title: The genotoxic effect of carcinogenic PAHs, their artificial and environmental mixtures (EOM) on human diploid lung fibroblasts.
abstract: The aim of this study was to investigate the genotoxic effect of single carcinogenic polycyclic aromatic hydrocarbons (c-PAHs), as well as their binary and artificial mixtures containing c-PAHs in the same relative proportions as real complex mixtures, and to investigate the genotoxic effect of environmental mixtures such as EOM (extractable organic matter adsorbed to respirable air particles). We used normal human diploid lung fibroblasts (HEL) in a confluent ("quiescent") state as being closer to in vivo conditions. We determined DNA adducts by (32)P -postlabelling and the expression of p53 and p21(WAF1) proteins by Western blotting. Our results showed that the DNA binding potencies of c-PAHs in quiescent HEL cells were as follows: DB[al]P>>B[a]P>>B[b]F approximately CHRY approximately B[k]F approximately B[a]A approximately DB[ah]A approximately I[cd]P > B[ghi]P. Both DB[al]P and B[a]P were chosen to assess the genotoxic effect of other c-PAHs when present in binary mixtures. In all cases, co-treatment resulted in significantly lower DNA adduct levels as compared with adduct levels resulting from single compounds. The highest inhibitory effect was found in the presence of DB[ah]A followed by B[k]F. In both artificial and EOM mixtures a strong inhibitory effect of other compounds was observed. BPDE-adduct levels were up to five-fold lower in an artificial c-PAH mixture and up to 10-fold lower in an EOM sample than expected from applying B[a]P alone at the same concentration. These results suggest a competition for the metabolic activation enzymes, their saturation and/or inactivation by PAH metabolites. We did not observe the decrease of DNA adducts and induced levels of both p53 and p21(WAF1) proteins during the post-treatment period up to 6 days following exposure. We suggest that parent c-PAHs accumulate inside cells and that they are further activated by newly synthesised enzymes during the post-treatment period. Therefore, unaltered c-PAHs may act as a reservoir inside the cells for the long-term production of active metabolites. The total DNA adduct levels induced in HEL cells after exposure to EOM were generally low as compared with exposure to B[a]P alone under the same conditions. Human diploid lung fibroblasts possess low metabolic capacity than other human target cells. Therefore, it may be easier to competitively inhibit the metabolising enzymes in this cell system. We hypothesise that this finding might explain the generally low PAH-DNA adduct levels detected in human studies using surrogate cells such as blood WBC or lymphocytes that also possess low metabolic capacity.
authors.affiliation: Laboratory of Genetic Ecotoxicology, Institute of Experimental Medicine, Academy of Sciences of the Czech Republic, Vídenskā 1083, 142 20 Prague 4, Czech Republic. bbinkova@biomed.cas.cz
authors.forename: Blanka
authors.initials: B
authors.lastname: Binková
authors.forename: Radim J
authors.initials: RJ
authors.lastname: Srám
mesh_terms: D015123:7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide / Q000032:analysis; D001564:Benzo(a)pyrene / Q000633:toxicity; D002274:Carcinogens, Environmental / Q000633:toxicity*; D002478:Cells, Cultured; D004247:DNA / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism; D018736:DNA Adducts / Q000032:analysis / Q000187:drug effects; D004171:Diploidy; D004305:Dose-Response Relationship, Drug; D004781:Environmental Exposure; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D008168:Lung / Q000166:cytology* / Q000196:embryology; D011084:Polycyclic Aromatic Hydrocarbons / Q000633:toxicity*; D013329:Structure-Activity Relationship; D013997:Time Factors
chemical_list: D002274:Carcinogens, Environmental; D018736:DNA Adducts; D011084:Polycyclic Aromatic Hydrocarbons; D001564:Benzo(a)pyrene; D015123:7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide; D004247:DNA