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Abstract: 32P-postlabelling and PAH-ELISA using the antiserum #29 were employed to analyze DNA adducts in venous and umbilical cord blood and the placenta of 79 mothers giving birth to 80 living babies in Prague (Czech Republic). Ambient air exposure was measured by stationary measurements of basic air pollutants (PM2.5, c-PAHs) during the entire pregnancy. Tobacco smoke exposure was assessed by questionnaire data and by plasma cotinine levels. The total DNA adduct levels in the lymphocytes of mothers and newborns were elevated by 30–40% (p <0.001) compared with the placenta. B[a]P-like DNA adduct (adduct with the identical chromatographic mobility on TLC as major BPDE derived DNA adduct) levels were elevated in the blood of mothers compared with the placenta and the blood of newborns (p <0.05 and p <0.01). In tobacco smoke-exposed mothers, higher DNA adduct levels in the blood of mothers and newborns compared with the placenta were found (p <0.001), whereas the total and B[a]P-like adduct levels were comparable in the blood of mothers and newborns. B[a]P-like adducts were elevated in the blood of mothers unexposed to tobacco smoke compared with that of corresponding newborns and the placenta (p <0.01). Total and B[a]P-like DNA adducts were increased in the placenta of tobacco smoke-exposed compared with unexposed mothers (p <0.001 and p <0.01). In lymphocytes of tobacco smoke-exposed mothers, the comparison of total adduct levels (1.18±0.67 vs. 0.92±0.28) and B[a]P-like DNA adducts (0.22±0.12adducts/108 nucleotides vs. 0.15±0.06adducts/108 nucleotides) with newborns indicated a 30–40% increase of adducts in mothers. Almost equal PAH–DNA adduct levels were detected by anti-BPDE-DNA ELISA in the placenta of tobacco smoke-exposed and -unexposed mothers. Our results suggest a protective effect of the placental barrier against the genotoxic effect of some tobacco smoke components between the circulation of mother and child. We found a correlation between adduct levels in the blood of mothers and newborns. [Copyright &y& Elsevier]

Abstract: The effect of exposure to organic compounds adsorbed onto respirable air particles (<2.5μm) on DNA adducts in lymphocytes was studied in a group of non-smoking policemen (N =109, aged 35±0.9 years) working in the downtown area of Prague and spending >8h daily outdoors. Personal exposure to carcinogenic polycyclic aromatic hydrocarbons (c-PAHs) adsorbed on respirable particles was monitored in each subject for 48h before biological sampling. DNA adducts were analyzed by a 32P-postlabelling assay, and total DNA adduct levels and B[a]P-like spots were determined. Further biomarkers included cotinine levels in urine to control for exposure to tobacco smoke, plasma levels of vitamins A, E and C and polymorphisms of metabolic genotypes (GSTM1, GSTP1, GSTT1, CYP 1A1-Msp I and Ile/Val, MTHFR, MS), DNA repair genotypes (XRCC1, hOGG1 and XPD exons 6 and 23) and the p53 gene (p53 Msp I and BstU I). All the biomarkers of exposure and effect were analyzed repeatedly during a period of one year at 2–3 month intervals (January, March, June, September 2004) to cover periods with high (winter) and low (summer) levels of air pollution. The highest personal exposure to c-PAHs was found in January (8.1±8.8ng/m3), while the other three sampling periods exhibited 3–4-fold lower c-PAH exposure. The total DNA adducts were only slightly elevated in January (2.08±1.60) compared to March (1.66±0.65), June (1.96±1.73) and September (1.77±1.77). B[a]P-like DNA adducts, however, were significantly higher in January than in the March and June sampling periods (0.26±0.14 vs. 0.19±0.12 and 0.22±0.13, respectively; p <0.0001 and p =0.017) indicating that c-PAH exposure probably plays a crucial role in DNA adduct formation in lymphocytes. No effect of individual metabololic or DNA repair genotypes on DNA adduct levels was observed. However, the combination of two genotypes encoding enzymes metabolizing c-PAHs – CYP 1A1 and GSTM1 – was associated with the levels of total and B[a]P-like DNA adducts under conditions of increased exposure to c-PAHs. Our study suggests that DNA adducts in the lymphocytes of subjects exposed to increased c-PAH levels are an appropriate biomarker of a biologically effective dose, directly indicating whether or not the extent of exposure to these compounds is related to an increased mutagenic and carcinogenic risk. [Copyright &y& Elsevier]

