The effect of high polycyclic aromatic hydrocarbon exposure on biological aging indicators.

BACKGROUND: Aging represents a serious health and socioeconomic concern for our society. However, not all people age in the same way and air pollution has been shown to largely impact this process. We explored whether polycyclic aromatic hydrocarbons (PAHs), excellent fossil and wood burning tracers, accelerate biological aging detected by lymphocytes DNA methylation age (DNAmAge) and telomere length (TL), early nuclear DNA (nDNA) hallmarks of non-mitotic and mitotic cellular aging, and mitochondrial DNA copy number (mtDNAcn). METHODS: The study population consisted of 49 male noncurrent-smoking coke-oven workers and 44 matched controls. Occupational and environmental sources of PAH exposures were evaluated by structured questionnaire and internal dose (urinary 1-pyrenol). We estimated Occup_PAHs, the product of 1-pyrenol and years of employment as coke-oven workers, and Environ_PAHs, from multiple items (diet, indoor and outdoor). Biological aging was determined by DNAmAge, via pyrosequencing, and by TL and mtDNAcn, via quantitative polymerase chain reaction. Genomic instability markers in lymphocytes as target dose [anti-benzo[a]pyrene diolepoxide (anti-BPDE)-DNA adduct], genetic instability (micronuclei), gene-specific (p53, IL6 and HIC1) and global (Alu and LINE-1 repeats) DNA methylation, and genetic polymorphisms (GSTM1) were also evaluated in the latent variable nDNA_changes. Structural equation modelling (SEM) analysis evaluated these multifaceted relationships. RESULTS: In univariate analysis, biological aging was higher in coke-oven workers than controls as detected by higher percentage of subjects with biological age older than chronological age (AgeAcc ≥ 0, p = 0.007) and TL (p = 0.038), mtDNAcn was instead similar. Genomic instability, i.e., genotoxic and epigenetic alterations (LINE-1, p53 and Alu) and latent variable nDNA_changes were higher in workers (p < 0.001). In SEM analysis, DNAmAge and TL were positively correlated with Occup_PAHs (p < 0.0001). Instead, mtDNAcn is positively correlated with the latent variable nDNA_changes (p < 0.0001) which is in turn triggered by Occup_PAHs and Environ_PAHs. CONCLUSIONS: Occupational PAHs exposure influences DNAmAge and TL, suggesting that PAHs target both non-mitotic and mitotic mechanisms and made coke-oven workers biologically older. Also, differences in mtDNAcn, which is modified through nDNA alterations, triggered by environmental and occupational PAH exposure, suggested a nuclear-mitochondrial core-axis of aging. By decreasing this risky gerontogenic exposure, biological aging and the consequent age-related diseases could be prevented.

Association between a urinary biomarker for exposure to PAH and blood level of the acute phase protein serum amyloid A in coke oven workers.

BACKGROUND: Coke oven workers are exposed to both free and particle bound PAH. Through this exposure, the workers may be at increased risk of cardiovascular diseases. Systemic levels of acute phase response proteins have been linked to cardiovascular disease in epidemiological studies, suggesting it as a marker of these conditions. The aim of this study was to assess whether there was association between PAH exposure and the blood level of the acute phase inflammatory response marker serum amyloid A (SAA) in coke oven workers. METHODS: A total of 87 male Polish coke oven workers from two different plants comprised the study population. Exposure was assessed by means of the individual post-shift urinary excretion of 1-hydroxypyrene, as internal dose of short-term PAH exposure, and by anti-benzo[a]pyrene diolepoxide (anti-B[a]PDE)-DNA), as a biomarker of long-term PAH exposure. Blood levels of acute phase proteins SAA and CRP were measured by immunoassay. C-reactive protein (CRP) levels were included to adjust for baseline levels of SAA. RESULTS: Multiple linear regression showed that the major determinants of increased SAA levels were urinary 1-hydroxypyrene (beta = 0.56, p = 0.030) and serum CRP levels (beta = 7.08; p < 0.0001) whereas anti-B[a]PDE-DNA, the GSTM1 detoxifying genotype, diet, and smoking were not associated with SAA levels. CONCLUSIONS: Urinary 1-hydroxypyrene as biomarker of short-term PAH exposure and serum levels of CRP were predictive of serum levels of SAA in coke oven workers. Our data suggest that exposure of coke oven workers to PAH can lead to increased systemic acute response and therefore potentially increased risk of cardiovascular disease.

