The environmental pollutant benzo[a]pyrene (BaP) is a human carcinogen that reacts with DNA after metabolic activation catalysed by cytochromes P450 (CYP) 1A1 and 1B1 together with microsomal epoxide hydrolase. The azo dye Sudan I is a potent inducer of CYP1A1/2. Here, Wistar rats were either treated with single doses of BaP (150 mg/kg bw) or Sudan I (50 mg/kg bw) alone or with both compounds in combination to explore BaP-derived DNA adduct formation in vivo. Using 32P-postlabelling, DNA adducts generated by BaP-7,8-dihydrodiol-9,10-epoxide were found in livers of rats treated with BaP alone or co-exposed to Sudan I. During co-exposure to Sudan I prior to BaP treatment, BaP-DNA adduct levels increased 2.1-fold in comparison to BaP treatment alone. Similarly, hepatic microsomes isolated from rats exposed to Sudan I prior to BaP treatment were also the most effective in generating DNA adducts in vitro with the activated metabolites BaP-7,8-dihydrodiol or BaP-9-ol as intermediates. DNA adduct formation correlated with changes in the expression and/or enzyme activities of CYP1A1, 1A2 and 1B1 in hepatic microsomes. Thus, BaP genotoxicity in rats in vivo appears to be related to the enhanced expression and/or activity of hepatic CYP1A1/2 and 1B1 caused by exposure of rats to the studied compounds. Our results indicate that the industrially employed azo dye Sudan I potentiates the genotoxicity of the human carcinogen BaP, and exposure to both substances at the same time seems to be hazardous to humans.

• Klíčová slova
• DNA-adducts, Sudan I, benzo[a]pyrene, cytochromes P450 1A1 and 1A2 and 1B1, genotoxicity, microsomal epoxide hydrolase,
• MeSH
• adukty DNA toxicita   MeSH
• barvicí látky toxicita   MeSH
• benzopyren toxicita   MeSH
• cytochrom P-450 CYP1A1 metabolismus   MeSH
• jaterní mikrozomy účinky léků   MeSH
• játra účinky léků   MeSH
• karcinogeny životního prostředí toxicita   MeSH
• krysa rodu Rattus MeSH
• naftoly toxicita   MeSH
• potkani Wistar MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• 1-phenylazo-2-naphthol MeSH Prohlížeč
• adukty DNA MeSH
• barvicí látky MeSH
• benzo(a)pyrene-DNA adduct MeSH Prohlížeč
• benzopyren MeSH
• cytochrom P-450 CYP1A1 MeSH
• karcinogeny životního prostředí MeSH
• naftoly MeSH

Endocrine disruptors (EDs) are compounds that interfere with the balance of the endocrine system by mimicking or antagonising the effects of endogenous hormones, by altering the synthesis and metabolism of natural hormones, or by modifying hormone receptor levels. The synthetic estrogen 17α-ethinylestradiol (EE2) and the environmental carcinogen benzo[a]pyrene (BaP) are exogenous EDs whereas the estrogenic hormone 17β-estradiol is a natural endogenous ED. Although the biological effects of these individual EDs have partially been studied previously, their toxicity when acting in combination has not yet been investigated. Here we treated Wistar rats with BaP, EE2 and estradiol alone or in combination and studied the influence of EE2 and estradiol on: (i) the expression of cytochrome P450 (CYP) 1A1 and 1B1 in rat liver on the transcriptional and translational levels; (ii) the inducibility of these CYP enzymes by BaP in this rat organ; (iii) the formation of BaP-DNA adducts in rat liver in vivo; and (iv) the generation of BaP-induced DNA adducts after activation of BaP with hepatic microsomes of rats exposed to BaP, EE2 and estradiol and with recombinant rat CYP1A1 in vitro. BaP acted as a strong and moderate inducer of CYP1A1 and 1B1 in rat liver, respectively, whereas EE2 or estradiol alone had no effect on the expression of these enzymes. However, when EE2 was administered to rats together with BaP, it significantly decreased the potency of BaP to induce CYP1A1 and 1B1 gene expression. For EE2, but not estradiol, this also correlated with a reduction of BaP-induced CYP1A1 enzyme activity in rat hepatic microsomes. Further, while EE2 and estradiol did not form covalent adducts with DNA, they affected BaP-derived DNA adduct formations in vivo and in vitro. The observed decrease in BaP-DNA adduct levels in rat liver in vivo resulted from the inhibition of CYP1A1-mediated BaP bioactivation by EE2 and estradiol. Our results indicate that BaP genotoxicity mediated through its activation by CYP1A1 in rats in vivo is modulated by estradiol and its synthetic derivative EE2.

