Assessment of 9-OH- and 7,8-diol-benzo[a]pyrene in Blood as Potent Markers of Cognitive Impairment Related to benzo[a]pyrene Exposure: An Animal Model Study.
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ABSTRACT:
SUBMITTER: Cherif LS
PROVIDER: S-EPMC7998639 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Benzo[a]pyrene 7,8-diol 9,10-epoxide adducts in DNA are implicated in mutagenesis, and their formation from the diol epoxides and subsequent incorrect replication by human DNA polymerases provide an attractive mechanism for the induction of cancer by this highly carcinogenic hydrocarbon and its diol epoxide metabolites. Here, we describe the crystal structure of such an adduct at the exocyclic amino group of a purine nucleoside. The present adduct derives from trans opening at C10 of the (-)-(7S,8R)-diol (9R,10S)-epoxide enantiomer by the exocyclic N(2)-amino group of deoxyguanosine. In the crystal, the pyrene rings of adjacent molecules stack with each other, but the guanine bases do not stack either intermolecularly with each other or intramolecularly with the pyrene. The most notable features of the molecular structure are (i) independent and unambiguous proof of the absolute configuration of the adduct based on the spatial relationship between the known chiral carbon atoms of the deoxyribose and the four asymmetric centers in the hydrocarbon moiety; (ii) visualization of the relative orientations of the pyrene and guanine ring systems as well as the conformation of the partially saturated hydrocarbon ring (comprising carbon atoms 7, 8, 9, and 10), both of which conformational features in the crystal are in good agreement with deductions from NMR and CD measurements in solution; and (iii) the presence in the crystal of a syn glycosidic torsion angle, a conformation that is unusual in B-DNA but that may be involved in error-prone replication of these benzo[a]pyrene 7,8-diol 9,10-epoxide deoxyguanosine adducts by DNA polymerases.
| S-EPMC341736 | biostudies-literature
Project description:Polycyclic aromatic hydrocarbons (PAH) are environmental and tobacco carcinogens. Human aldo-keto reductases catalyze the metabolic activation of proximate carcinogenic PAH trans-dihydrodiols to yield electrophilic and redox-active o-quinones. Benzo[a]pyrene-7,8-dione a representative PAH o-quinone is reduced back to the corresponding catechol to generate a futile redox-cycle. We investigated whether sulfonation of PAH catechols by human sulfotransferases (SULT) could intercept the catechol in human lung cells. RT-PCR identified SULT1A1, -1A3, and -1E1 as the isozymes expressed in four human lung cell lines. The corresponding recombinant SULTs were examined for their substrate specificity. Benzo[a]pyrene-7,8-dione was reduced to benzo[a]pyrene-7,8-catechol by dithiothreitol under anaerobic conditions and then further sulfonated by the SULTs in the presence of 3'-[(35)S]phosphoadenosine 5'-phosphosulfate as the sulfonate group donor. The human SULTs catalyzed the sulfonation of benzo[a]pyrene-7,8-catechol and generated two isomeric benzo[a]pyrene-7,8-catechol O-monosulfate products that were identified by reversed phase HPLC and by LC-MS/MS. The various SULT isoforms produced the two isomers in different proportions. Two-dimensional (1)H and (13)C NMR assigned the two regioisomers of benzo[a]pyrene-7,8-catechol monosulfate as 8-hydroxy-benzo[a]pyrene-7-O-sulfate (M1) and 7-hydroxy-benzo[a]pyrene-8-O-sulfate (M2), respectively. The kinetic profiles of three SULTs were different. SULT1A1 gave the highest catalytic efficiency (k(cat)/K(m)) and yielded a single isomeric product corresponding to M1. By contrast, SULT1E1 showed distinct substrate inhibition and formed both M1 and M2. Based on expression levels, catalytic efficiency, and the fact that the lung cells only produce M1, it is concluded that the major isoform that can intercept benzo[a]pyrene-7,8-catechol is SULT1A1.