Abstract: Acellular assay of calf thymus DNA±rat liver microsomal S9 fraction coupled with 32P-postlabelling was used to study the genotoxic potential of organic compounds bound onto PM10 particles collected in three European cities—Prague (CZ), Kosice (SK) and Sofia (BG) during summer and winter periods. B[a]P alone induced DNA adduct levels ranging from 4.8 to 768 adducts/108 nucleotides in the concentration dependent manner. However, a mixture of 8 c-PAHs with equimolar doses of B[a]P induced 3.7–757 adducts/108 nucleotides, thus suggesting the inhibition of DNA adduct forming activity by interaction among various PAHs. Comparison of DNA adduct levels induced by various EOMs indicates higher variability among seasons than among localities. DNA adduct levels for Prague collection site varied from 19 to 166 adducts/108 nucleotides, for Kosice from 22 to 85 and for Sofia from 6 to 144 adducts/108 nucleotides. Bioactivation with S9 microsomal fraction caused 2- to 7-fold increase in DNA adduct levels compared to −S9 samples, suggesting a crucial role of indirectly acting genotoxic EOM components, such as PAHs. We have demonstrated for the first time a significant positive correlation between B[a]P content in EOMs and total DNA adduct levels detected in the EOM treated samples (R =0.83; p =0.04). These results suggest that B[a]P content in EOM is an important factor for the total genotoxic potential of EOM and/or B[a]P is a good indicator of the presence of other genotoxic compounds causing DNA adducts. Even stronger correlation between the content of genotoxic compounds in EOMs and total DNA adduct levels detected (R =0.94; p =0.005) was found when eight c-PAHs were taken into the consideration. Our findings support a hypothesis that a relatively limited number of EOM components is responsible for a major part of its genotoxicity detectable as DNA adducts by 32P-postlabelling. [Copyright &y& Elsevier]

Abstract: The potential of two asbestos substitute mineral fibres – rock (stone) wool RW1 and glass wool MMVF10 – to induce gene mutations, DNA strand breaks, inflammation and oxidative stress has been studied in rats. Male homozygous λ-lacI transgenic F344 rats were intratracheally instilled with single doses of 1 and 2mg/animal of fibres or with multiple doses of 2mg/animal administered weekly on four consecutive weeks (8mg in total). Exposure to RW1 fibres for 16 weeks significantly increased mutant frequency (MF) in the lung in a dose-dependent manner, while MMVF10 fibres did not exhibit any increase of MF at any dose. RW1 fibres gave a significant increase of MF at a dose of 1mg. Four weeks after instillation, neither the single nor the multiple doses significantly increased MF for both fibre types. To investigate mechanisms for induction of mutations, other genotoxicity markers and parameters of inflammatory and oxidative damage were determined in relation to MF. A weak correlation of mutagenicity data with other genotoxicity parameters studied was observed. DNA strand breaks as measured by comet assay were increased in alveolar macrophages and lung epithelial cells of RW1 and MMVF10 treated rats. RW1 fibres caused more extensive lung inflammation as measured by release of neutrophils into broncho-alveolar lavage fluid than MMVF10 fibres. The effects were observed 16 weeks post-exposure, indicating a persistence of the pathogenic process during the exposure period. Only minor differences in the extent of inflammatory processes were observed between the doses of 2mg and 4×2mg, suggesting that any threshold for inflammation lies below the dose of 2mg. With the exception of the highest dose of MMVF10 fibres after 16 weeks of exposure, no significant increase of oxidative damage as measured by levels of malondialdehyde in lung tissue was observed. MMVF10 fibres caused weaker inflammation in the lung of rats and did not exhibit any mutagenic effect. We conclude that a weak but chronic inflammation (more likely than acute inflammation or direct oxidative damage) in the lung tissue of fibre treated rats characterized by moderate influx of inflammatory cells into BAL is probably responsible for the observed mutagenic effect of RW1 fibres. [Copyright &y& Elsevier]

