans on a daily basis, and are suspected to be related to the human risk of colon, breast, and prostate cancers. The mutagenicity of PhIP in Big Blue© rats was shown to be tissue-, sex-, and dose-dependent. PhIP was found to be a potent mutagen in the colon, followed by the cecum, prostate, and kidney. Compared with the background mutational spectra, the PhlP-induced spectra were characterized by an elevated proportion o f-1 frameshifts, consisting mainly of deletions of single G:C base pair. However, the induced spectra varied among tissues. A sex-dependent induction of mutation by PhIP was observed in the kidney such that the PhlP-induced mutation frequency was twice as high in male rats as in female rats; the biological significance of this difference is not clear. In contrast, although PhIP has been shown to induce colon tumors preferentially in male rats, and only rarely in female rats, no difference in mutational response was detected between the colons of male and female rats treated with PhIP. Experiments were performed to examine the in vivo effect of CLA on mutagenesis. Similar to what is seen for the mutagenicity of PhIP, the modification by CLA depends on tissue, sex, and dose of administration. CLA showed a modest protection against PhlP-induced mutagenesis in the distal part of the colon, in the prostate, and in the kidney of female rats. However, significant changes in the overall PhlP-induced mutation spectrum were seen only in the prostate. The antimutagenic effect of CLA may be directly responsible for its cancer prevention «^lability, since PhlP-induced aberrant crypt foci in the colon of male rats were completely inhibited by CLA However, CLA was not totally innocuous. When supplemented at 0.5%, CLA acted as a comutagen of PhIP, increasing the PhlP-induced MF in the cecum, although this effect was not observed when CLA was supplemented at 1%. The differences in effect may be related to the antioxidant or pro-oxidant activities of CLA isomers under experimental conditions. Due to the artificial nature of the lambda/LIZ lacI transgene and the possible absence of DNA repair in this transgene, the suitability of the Big Blue© transgenic assay as a mutational test system has been questioned. We examined the repair of UV- and benzo(α)pyrene diol epoxide-induced DNA damage in this non-transcribed lambda construct of the Big Blue© rat-2 transgenic cell line and demonstrated that DNA damage is indeed repaired in this transgenic construct. Lastly, since CLA altered the mutational spectra in the prostate in a way consistent with an effect of mismatch repair, the possibility of an effect of CLA on mismatch repair was explored in bacteria. Although CLA was found to increase mutant frequency in a mismatch repair proficient E. coli strain, but not in deficient strains, the mechanism by which CLA operates remains unclear. Altogether, the data demonstrate the mutagenicity of PhIP and its modulation by CLA as a function of tissue, sex, and dose of administration, and support the application of the Big Blue© transgenic assay as a screening tool for mutagens and chemopreventive agent



Mutagenesis, Cancer, Genetic aspects