| S-EPMC3436139 | biostudies-literature
Project description:Metabolic activation of the proximate carcinogen benzo[a]pyrene-7,8-trans-dihydrodiol (B[a]P-7,8-trans-dihydrodiol) by aldo-keto reductases (AKRs) leads to B[a]P-7,8-dione that is both electrophilic and redox-active. B[a]P-7,8-dione generates reactive oxygen species resulting in oxidative DNA damage in human lung cells. However, information on the formation of stable B[a]P-7,8-dione-DNA adducts in these cells is lacking. We studied stable DNA adduct formation of B[a]P-7,8-dione in human lung adenocarcinoma A549 cells, human bronchoalveolar H358 cells, and immortalized human bronchial epithelial HBEC-KT cells. After treatment with 2 μM B[a]P-7,8-dione, the cellular DNA was extracted from the cell pellets subjected to enzyme hydrolysis and subsequent analysis by LC-MS/MS. Several stable DNA adducts of B[a]P-7,8-dione were only detected in A549 and HBEC-KT cells. In A549 cells, the structures of stable B[a]P-7,8-dione-DNA adducts were identified as hydrated-B[a]P-7,8-dione-N(2)-2'-deoxyguanosine and hydrated-B[a]P-7,8-dione-N1-2'-deoxyguanosine. In HBEC-KT cells, the structures of stable B[a]P-7,8-dione-DNA adducts were identified as hydrated-B[a]P-7,8-dione-2'-deoxyadenosine, hydrated-B[a]P-7,8-dione-N1- or N3-2'-deoxyadenosine, and B[a]P-7,8-dione-N1- or N3-2'-deoxyadenosine. In each case, adduct structures were characterized by MS(n) spectra. Adduct structures were also compared to those synthesized from reactions of B[a]P-7,8-dione with either deoxyribonucleosides or salmon testis DNA in vitro but were found to be different.
| S-EPMC3660951 | biostudies-literature
Project description:Polycyclic aromatic hydrocarbons (PAHs) are environmental and tobacco carcinogens. Proximate carcinogenic PAH trans-dihydrodiols are activated by human aldo-keto reductases (AKRs) to yield electrophilic and redox-active o-quinones. Interconversion among benzo[a]pyrene (B[a]P)-7,8-dione, a representative PAH o-quinone, and its corresponding catechol generates a futile redox-cycle with the concomitant production of reactive oxygen species (ROS). We investigated whether glucuronidation of B[a]P-7,8-catechol by human UDP glucuronosyltransferases (UGTs) could intercept the catechol in three different human lung cells. RT-PCR showed that UGT1A1, 1A3, and 2B7 were only expressed in human lung adenocarcinoma A549 cells. The corresponding recombinant UGTs were examined for their kinetic constants and product profile using B[a]P-7,8-catechol as a substrate. B[a]P-7,8-dione was reduced to B[a]P-7,8-catechol by dithiothreitol under anaerobic conditions and then further glucuronidated by the UGTs in the presence of uridine-5'-diphosphoglucuronic acid as a glucuronic acid group donor. UGT1A1 catalyzed the glucuronidation of B[a]P-7,8-catechol and generated two isomeric O-monoglucuronsyl-B[a]P-7,8-catechol products that were identified by RP-HPLC and by LC-MS/MS. By contrast, UGT1A3 and 2B7 catalyzed the formation of only one monoglucuronide, which was identical to that formed in A549 cells. The kinetic profiles of three UGTs followed Michaelis-Menten kinetics. On the basis of the expression levels of UGT1A3 and UGT2B7 and the observation that a single monoglucuronide was produced in A549 cells, we suggest that the major UGT isoforms in A549 cells that can intercept B[a]P-7,8-catechol are UGT1A3 and 2B7.
| S-EPMC3829198 | biostudies-literature
Project description:Poly(ADP-ribosyl)ation (PARylation) is a complex and reversible posttranslational modification catalyzed by poly(ADP-ribose)polymerases (PARPs), which orchestrates protein function and subcellular localization. The function of PARP1 in genotoxic stress response upon induction of oxidative DNA lesions and strand breaks is firmly established, but its role in the response to chemical-induced, bulky DNA adducts is understood incompletely. To address the role of PARP1 in the response to bulky DNA adducts, we treated human cancer cells with benzo[a]pyrene 7,8-dihydrodiol-9,10-epoxide (BPDE), which represents the active metabolite of the environmental carcinogen benzo[a]pyrene [B(a)P], in nanomolar to low micromolar concentrations. Using a highly sensitive LC-MS/MS method, we revealed that BPDE induces cellular PAR formation in a time- and dose-dependent manner. Consistently, PARP1 activity significantly contributed to BPDE-induced genotoxic stress response. On one hand, PARP1 ablation rescued BPDE-induced NAD+ depletion and protected cells from BPDE-induced short-term toxicity. On the other hand, strong sensitization effects of PARP inhibition and PARP1 ablation were observed in long-term clonogenic survival assays. Furthermore, PARP1 ablation significantly affected BPDE-induced S- and G2-phase transitions. Together, these results point towards unresolved BPDE-DNA lesions triggering replicative stress. In line with this, BPDE exposure resulted in enhanced formation and persistence of DNA double-strand breaks in PARP1-deficient cells as evaluated by microscopic co-localization studies of 53BP1 and ?H2A.X foci. Consistently, an HPRT mutation assay revealed that PARP inhibition potentiated the mutagenicity of BPDE. In conclusion, this study demonstrates a profound role of PARylation in BPDE-induced genotoxic stress response with significant functional consequences and potential relevance with regard to B[a]P-induced cancer risks.