Abstract: In an attempt to examine the interaction of man-made mineral fibres with benzo[a]pyrene (B[a]P), homozygous λ-lacI transgenic F344 rats were intratracheally treated with rock (stone) wool RW1 and glass wool MMVF 10 fibres together with B[a]P. To analyze the induction of gene mutations by fibres and B[a]P in lung, single doses of 1 and 2mg fibres/animal or multiple doses of 2mg fibres/animal were administered weekly on 4 consecutive weeks (total dose 8mg/animal). B[a]P (10mg/animal) was administered either simultaneously with fibres (for single dose treatment with fibres) or together with the last fiber treatment (for multiple dose treatment with fibres). Animals were scarified 4 weeks after the last treatment. Benzo[a]pyrene administered simultaneously with RW1 fibres exhibited a strong synergistic effect on mutagenicity, the observed mutant frequency (MF) being more than three-fold higher than the net sum of the MF induced after separate administration of both agents. Our data suggest that DNA adducts induced by simultaneous B[a]P and fiber treatment lead to a strong increase in mutatant frequencies. [Copyright &y& Elsevier]

In order to get more insight into the mechanism of asbestos-related lung cancer, the mutagenic potential of asbestos was examined in vivo in rat lung. Groups of five transgenic λ-lacI (Big Blue™) rats were intratracheally instilled with single doses of 1 or 2 mg, or with four weekly doses of 2 mg, per animal of the amosite asbestos. Sixteen weeks after instillation, the mutation frequency was found to be increased in lung DNA by 2-fold at doses of 2 mg (P = 0.035) and of 4 × 2 mg (P = 0.007) amosite. No significant changes were observed after 4 weeks of exposure. In separate experiments, wild-type F344 rats were treated by the same regimen as described above and markers of inflammation, genotoxicity, cell proliferation and lung tissue damage were analysed. Our results indicate a weak but persistent inflammation and cell proliferation which possibly plays a major role in the observed mutagenic effect. [Copyright &y& Elsevier]

To study the suspected mechanism of the interaction between tobacco smoking and asbestos exposure in the modulation of cancer risk, the mutagenic potential of asbestos in combination with the tobacco smoke carcinogen benzo[a]pyrene (B[a]P) was examined in vivo in the rat lung. B[a]P was administered intratracheally in one set of experiments, or by two daily intraperitoneal injections in another set of experiments, to λlacI transgenic rats, together with 1, 2 or 4 × 2 mg amosite in one experiment. In the first experiment, the combined action of amosite and B[a]P caused a synergistic (superadditive) increase of mutation frequency in the lung, as compared to groups treated only with asbestos or B[a]P. In the second experiment, i.p. treatment with B[a]P did not significantly alter the mutation frequency induced by amosite, neither after 4 nor after 16 weeks of exposure. The B[a]P-DNA adduct levels were unaffected by amosite co-treatment in both experiments. We assume that the synergistic increase of mutation frequency after intratracheal treatment was due to the mitogenic activities of B[a]P and of amosite. In conclusion, our findings indicate that a weak and delayed mutagenic effect of amosite in rat lung observed in another study was strongly enhanced by the concomitant action of B[a]P. The striking enhancement effect of B[a]P may provide a basis for understanding the suspected synergism of smoking on asbestos carcinogenesis. [Copyright &y& Elsevier]

Kearney's bluestar ( Amsonia kearneyana) is a highly endangered herbaceous perennial in the family Apocynaceae. The species is found only in the Baboquivari Mountains of southern Arizona. We report the isolation and development of 12 microsatellite loci for Kearney's bluestar. Numbers of alleles ranged from two to four and observed heterozygosities ranged from 0.20 to 0.80 in the nine loci found to be polymorphic in the test population. All loci were also tested for cross-amplification in five other Amsonia species representing two subgenera from the southwestern United States. Some loci that were not polymorphic in the Kearney's bluestar were polymorphic in other species. [ABSTRACT FROM AUTHOR]