• Klíčová slova
• 17alpha-ethinylestradiol, Benzo[a]pyrene, Cytochrome P450, DNA-adducts, Endocrine disruptors, Estradiol,
• MeSH
• benzopyren toxicita   MeSH
• cytochrom P-450 CYP1A1 biosyntéza   genetika   MeSH
• endokrinní disruptory toxicita   MeSH
• estradiol toxicita   MeSH
• ethinylestradiol toxicita   MeSH
• jaterní mikrozomy účinky léků   enzymologie   MeSH
• krysa rodu Rattus MeSH
• potkani Wistar MeSH
• regulace genové exprese enzymů * účinky léků   MeSH
• synergismus léků MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• benzopyren MeSH
• cytochrom P-450 CYP1A1 MeSH
• endokrinní disruptory MeSH
• estradiol MeSH
• ethinylestradiol MeSH

We investigated the toxicity of benzo[a]pyrene (B[a]P), 1-nitropyrene (1-NP) and 3-nitrobenzanthrone (3-NBA) in A549 cells. Cells were treated for 4 h and 24 h with: B[a]P (0.1 and 1 μM), 1-NP (1 and 10 μM) and 3-NBA (0.5 and 5 μM). Bulky DNA adducts, lipid peroxidation, DNA and protein oxidation and mRNA expression of CYP1A1, CYP1B1, NQO1, POR, AKR1C2 and COX2 were analyzed. Bulky DNA adducts were induced after both treatment periods; the effect of 1-NP was weak. 3-NBA induced high levels of bulky DNA adducts even after 4-h treatment, suggesting rapid metabolic activation. Oxidative DNA damage was not affected. 1-NP caused protein oxidation and weak induction of lipid peroxidation after 4-h incubation. 3-NBA induced lipid peroxidation after 24-h treatment. Unlike B[a]P, induction of the aryl hydrocarbon receptor, measured as mRNA expression levels of CYP1A1 and CYP1B1, was low after treatment with polycyclic aromatic hydrocarbon (PAH) nitro-derivatives. All test compounds induced mRNA expression of NQO1, POR, and AKR1C2 after 24-h treatment. AKR1C2 expression indicates involvement of processes associated with reactive oxygen species generation. This was supported further by COX2 expression induced by 24-h treatment with 1-NP. In summary, 3-NBA was the most potent genotoxicant, whereas 1-NP exhibited the strongest oxidative properties.