| S-EPMC5866831 | biostudies-other
Project description:Genotoxic agents cause cellular DNA damage and stress responses, including transcriptional changes. Here we focused on the early transcriptional responses of human cells to benzo(a)pyrene diol epoxide (BPDE), which causes bulky DNA adduct damage. Human amnion epithelial FL cells were exposed to three doses of BPDE (5, 50, and 500 nM) and the vehicle control DMSO, and differential gene expression profiles were obtained 4 h after exposure using oligonucleotide microarrays followed by validation with quantitative real-time RT-PCR. Compared with a few genes affected by the low and medium-dose exposure, extensive and robust changes in gene expression were induced by the high-dose BPDE. We found that the expression of cell cycle-regulators, signaling molecules and transcription factors were significantly altered and important signaling pathways related to cell survival or apoptosis were affected by BPDE. Several genes and related regulatory pathways that were previously not known to be responsive to this genotoxic agent have now been implicated, which helps to draw the whole picture of how cells respond to environmental chemical exposure via transcriptional regulation. Keywords: Dose response
2008-01-01 | GSE7675 | GEO
Project description:The catalytic intermediates of DNA topoisomerase I (top1) are cleavage complexes that can relax DNA supercoiling (intramolecular reaction) or mediate recombinations (intermolecular religation). We report here that DNA adducts formed from benzo[a]pyrene bay-region diol epoxides can markedly affect top1 activity. Four oligonucleotide 22-mers of the same sequence were synthesized, each of which contained a stereoisomerically unique benzo[a]pyrene 7, 8-diol 9,10-epoxide adduct at the 2-amino group of a central 2'-deoxyguanosine residue. These four adducts correspond to either cis or trans opening at C-10 of the (+)-(7R, 8S, 9S, 10R)- or (-)-(7S, 8R, 9R, 10S)-7,8-diol 9,10-epoxides. Their solution conformations in duplex DNA (intercalated and minor-groove bound for the cis and trans opened adducts respectively) can be deduced from previous NMR studies. All four adducts completely suppress top1 cleavage activity at the alkylation site and induce the formation of new top1cleavage complexes on both strands of the DNA 3-6 bases away from the alkylation site. The trans opened adduct from the highly carcinogenic (+)-diol epoxide is the most active in inducing top1 cleavage independently of camptothecin, demonstrating that minor groove alkylation can efficiently poison top1. We also found that this isomer of the diol epoxide induces the formation of top1-DNA complexes in mammalian cells, which suggests a possible relationship between induction of top1 cleavage complexes and carcinogenic activity of benzo[a]pyrene diol epoxides.
| S-EPMC15750 | biostudies-literature
Project description:Benzo[a]pyrene (B[a]P) is a polycyclic aromatic hydrocarbon (PAH) commonly found in cigarette smoke, automobile exhaust fumes, grilled meat, and smoked food among others. Exposure to B[a]P is associated with a range of toxic effects including developmental, neurological, oxidative, inflammatory, mutagenic, carcinogenic and mortal. Efficient and more affordable experimental models like Drosophila melanogaster could provide more insight into the mechanism of PAH toxicity and help develop new strategies for prevention, diagnosis and treatment of PAH-related conditions. In this study, we examined the induction of some biochemical changes along with mortality and functional senescence by B[a]P and its metabolite, benzo[a]pyrene- 7,8-dihydrodiol-910-epoxide (BPDE) in the Canton-S strain of Drosophila melanogaster, with the aim to establish an alternative assay medium for B[a]P toxicity in flies. Flies were exposed to 2-200 μM of B[a]P and 1-10 μM of BPDE through diet for a seven-day survival assay followed by a four-day treatment to determine the effects of the compounds on negative geotaxis, fecundity and some biochemical parameters of oxidative damage. BPDE significantly reduced the survival rate of flies along the 7 days of exposure whereas B[a]P did not cause any significant change in the survival rate of flies. B[a]P and BPDE significantly reduced the climbing ability of flies after 4 days of exposure. Rate of emergence of flies significantly reduced at 10-200 μM of B[a]P and 5-10 μM of BPDE. Both compounds caused various levels of alterations in the values of reduced glutathione (GSH), total thiol (T-SH), glutathione-S-transferase (GST), catalase (CAT), hydrogen peroxide (H2O2), nitric oxide (NO) and acetylcholinesterase (AChE) of the flies. The compounds also exhibited high binding affinities and molecular interactions with the active site amino acid residues of Drosophila GST and the inhibitor binding site of Drosophila AChE in an in silico molecular docking analysis, with BPDE forming stable hydrogen bonds with AChE. Hence, the Canton-S strain of Drosophila melanogaster could offer a simple and affordable assay medium to study B[a]P toxicity.