Abstract: Principal aims of this study were at first, to find a relevant human derived cell line to investigate the genotoxic potential of PAH-containing complex mixtures and second, to use this cell system for the analysis of DNA adduct forming activity of organic compounds bound onto PM10 particles. Particles were collected by high volume air samplers during summer and winter periods in three European cities (Prague, Kosice, and Sofia), representing different levels of air pollution. The genotoxic potential of extractable organic matter (EOM) was compared with the genotoxic potential of individual carcinogenic polycyclic aromatic hydrocarbons (c-PAHs) as well as their artificial mixtures. Metabolically competent human hepatoma HepG2 cells, confluent cultures of human diploid lung fibroblasts (HEL), and the human monocytic leukemia cell line THP-1 were used as models. DNA adducts were analyzed by 32P-postlabeling. The total DNA adduct levels induced in HepG2 cells after exposure to EOMs were higher than in HEL cells treated under the same conditions (15–190 versus 2–15adducts/108 nucleotides, in HepG2 and HEL cells, respectively). THP-1 cells exhibited the lowest DNA adduct forming activity induced by EOMs (1.5–3.7adducts/108 nucleotides). A direct correlation between total DNA adduct levels and c-PAH content in EOM was found for all EOMs in HepG2 cells incubated with 50μg EOM/ml (R =0.88; p =0.0192). This correlation was even slightly stronger when B[a]P content in EOMs and B[a]P-like adduct spots were analyzed (R =0.90; p =0.016). As THP-1 cells possess a limited metabolic capacity for most c-PAHs to form DNA reactive intermediates and are also more susceptible to toxic effects of PAHs and various EOM components, this cell line seemed to be an inappropriate system for genotoxicity studies of PAH-containing complex mixtures. The seasonal variability of genotoxic potential of extracts was stronger than variability among the three localities studied. In HepG2 cells, the highest DNA adduct levels were induced by EOM collected in Prague in the winter period, followed by Sofia and Kosice. However, in the summer sampling period, the order was quite opposite: Kosice>Sofia>Prague. When the EOM content per m3 of air was taken into consideration in order to compare real exposures of humans to genotoxic compounds in all three localities, extracts from respirable dust particles collected in Sofia exhibited the highest genotoxicity regardless of the sampling period. The results indicate that most of DNA adducts detected in cells incubated with EOMs have their origin in low concentrations of c-PAHs representing 0.03–0.17% of EOM total mass. Finally, our results suggest that HepG2 cells have a metabolic capacity for PAHs similar to human hepatocytes and represent therefore the best in vitro model for investigating the genotoxic potential of complex mixtures containing PAHs among the three cell lines tested in this study. [Copyright &y& Elsevier]

We developed 26 Swainson's hawk ( Buteo swainsoni) microsatellite primers from CA, AAT, CATC and GAGAA enriched genomic libraries. Primers were tested in 357 Swainson's hawks from western North America as well as seven other Buteo species. These markers will have broad application in investigations of Buteo population structure and genetic diversity. [ABSTRACT FROM AUTHOR]

Little is known about the impact of genetic variation on the genetic damage induced by urban air pollution or environmental tobacco smoke (ETS) in exposed populations. The levels of bulky DNA adducts (32P-postlabelling, nuclease P1 enrichment) and chromosomal aberrations were measured in lymphocytes of 194 non-smoking students living in the city of Athens, and the rural region of Halkida, Greece. In these individuals personal exposure to PAH was also measured. Furthermore, genetic polymorphisms were examined in cytochromes P450 1A1, 1B1, in the GSTM1, GSTP1 and GSTT1 as well as in microsomal epoxy hydrolase (EPHX) genes. Subjects with the CYP1&ast;2A mutant genotype also suffering significant ETS exposure tended to exhibit higher adduct levels and % aberrant cells. In contrast, CYP1B1 polymorphisms seemed to have an impact on the DNA adduct levels only among individual with negligible ETS exposure. Subjects carrying both the CYP1&ast;2A mutant genotype and the GSTM1 null genotype tended to have higher DNA adduct levels. A similar effect was also observed with the combined CYP1A1&ast;2A/GSTP1 (Ile/Val) and the CYP1A1&ast;2A/mEH “slow” polymorphisms. In both cases, the effect was more pronounced among subjects with higher levels of ETS exposure. Stepwise restriction of the observations to subjects characterised by (a) GSTP1 mutant, (b) GSTM1 null, (c) mEH “slow” (His139His) genotypes and (d) ETS exposure resulted in a significant trend of increasing DNA adduct levels only among individuals with at least one CYP1A1&ast;2A mutated allele, illustrating the importance and complexity of gene-exposure and gene–gene interactions in determining the level of genotoxic damage on an individual levels. [Copyright &y& Elsevier]