• Klíčová slova
• 1-nitropyrene, 3-nitrobenzanthrone, benzo[a]pyrene, bulky DNA adducts, gene expression, oxidative damage,
• MeSH
• adukty DNA účinky léků   genetika   MeSH
• benz(a)anthraceny toxicita   MeSH
• benzopyren toxicita   MeSH
• buňky A549 MeSH
• cyklooxygenasa 2 genetika   MeSH
• cytochrom P-450 CYP1A1 genetika   MeSH
• cytochrom P450 CYP1B1 genetika   MeSH
• hydroxysteroiddehydrogenasy genetika   MeSH
• lidé MeSH
• NAD(P)H dehydrogenasa (chinon) genetika   MeSH
• pneumocyty účinky léků   metabolismus   MeSH
• poškození DNA účinky léků   genetika   MeSH
• pyreny toxicita   MeSH
• systém (enzymů) cytochromů P-450 genetika   MeSH
• výfukové emise vozidel toxicita   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• 1-nitropyrene MeSH Prohlížeč
• 3-nitrobenzanthrone MeSH Prohlížeč
• adukty DNA MeSH
• AKR1C2 protein, human MeSH Prohlížeč
• benz(a)anthraceny MeSH
• benzopyren MeSH
• cyklooxygenasa 2 MeSH
• CYP1A1 protein, human MeSH Prohlížeč
• CYP1B1 protein, human MeSH Prohlížeč
• cytochrom P-450 CYP1A1 MeSH
• cytochrom P450 CYP1B1 MeSH
• hydroxysteroiddehydrogenasy MeSH
• NAD(P)H dehydrogenasa (chinon) MeSH
• NQO1 protein, human MeSH Prohlížeč
• POR protein, human MeSH Prohlížeč
• PTGS2 protein, human MeSH Prohlížeč
• pyreny MeSH
• systém (enzymů) cytochromů P-450 MeSH
• výfukové emise vozidel MeSH

The tumour suppressor p53 is one of the most important cancer genes. Previous findings have shown that p53 expression can influence DNA adduct formation of the environmental carcinogen benzo[a]pyrene (BaP) in human cells, indicating a role for p53 in the cytochrome P450 (CYP) 1A1-mediated biotransformation of BaP in vitro. We investigated the potential role of p53 in xenobiotic metabolism in vivo by treating Trp53(+/+), Trp53(+/-) and Trp53(-/-) mice with BaP. BaP-DNA adduct levels, as measured by (32)P-postlabelling analysis, were significantly higher in liver and kidney of Trp53(-/-) mice than of Trp53(+/+) mice. Complementarily, significantly higher amounts of BaP metabolites were also formed ex vivo in hepatic microsomes from BaP-pretreated Trp53(-/-) mice. Bypass of the need for metabolic activation by treating mice with BaP-7,8-dihydrodiol-9,10-epoxide resulted in similar adduct levels in liver and kidney in all mouse lines, confirming that the influence of p53 is on the biotransformation of the parent compound. Higher BaP-DNA adduct levels in the livers of Trp53(-/-) mice correlated with higher CYP1A protein levels and increased CYP1A enzyme activity in these animals. Our study demonstrates a role for p53 in the metabolism of BaP in vivo, confirming previous in vitro results on a novel role for p53 in CYP1A1-mediated BaP metabolism. However, our results also suggest that the mechanisms involved in the altered expression and activity of the CYP1A1 enzyme by p53 in vitro and in vivo are different.

• Klíčová slova
• Benzo[a]pyrene, Carcinogen metabolism, Cytochrome P450, DNA adducts, Mouse models, Tumour suppressor p53,
• MeSH
• adukty DNA metabolismus   MeSH
• benzopyren metabolismus   farmakokinetika   MeSH
• cytochrom P-450 CYP1A1 metabolismus   MeSH
• jaterní mikrozomy účinky léků   metabolismus   MeSH
• karcinogeny životního prostředí metabolismus   farmakokinetika   MeSH
• ledviny účinky léků   metabolismus   MeSH
• metabolická aktivace MeSH
• metabolická inaktivace MeSH
• mutantní kmeny myší MeSH
• myši inbrední C57BL MeSH
• NAD(P)H dehydrogenasa (chinon) metabolismus   MeSH
• nádorový supresorový protein p53 genetika   metabolismus   MeSH
• poškození DNA účinky léků   genetika   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• adukty DNA MeSH
• benzo(a)pyrene-DNA adduct MeSH Prohlížeč
• benzopyren MeSH
• cytochrom P-450 CYP1A1 MeSH
• karcinogeny životního prostředí MeSH
• NAD(P)H dehydrogenasa (chinon) MeSH
• nádorový supresorový protein p53 MeSH
• Nqo1 protein, mouse MeSH Prohlížeč