Inter-laboratory consistency and variability in the buccal micronucleus cytome assay depends on biomarker scored and laboratory experience: results from the HUMNxl international inter-laboratory scoring exercise.

The buccal micronucleus cytome (BMNcyt) assay in uncultured exfoliated epithelial cells from oral mucosa is widely applied in biomonitoring human exposures to genotoxic agents and is also proposed as a suitable test for prescreening and follow-up of precancerous oral lesions. The main limitation of the assay is the large variability observed in the baseline values of micronuclei (MNi) and other nuclear anomalies mainly related to different scoring criteria. The aim of this international collaborative study, involving laboratories with different level of experience, was to evaluate the inter- and intra-laboratory variations in the BMNcyt parameters, using recently implemented guidelines, in scoring cells from the same pooled samples obtained from healthy subjects (control group) and from cancer patients undergoing radiotherapy (treated group). The results indicate that all laboratories correctly discriminated samples from the two groups by a significant increase of micronucleus (MN) and nuclear bud (NBUD) frequencies and differentiated binucleated (BN) cells, associated with the exposure to ionizing radiation. The experience of the laboratories was shown to play an important role in the identification of the different cell types and nuclear anomalies. MN frequency in differentiated mononucleated (MONO) and BN cells showed the greatest consistency among the laboratories and low variability was also detected in the frequencies of MONO and BN cells. A larger variability was observed in classifying the different cell types, indicating the subjectivity in the interpretation of some of the scoring criteria while reproducibility of the results between scoring sessions was very good. An inter-laboratory calibration exercise is strongly recommended before starting studies with BMNcyt assay involving multiple research centers.

Influence of genetic polymorphisms on biomarkers of exposure and effects in children living in Upper Silesia.

This article is a follow-up to our previous molecular epidemiology studies on the DNA damage in children from the Upper Silesia region of Poland. It is expected that metabolic and DNA repair gene polymorphisms may modulate individual susceptibility to environmental exposure. In this study, we investigate the association between polymorphisms of metabolising (CYP2D, EPHX1, GSTM1, GSTP1, GSTT1, NAT2) and DNA repair (XPD, XRCC1, XRCC3) genes and selected biomarkers of exposure and effect such as levels of 1-hydroxypyrene (1-OHP) and urinary mutagenicity, aromatic DNA adducts, sister chromatid exchange (SCE) and micronuclei (MN) in 74 children. Both 1-OHP concentration and urinary mutagenicity tested by TA98+S9 were significantly higher in individuals with EPHX1 (exon 4) Arg/Arg genotype than in individuals with other genotype. The EPHX1 (exon 3) significantly affected urinary mutagenicity tested with strain YG1024+S9. The urinary mutagenicity in individuals with Tyr/Tyr homozygotes was lower than in individuals with Tyr/His and His/His (1057±685 vs. 1432±1003 revertants/mol creatinine). XRCC3 Met/Met genotype was associated with significantly higher levels of 1-OHP in urine compared with only The/Met genotype. The PAH-DNA adduct levels in the subgroup with GSTM1 null genotype was 2-fold higher than in individuals with GSTM1 active (7.06±5.12 vs. 13.14±9.81 adduct/10(8) nucleotides). The mean level of aromatic DNA adducts in children with deletion of the GSTT1 gene was significantly higher compared with individuals with that gene present (8.03±6.23 vs. 14.66±10.70 adduct/10(8) nucleotides). Also the carriers of the XPD Lys/Lys genotype showed higher levels of DNA adducts than heterozygotes (13.16±9.70 vs. 6.81±5.86 adducts/10(8) nucleotides). Children carrying the XRCC3-241 Met/Met genotype exhibited a higher number of SCE in peripheral blood lymphocytes than carriers of Thr/Met allele (8.15±0.86 vs. 7.62±0.79 SCE/cell). It was also observed that children with the GSTP1 slow conjugator had significantly elevated MN in peripheral blood lymphocytes compared with fast conjugator (4.23±3.49 vs. 6.56±5.00 MN/1000 cells).

