Levels Of Aromatic DNA Adducts Research Articles

Aromatic DNA adducts in lymphocytes of humans working at high and low traffic density areas

Aromatic DNA adduct levels were determined by the 32P-postlabelling assay in lymphocytes isolated from newspaper vendors working at urban high traffic areas ( n = 31) and suburban low traffic areas ( n = 22) in Milan, Italy. The DNA adduct levels ranged from 0.7 to 6.7 10 8 nucleotides, while most of them were between 1.0 and 3.0 10 8 nucleotides. No difference was found between the DNA adduct levels of the high-exposed group ( 2.2 10 8 ) and the low-exposed group ( 2.2 10 8 ). The heavy smokers ( n = 8) had 23% higher DNA adduct level ( 2.7 10 8 ) than the non-smokers ( n = 37, 2.2 10 8 ) ( P = 0.27), but no correlation was found between the adduct level and the number of cigarettes/day. Analysis of variance of the DNA adduct levels among the 14 pairs of individuals working at the same news-stands revealed little effect of the environmental air exposure on the DNA adduct level.

Correlation of DNA adducts in blood mononuclear cells with tobacco carcinogen-induced damage in human lung

The formation of carcinogen-DNA adducts within the respiratory epithelium is thought to be a critical factor in the induction of lung cancer from tobacco smoke. A reliable surrogate measure of carcinogen damage to the lung would be of great value in molecular epidemiological studies of cancer risk. The validity of measurements of DNA adducts formed from hydrophobic aromatic hydrocarbons in peripheral blood mononuclear cells (MNCs) was investigated by comparing the levels of aromatic DNA adducts detected in lung tissue from 31 lung cancer patients with those detected in MNCs from the same individuals using the 32P-postlabeling assay. The associations of smoking history and intake of dietary antioxidants with adduct levels also were assessed. Tissue-specific, as well as common DNA adducts were detected in lung and blood; total MNC adduct levels were highly correlated with total lung adducts. After smoking cessation, adduct levels appeared to decay in both tissues at similar rates. Multivariate analyses (Poisson regression modeling) indicated that dietary antioxidant intake (carotenoids, vitamin A, and retinol) modified the levels of aromatic DNA adducts in both the lungs and blood. Of all models tested, the optimal one for predicting lung adduct levels included the measure of blood MNC adduct levels only. Therefore, blood MNCs are a valid surrogate tissue for estimating the burden of DNA adducts in respiratory tissue in molecular epidemiological studies.

Analysis of mutation at the hprt locus in human T lymphocytes

Studies of mutation at the hypoxanthine phosphoribosyl transferase ( hrpt) locus in human T-cells have the potential to elucidate the molecular basis of in vivo mutagenesis. reveal exposure dependent changes in ther background frequency of mutation, and provide knowledge on individual sensitivity. Styrene exposed lamination workers in Bohemia showed a significantly higher frequency of hprt mutant cells than Swedish control populations studied simultaneously. In a study of 47 healthy, non-smoking male bus maintenance workers exposed to diesel exhausts, soot and oil, and 22 unexposed controls, a significant correlation ( P = 0.008) was obtained between the levels of aromatic DNA adducts and frequencies of fcprf-mutant T-cells. In the group of workers with the highest exposure, subjects with glutathione 5-transferase (GSTM1) deficiency showed significantly higher ( p < 0.05) frequency of hprt mutant T-cells than GSTM1-positive subjects. The highest adduct levels were found in subjects with the combined genotype of GSTM1 and NAT2 deficiency (GSTMl-negative slow acetylators). These results indicate that GSTM1 and NAT2 genotypes may play a role in determining the individual levels of hprt mutation and DNA adducts. Using PCR-based screening methods, hprt mutations have been classified in 462 T-cell clones from 43 subjects in this study population. Deletions were found in 3% of the mutants, coding errors in 81% and splice mutations in 17%. Transitions and transversions were equally common, and all types of base substitutions were detected.

Lack of DNA adduct formation in mice treated with benzene

The potential of benzene to produce DNA adducts in B6C3F1 mice was investigated by the nuclease P1-enhanced 32P-postlabeling assay. The mice were daily treated i.p. with 500 mg/kg benzene in olive oil for 4 days, and the liver, mammary gland, and bone marrow were collected 24 h after the last treatment. Thin-layer chromatograms obtained with treated-tissue DNA specimens were qualitatively identical to those from corresponding olive oil-treated (control) tissue DNA. Quantitative evaluations revealed that there was no treatment-related increase in radioactivity on the chromatograms at or near the locations where the major in vitro adducts of phenol, hydroquinone and benzoquinone migrated. Benzene treatment, however, resulted in a decrease in the levels of certain endogenous adducts, the biological significance of which is unknown. Our results indicate that benzene treatment does not produce detectable levels of aromatic DNA adducts in mouse tissues.

Formation of aromatic DNA adducts in white blood cells in relation to urinary excretion of 1-hydroxypyrene during consumption of grilled meat.