DNA damage can impair normal cellular functions and result in various pathophysiological processes including cardiovascular diseases and cancer. We compared the genotoxic potential of diverse DNA damaging agents, and focused on their effects on the DNA damage response (DDR) and cell fate in human lung cells BEAS-2B. Polycyclic aromatic hydrocarbons [PAHs; benzo[ a ]pyrene (B[ a ]P), 1-nitropyrene (1-NP)] induced DNA strand breaks and oxidative damage to DNA; anticancer drugs doxorubicin (DOX) and 5-bromo-2′-deoxyuridine (BrdU) were less effective. DOX triggered the most robust p53 signaling indicating activation of DDR, followed by cell cycle arrest in the G2/M phase, induction of apoptosis and senescence, possibly due to the severe and irreparable DNA lesions. BrdU not only activated p53, but also increased the percentage of G1-phased cells and caused a massive accumulation of senescent cells. In contrast, regardless the activation of p53, both PAHs did not substantially affect the cell cycle distribution or senescence. Finally, a small fraction of cells accumulated only in the G2/M phase and exhibited increased cell death after the prolonged incubation with B[ a ]P. Overall, we characterized differential responses to diverse DNA damaging agents resulting in specific cell fate and highlighted the key role of DNA lesion type and the p53 signaling persistence. • PAHs induced considerably higher level of DNA damage than DOX and BrdU. • DOX treatment was more toxic and led to G2 arrest, senescence and apoptosis. • BrdU triggered senescence possibly due to an alternative mechanism. • Despite slight increase of apoptosis, PAHs displayed better survival of cells. [ABSTRACT FROM AUTHOR]

Abstract TiO 2 along with nano-TiO 2 are commonly found in consumer products. In vivo studies have observed an accumulation of nano-TiO 2 in macrophages. However, characteristics of nano-TiO 2 determining toxicity remain unclear. In our study, the cytotoxic effects of 14 diverse nano-TiO 2 on THP-1 macrophage-like cells were measured by 3 cytotoxicity assays (MTS, WST-1 and LDH). Total averaged cytotoxicity was calculated using principal component analysis. Characteristics of all 14 nano-TiO 2 included hydrodynamic diameter, zeta potential, shape, polydispersity index (PDI) and concentration; moreover, crystal form, specific surface area and crystallite size were measured for 10 nano-TiO 2. The variables affecting cytotoxicity were chosen using LASSO (least absolute shrinkage and selection operator). Except for concentration, PDI in media measured within 1 h after preparation of the nanomaterial dispersion was selected as a variable affecting cytotoxicity: stable dispersion resulted in higher cytotoxic effects. Crystallite size has been shown to have nonlinear effects (particles of sizes between 20 and 60 nm were cytotoxic while smaller and larger ones were not) and thus it has been excluded from LASSO. The shape (particles/fibre) and crystal form did not affect the cytotoxicity. PDI and the nonlinear effect of size could be an explanation for the inconsistencies of the cytotoxicity of nano-TiO 2 in various studies. Graphical abstract Unlabelled Image Highlights • Physical factors affecting cytotoxicity of nano-TiO 2 in THP-1 cells were studied • Concentration and polydispersity index measured within 1 h affected cytotoxicity • Particles of 20–60 nm were cytotoxic while smaller and larger ones were not • No significant difference between anatase/rutile crystallinity was found • Overall cytotoxic effects of nano-TiO 2 in THP-1 macrophage-like cells were low [ABSTRACT FROM AUTHOR]