UNLABELLED: This review summarizes the results found in studies investigating the enzymatic activation of two genotoxic nitro-aromatics, an environmental pollutant and carcinogen 3-nitrobenzanthrone (3-NBA) and a natural plant nephrotoxin and carcinogen aristolochic acid I (AAI), to reactive species forming covalent DNA adducts. Experimental and theoretical approaches determined the reasons why human NAD(P)H: quinone oxidoreductase (NQO1) and cytochromes P450 (CYP) 1A1 and 1A2 have the potential to reductively activate both nitro-aromatics. The results also contributed to the elucidation of the molecular mechanisms of these reactions. The contribution of conjugation enzymes such as N,O-acetyltransferases (NATs) and sulfotransferases (SULTs) to the activation of 3-NBA and AAI was also examined. The results indicated differences in the abilities of 3-NBA and AAI metabolites to be further activated by these conjugation enzymes. The formation of DNA adducts generated by both carcinogens during their reductive activation by the NOQ1 and CYP1A1/2 enzymes was investigated with pure enzymes, enzymes present in subcellular cytosolic and microsomal fractions, selective inhibitors, and animal models (including knock-out and humanized animals). For the theoretical approaches, flexible in silico docking methods as well as ab initio calculations were employed. The results summarized in this review demonstrate that a combination of experimental and theoretical approaches is a useful tool to study the enzyme-mediated reaction mechanisms of 3-NBA and AAI reduction.

• MeSH
• acetyltransferasy metabolismus   MeSH
• adukty DNA chemie   metabolismus   MeSH
• aromatické hydroxylasy metabolismus   MeSH
• benz(a)anthraceny chemie   metabolismus   MeSH
• biokatalýza MeSH
• enzymy metabolismus   MeSH
• kyseliny aristolochové chemie   metabolismus   MeSH
• lidé MeSH
• molekulární modely * MeSH
• NAD(P)H dehydrogenasa (chinon) metabolismus   MeSH
• sulfotransferasy metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• 3-nitrobenzanthrone MeSH Prohlížeč
• acetyltransferasy MeSH
• adukty DNA MeSH
• aristolochic acid I MeSH Prohlížeč
• aromatické hydroxylasy MeSH
• benz(a)anthraceny MeSH
• enzymy MeSH
• kyseliny aristolochové MeSH
• NAD(P)H dehydrogenasa (chinon) MeSH
• sulfotransferasy MeSH

3-Aminobenzanthrone (3-ABA) is a human metabolite of carcinogenic 3-nitrobenzanthrone (3-NBA), which occurs in diesel exhaust and air pollution. Understanding which cytochrome P450 (CYP) enzymes are involved in metabolic activation and/or detoxication of this toxicant is important in the assessment of an individual's susceptibility to this substance. The aim of this study was to evaluate the efficiency of rat hepatic CYPs to oxidize 3-ABA and to examine the metabolites formed during such an oxidation. The metabolites formed by CYPs in rat hepatic microsomes were separated by high performance liquid chromatography (HPLC). 3-ABA is oxidized by these enzymes to three metabolites, which were separated by HPLC as distinguish product peaks. Using co-chromatography with synthetic standards, two of them were identified to be oxidative metabolites of 3-ABA, N-hydroxy-3-ABA and 3-NBA. The structure of another 3-ABA metabolite remains to be characterized. To define the role of rat hepatic CYP enzymes in metabolism of 3-ABA, we investigated the modulation of its oxidation using different inducers of CYPs for treatment of rats to enrich the liver microsomes with individual CYPs. Based on these studies, we attribute most of 3-ABA oxidation in rat hepatic microsomes to CYP2B, followed by CYP1A, although a role of other hepatic CYPs cannot be ruled out. Inhibition of 3-ABA oxidation by selective inhibitors of individual CYPs, supported this finding.

• Klíčová slova
• 3-aminobenzanthrone, 3-nitrobenzanthrone, HPLC, cytochrome P450, induction, inhibition, oxidation,
• Publikační typ
• časopisecké články MeSH