Urinary Mercapturic Acids to Assess Exposure to Benzene and Other Volatile Organic Compounds in Coke Oven Workers.

Coke production was classified as carcinogenic to humans by the International Agency for Research on Cancer. Besides polycyclic aromatic hydrocarbons, coke oven workers may be exposed to benzene and other volatile organic compounds (VOCs). The aim of this study was to assess the exposure to several VOCs in 49 coke oven workers and 49 individuals living in the same area by determining urinary mercapturic acids. Active tobacco smoking was an exclusion criterion for both groups. Mercapturic acids were investigated by a validated isotopic dilution LC-MS/MS method. Linear models were built to correct for different confounding variables. Urinary levels of N-acetyl-S-phenyl-L-cysteine (SPMA) (metabolite of benzene), N-acetyl-S-(2-hydroxy-1/2-phenylethyl)-L-cysteine (PHEMA) (metabolite of styrene), N-acetyl-S-(2-cyanoethyl)-L-cysteine (CEMA) (metabolite of acrylonitrile), N-acetyl-S-[1-(hydroxymethyl)-2-propen-1-yl)-L-cysteine and N-acetyl-S-(2-hydroxy-3-buten-1-yl)-L-cysteine (MHBMA) (metabolites of 1,3-butadiene) were 2-10 fold higher in workers than in controls (p < 0.05). For SPMA, in particular, median levels were 0.02 and 0.31 µg/g creatinine in workers and controls, respectively. Among workers, coke makers were more exposed to PHEMA and SPMA than foremen and engine operators. The comparison with biological limit values shows that the exposure of workers was within 20% of the limit values for all biomarkers, moreover three subjects exceeded the restrictive occupational limit value recently proposed by the European Chemicals Agency (ECHA) for SPMA.

Polycyclic aromatic hydrocarbons bound to outdoor and indoor airborne particles (PM2.5) and their mutagenicity and carcinogenicity in Silesian kindergartens, Poland.

Assessment of exposure to polycyclic aromatic hydrocarbons (PAHs) is important due to the widespread presence of PAHs in the environment and their toxicological relevance, especially to susceptible populations such as children and their health. The aim of this study is to compare indoor and outdoor concentrations of particulate matter with a diameter of 2.5 µm or less (PM2.5) and 15 individual PAHs, as well as contribution of the analyzed PAHs to mutagenic and carcinogenic activity. Samples were collected during spring season in two sites in southern Poland (Silesia) representing urban and rural areas. Indoor samples of PM2.5 were sampled in kindergartens. At the same time, in the vicinity of the kindergarten buildings, the collection of the outdoor PM2.5 samples was carried out. Mutagenic (MEQ) and carcinogenic (TEQ) equivalents related to BaP and the percentage share expressed as mutagenic (MP) and carcinogenic (CP) potential of each individual compound to the total mutagenic/carcinogenic potential of the PAH mixture were calculated. The obtained results show that high concentrations of PM2.5 (above 25 µg/m3) and 15 PM2.5-bound PAHs in outdoor and indoor air were similar in the two studied areas. In overall PAHs mutagenic and carcinogenic potential, the percentage share of benzo(a)pyrene (BaP) was dominant and varied from 49.0-54.5% to 62.5-70.0%, respectively. The carried out study indicates the necessity of reducing PAH emission from solid fuel combustion, which is reflected in PM2.5-bound PAHs concentrations and their diagnostic ratios. In the recent years, health effects on children resulting from their activity pattern and air quality in the public places have been a serious problem.

Indoor air quality in urban and rural kindergartens: short-term studies in Silesia, Poland.