Abstract: Oxidative damage to macromolecules may have numerous negative health consequences. We measured oxidative damage to DNA, proteins and lipids in 80 newborns and 79 mothers, analyzed the effect of mother''s tobacco smoke exposure on oxidative stress, and assessed correlations between oxidative stress markers and bulky and PAH (polycyclic aromatic hydrocarbons)-specific DNA adducts. Mean levels (±S.D.) of 8-oxodeoxyguanosine (8-oxodG) per 105 dG in the placenta were 2.85±0.78; we did not see a difference between 8-oxodG levels in newborns born to mothers exposed and unexposed to tobacco smoke. Protein carbonyl levels, a marker of protein oxidation, were comparable in the umbilical cord and in maternal venous blood plasma (17.4±3.2 and 17.6±4.2nmol/ml plasma in newborns and mothers, respectively, p =0.66). Lipid peroxidation measured as levels of 15-F2t-isoprostane (15-F2t-IsoP) in plasma was significantly higher in newborns than in mothers (362±129 and 252±130pg/ml in newborns and mothers, respectively, p <0.001). We did not find any effect of tobacco smoke exposure on either biomarker in any group. Levels of both protein carbonyls and 15-F2t-IsoP in cord blood significantly correlated with those in maternal plasma (p <0.001). 8-oxodG levels positively correlated with plasma carbonyls in cord plasma, as well as with cotinine levels (marker of tobacco smoke exposure) in maternal plasma. 8-oxodG levels also correlated with bulky DNA adducts in lymphocyte DNA of newborns and mothers and with PAH-DNA adducts in the placenta. Our results showed higher lipid peroxidation in newborns than in mothers, close correlation of analyzed oxidative stress markers between newborns and mothers, and a relationship between oxidative stress and induction of DNA adducts. [Copyright &y& Elsevier]

Abstract: Genetic polymorphisms were examined using direct sequencing of the hypervariable region II (HVRII) in the D-loop of mtDNA in the cord blood of 355 children living in two areas of the Czech Republic – the industrial district of Teplice and the agricultural district of Prachatice. The incidence of the most frequent nucleotide variants of HVRII, C150T (10.1%), T152C (19.7%), T195C (19.7%) and 309.nC (41.4% for 309.2C and 13.8% for 309.3C), and the respiratory morbidity at the ages of 0–2 years and 2–6 years were investigated, considering many other factors such as locality, gender, ethnicity, heating by coal in household, maternal age, asthma bronchiale, allergic rhinitis, pollinosis, conjunctivitis and maternal tobacco exposure during and after pregnancy. We found that the T195C transversion in HVRII is connected with an increased risk of early childhood (0–2 years) bronchitis (RR 1.38, p =0.034, 95% CI 1.04–1.85) and with increased risk of otitis media in children aged 2–6 years (RR 1.62, p =0.032, 95% CI 1.04–2.53). Another polymorphism, 309.nC, is associated with an increased risk of bronchitis in children aged 2–6 years (RR 1.46, p =0.030, 95% CI 1.04–2.06). The results indicate that genetic polymorphisms in mtDNA may be an important factor not only for various types of cancers and neurodegenerative diseases, but also for respiratory morbidity in children. [Copyright &y& Elsevier]

Abstract: Methylated chrysenes (MeChry) are important cigarette smoke constituents and 5-MeChry has been listed as possibly carcinogenic to humans. Although a major attention has been in past paid especially to mutagenic, tumor-initiating effects of MeChry, little is known about toxic effects of MeChry related to tumor promotion. As the position of methyl group has been repeatedly observed to determine genotoxic effects of MeChry, we examined both genotoxic and nongenotoxic effects of MeChry, using rat liver cell lines as experimental models. All six MeChry were relatively efficient aryl hydrocarbon receptor (AhR) agonists, with 3- and 6-MeChry being the most potent inducers of the AhR-mediated reporter gene activity. All six compounds disrupted contact inhibition in rat liver epithelial WB-F344 cells, a process previously reported to be AhR-dependent, suggesting that MeChry may interfere with cell cycle control in an AhR-dependent manner. In contrast, only 5- and 6-MeChry were found to acutely inhibit gap junctional intercellular communication (GJIC), another parameter correlating with tumor promoting effects of xenobiotics. Both 5- and 6-MeChry were efficient inducers of mRNA expression of enzymes involved in metabolic activation of polycyclic aromatic hydrocarbons, including cytochromes P450 1A1/1B1 and aldo-keto reductase 1C9. However, only 5-MeChry, and not 6-MeChry, induced significant formation of DNA adducts in rat liver epithelial cells, which corresponded with its ability to induce high accumulation of cells in S-phase. On the other hand, 5-MeChry induced neither apoptosis related to DNA damage nor phosphorylation of p53 tumor suppressor. Taken together, our results suggest that methyl group position may affect both genotoxic and nongenotoxic effects of MeChry, such as formation of DNA adducts and inhibition of GJIC. All MeChry showed a potency to disrupt cell proliferation control, while 5-MeChry was a single compound inducing DNA damage, disruption of cell cycle control and inhibition of GJIC in rat liver cells. [Copyright &y& Elsevier]