| S-EPMC7985712 | biostudies-literature
Project description:Genotoxic agents cause cellular DNA damage and stress responses, including transcriptional changes. Here we focused on the early transcriptional responses of human cells to benzo(a)pyrene diol epoxide (BPDE), which causes bulky DNA adduct damage. Human amnion epithelial FL cells were exposed to three doses of BPDE (5, 50, and 500 nM) and the vehicle control DMSO, and differential gene expression profiles were obtained 4 h after exposure using oligonucleotide microarrays followed by validation with quantitative real-time RT-PCR. Compared with a few genes affected by the low and medium-dose exposure, extensive and robust changes in gene expression were induced by the high-dose BPDE. We found that the expression of cell cycle-regulators, signaling molecules and transcription factors were significantly altered and important signaling pathways related to cell survival or apoptosis were affected by BPDE. Several genes and related regulatory pathways that were previously not known to be responsive to this genotoxic agent have now been implicated, which helps to draw the whole picture of how cells respond to environmental chemical exposure via transcriptional regulation. Experiment Overall Design: Human amnion epithelial FL cells were exposed to vehicle control (dimethyl sulfoxide) and increasing doses (5, 50, 500 nM) of anti-benzo(a)pyrene diol epoxide (anti-BPDE), respectively. The transcriptomes of the three treatments were compared to that of the control, respectively, to test the hypothesis that a characterized differential expression profile would be generated by exposure to various doses of this genotoxic agent.
2008-04-08 | E-GEOD-7675 | biostudies-arrayexpress
Project description:Tobacco smoke is an important risk factor for various human cancers, including esophageal cancer. How benzo [a]pyrene diol epoxide (BPDE), a carcinogen present in tobacco smoke as well as in environmental pollution, induces esophageal carcinogenesis has yet to be defined. In this study, we investigated the molecular mechanism responsible for BPDE-suppressed expression of retinoic acid receptor-beta2 (RAR-beta2) in esophageal cancer cells. We treated esophageal cancer cells with BPDE before performing methylation-specific polymerase chain reaction (MSP) to find that BPDE induced methylation of the RAR-beta2 gene promoter. We then performed chromatin immunoprecipitation (ChIP) assays to find that BPDE recruited genes of the methylation machinery into the RAR-beta2 gene promoter. We found that BPDE recruited DNA (cytosine-5-)-methyltransferase 3 alpha (DNMT3A), but not beta (DNMT3B), in a time-dependent manner to methylate the RAR-beta2 gene promoter, which we confirmed by reverse transcription-polymerase chain reaction (RT-PCR) analysis of the reduced RAR-beta2 expression in these BPDE-treated esophageal cancer cell lines. However, BPDE did not significantly change DNMT3A expression, but it slightly reduced DNMT3B expression. DNA methylase inhibitor 5-aza-2'-deoxycytidine (5-Aza) and DNMT3A small hairpin RNA (shRNA) vector antagonized the effects of BPDE on RAR-beta2 expressions. Transient transfection of the DNMT3A shRNA vector also antagonized BPDE's effects on expression of RAR-beta2, c-Jun, phosphorylated extracellular signal-regulated protein kinases 1/2 (ERK1/2), and cyclooxygenase-2 (COX-2), suggesting a possible therapeutic effect. The results of this study form the link between the esophageal cancer risk factor BPDE and the reduced RAR-beta2 expression.
| S-EPMC2867823 | biostudies-literature