More than 80% of people living in urban areas who monitor air pollution are exposed to air quality levels that exceed limits defined by the World Health Organization (WHO). Although all regions of the world are affected, populations in low-income cities are the most impacted. According to average annual levels of fine particulate matter (PM2.5, ambient particles with aerodynamic diameter of 2.5 µm or less) presented in the urban air quality database issued by WHO in 2016, as many as 33 Polish cities are among the 50 most polluted cities in the European Union (EU), with Silesian cities topping the list. The aim of this study was to characterize the indoor air quality in Silesian kindergartens based on the concentrations of gaseous compounds (SO2, NO2), PM2.5, and the sum of 15 PM2.5-bound polycyclic aromatic hydrocarbons (PAHs), including PM2.5-bound benzo(a)pyrene (BaP), as well as the mutagenic activity of PM2.5 organic extracts in Salmonella assay (strains: TA98, YG1024). The assessment of the indoor air quality was performed taking into consideration the pollution of the atmospheric air (outdoor). I/O ratios (indoor/outdoor concentration) for each investigated parameter were also calculated. Twenty-four-hour samples of PM2.5, SO2, and NO2 were collected during spring in two sites in southern Poland (Silesia), representing urban and rural areas. Indoor samples were taken in naturally ventilated kindergartens. At the same time, in the vicinity of the kindergarten buildings, the collection of outdoor samples of PM2.5, SO2, and NO2 was carried out. The content of BaP and the sum of 15 studied PAHs was determined in each 24-h sample of PM2.5 (indoor and outdoor). In the urban site, statistically lower concentrations of SO2 and NO2 were detected indoors compared to outdoors, whereas in the rural site, such a relationship was observed only for NO2. No statistically significant differences in the concentrations of PM2.5, PM2.5-bound BaP, and Σ15 PAHs in kindergartens (indoor) versus atmospheric (outdoor) air in the two studied areas were identified. Mutagenic effect of indoor PM2.5 samples was twice as low as in outdoor samples. The I/O ratios indicated that all studied air pollutants in the urban kindergarten originated from the ambient air. In the rural site concentrations of SO2, PM2.5 and BaP in the kindergarten were influenced by internal sources (gas and coal stoves).

Mitochondrial DNA copy number and exposure to polycyclic aromatic hydrocarbons.

BACKGROUND: Increased mitochondrial DNA copy number (mtDNAcn) is a biologic response to mtDNA damage and dysfunction, predictive of lung cancer risk. Polycyclic aromatic hydrocarbons (PAHs) are established lung carcinogens and may cause mitochondrial toxicity. Whether PAH exposure and PAH-related nuclear DNA (nDNA) genotoxic effects are linked with increased mtDNAcn has never been evaluated. METHODS: We investigated the effect of chronic exposure to PAHs on mtDNAcn in peripheral blood lymphocytes (PBLs) of 46 Polish male noncurrent smoking coke-oven workers and 44 matched controls, who were part of a group of 94 study individuals examined in our previous work. Subjects' PAH exposure and genetic alterations were characterized through measures of internal dose (urinary 1-pyrenol), target dose [anti-benzo[a]pyrene diolepoxide (anti-BPDE)-DNA adduct], genetic instability (micronuclei and telomere length), and DNA methylation (p53 promoter) in PBLs. mtDNAcn (MT/S) was measured using a validated real-time PCR method. RESULTS: Workers with PAH exposure above the median value (>3 µmol 1-pyrenol/mol creatinine) showed higher mtDNAcn [geometric means (GM) of 1.06 (unadjusted) and 1.07 (age-adjusted)] compared with controls [GM 0.89 (unadjusted); 0.89 (age-adjusted); (P = 0.029 and 0.016)], as well as higher levels of genetic and chromosomal [i.e., anti-BPDE-DNA adducts (P < 0.001), micronuclei (P < 0.001), and telomere length (P = 0.053)] and epigenetic [i.e., p53 gene-specific promoter methylation (P < 0.001)] alterations in the nDNA. In the whole study population, unadjusted and age-adjusted mtDNAcn was positively correlated with 1-pyrenol (P = 0.043 and 0.032) and anti-BPDE-DNA adducts (P = 0.046 and 0.049). CONCLUSIONS: PAH exposure and PAH-related nDNA genotoxicity are associated with increased mtDNAcn. IMPACT: The present study is suggestive of potential roles of mtDNAcn in PAH-induced carcinogenesis.