Abstract: The main aim of this study was to compare the genotoxic potential of organic extracts from urban air particles collected in three different sampling periods in the center of Prague (Czech Republic). For this purpose, we analyzed the DNA adduct forming activity of extractable organic matter (EOM) from urban air particles <10μm (PM10) in the human hepatoma cell line HepG2. DNA adducts were analyzed by 32P-postlabelling with nuclease P1 enrichment. PM10 concentrations were 36.9μg/m3, 62.6μg/m3 and 39.0μg/m3, in summer 2000, winter 2001 and winter 2005, respectively. The corresponding EOM contents were 5.0μg/m3 (13.9% of PM10), 14.9μg/m3 (23.8%) and 6.7μg/m3 (17.2%). The total DNA adduct levels induced by 10μg EOM/ml were 4.7, 19.5 and 37.2 adducts/108 nucleotides in summer 2000, winter 2001 and winter 2005, respectively. However, when the EOM quantities per cubic meter of air were taken into consideration, the summer sample exhibited a 10-fold lower genotoxicity than did those of winter, while the difference between the winter samples was not significant: 23.4 in summer 2000, 291 in winter 2001 and 249 in winter 2005 (in relative units). Although the PM10 concentration in air and the EOM content in particles in winter 2005 were significantly lower than in winter 2001, the genotoxic potential of the ambient air in these samples was almost equal. There were significant positive correlations between the B[a]P and c-PAH content in EOM from various sampling periods and the total DNA adduct levels detected in the EOM-treated samples. These findings support the hypothesis that the B[a]P and c-PAH content in EOM is the most important factor that determines its genotoxic potential. Thus, estimating the genotoxic potential of the ambient air and predicting health risk should be based mainly on the c-PAH concentration and the biological activity of the extracts, while the mass of particles and the EOM content do not seem to be crucial determinants of ambient air genotoxicity. [Copyright &y& Elsevier]

Abstract: Polycyclic aromatic hydrocarbons (PAHs) with molecular weight 278 are a group of PAHs that are mostly not covered by the current monitoring programs, despite their relative abundance in environmental samples and possible carcinogenicity. Although benzo[g]chrysene (BgChry) and dibenz[a,h]anthracene (DBahA) have been for a long time studied as genotoxic, tumour-initiating compounds, little is known about the potential tumour-promoting effects of this group of PAHs. In the present study, we investigated their impact on activation of the aryl hydrocarbon receptor (AhR), induction of enzymes involved in metabolic activation of PAHs, disruption of cell cycle control in confluent cell population and inhibition of gap junctional intercellular communication (GJIC), using the rat liver epithelial cell line WB-F344 as a model of liver progenitor cells. We found that BgChry was the weakest inducer of the AhR-mediated activity, while relative potencies of benzo[b]chrysene (BbChry) and benzo[c]chrysene (BcChry) were comparable to the previously reported values for dibenzanthracenes. All compounds increased expression of cytochromes P450 1A1 and 1B1, and aldo-keto reductase 1C9. BgChry was found to induce high amounts of DNA adducts, which corresponded with induction of p53 phosphorylation at Ser15, apoptosis and accumulation of cells in S-phase of cell cycle, leading to a decrease in cell numbers. All other compounds were found to stimulate cell proliferation in contact-inhibited WB-F344 cells in a dose-dependent manner. We found that only BgChry, and to a lesser extent also BcChry, inhibited GJIC at high concentrations. Taken together, dibenzanthracenes and benzochrysenes, with exception of BgChry, seem to act primarily through deregulation of cell proliferation in liver epithelial cells, which is related to their relatively high AhR-mediated activity. The disruption of cell cycle control might contribute to their carcinogenic effects, as well as to carcinogenicity of complex environmental mixtures containing high levels of PAHs with molecular weight 278. [Copyright &y& Elsevier]