With the aim of studying the effect of oral exposure to polycyclic aromatic hydrocarbons (PAH) on human DNA-adduct formation in mononuclear cells and excretion of 1-hydroxypyrene in urine, we examined the effect of consumption of charcoal-broiled hamburgers. Hamburgers were grilled and samples were homogenized, saponified, extracted with hexane and analysed for PAH content by HPLC. The mean levels of benzo[a]pyrene and pyrene in the grilled hamburgers were 8.6 and 26.5 micrograms/kg respectively. Twenty one healthy non-smoking individuals consumed two hamburgers (170 g) per day for 5 days. 32P-Postlabelling analysis was performed on DNA samples of mononuclear cells of the subjects. The excretion of 1-hydroxypyrene in urine was studied as a marker of endogenous exposure to PAH. In the DNA samples of eight of the 21 subjects, on day 3 of the consumption period a predominant adduct spot could be detected with similar chromatographic properties to a benzo[a]pyrenediolepoxide--deoxyguanosine standard, the levels varying between 3 and 103 adducts/10(10) nucleotides. Analysis of the urine samples revealed maximal 1-hydroxypyrene excretion on day 3 in all nine subjects who collected urine daily during the consumption week, with an average level of 5.2 nmol/24 h. In a subsequent study in which six volunteers consumed charcoal-broiled hamburgers with lower levels of benzo[a]pyrene and pyrene, no aromatic DNA adducts in mononuclear cells or increased 1-hydroxypyrene levels in urine were detected. In conclusion, oral intake of PAH may dose-dependent induce elevated levels of aromatic DNA adducts in mononuclear cells and of 1-hydroxypyrene in urine, indicating substantial bioactivation of PAH, in particular via this route.

DNA adducts among personnel servicing and loading diesel vehicles.

The levels of aromatic DNA adducts were compared by the 32P-postlabeling assay between the lymphocytes isolated from bus maintenance and truck terminal workers, using hospital mechanics as a control group. The adduct levels were elevated in all the bus and truck terminal workers. Within the bus maintenance personnel, garage workers had the highest levels of adducts. Within the terminal workers those driving diesel forklifts had the highest adduct levels. The results suggest that diesel exhaust contributes to the level of adducts.

Seasonal variation of aromatic DNA adducts in human lymphocytes and granulocytes.

DNA adducts were measured by 32P-postlabelling in lymphocytes and granulocytes of 75 healthy men exposed occupationally and environmentally to high concentrations of aromatic compounds in the ambient air. Volunteers enrolled in the study were men working at the coke batteries and nonoccupationally exposed inhabitants of Silesia, a highly industrialized region in southern Poland. Blood samples were drawn twice: in February and September 1992. Seasonal variations in the levels of DNA adducts were found only in lymphocytes: 3.6- and 8.7-fold in the occupationally and environmentally exposed groups respectively. In smokers the seasonal variation was as large as 12.8-fold in the environmentally exposed group. No seasonal variations were observed in granulocytes. The observed seasonal variation in the level of aromatic DNA adducts coincided with winter/summer differences in the concentrations of benzo[a]pyrene in the ambient air of Silesia. The study recommends the application of lymphocytes for adduct studies in chronic exposures and provides evidence on the repair of aromatic adducts in lymphocytes.

32P-postlabelling analysis of DNA from human tissues

DNA extracted from human lung, bladder, liver, pancreas, cervix and breast tissue samples taken at autopsy (37 sample sets) was analysed by the nuclease P1 enhancement modification of the 32P-postlabelling assay for levels of aromatic carcinogen DNA adducts. Results were combined with those from a previous study for statistical analysis of 56 sample set (32 male + 24 female). A strong trend was seen for increased adduct levels in the lung DNA of smokers and a weak association for the bladder DNA of smokers compared to non-smokers. Aromatic adducts were also detected in other tissues.

Aromatic DNA-carcinogen adducts in beluga whales from the Canadian Arctic and Gulf of St Lawrence

Aromatic DNA adduct levels in liver tissues of beluga whales ( Delphinapterus leucas) from two sites in the Canadian Arctic and from the St Lawrence estuary (N.W. Atlantic Ocean) were determined using the highly sensitive 32P-postlabelling technique. Detectable levels of aromatic DNA adducts (16–158 nmole adducts/mole total nucleotides) were found in all animals tested. It is of interest that the average level of adducts in animals from the St Lawrence estuary, which contains relatively high levels of contaminants including polycyclic aromatic hydrocarbons, was comparable to those found in animals from the two remote locations. These observations are of major significance in view of the controversy on the role of polycyclic aromatic hydrocarbons in producing cancer in beluga whales in the Gulf of St Lawrence. (The point made is that PAH may not be the primary reason and other causes should be addressed.) Also given that the study was carried out on animals taken from regions known to be relatively clean and highly contaminated, the results are of major importance for the scientific and regulatory communities with interests in the use of biochemical monitoring techniques. Also, the presence of adducts in whales from the Canadian Arctic suggests the existence of non pollution-related mechanisms for genetic damage.

Carcinogenic and genotoxic activity of extracts from contaminated sediments in Western Lake Ontario

In order to investigate the putative association between chemical contamination in western Lake Ontario and high prevalences of fish tumors, sediments from Hamilton Harbour and Oakville Creek in Lake Ontario and reference sites in non-polluted areas of Ontario, Canada were collected and extracted for organic contaminants. Sediment extracts from Hamilton Harbour had the highest concentrations of polychlorinated biphenyls and organochlorine insecticides (ppb) and contained very high concentrations of polynuclear aromatic hydrocarbons (ppm); although the levels of these compounds varied widely with sampling location in the harbor. A sediment extract from Hamilton Harbour was mutagenic in the Ames bacterial assay, both with and without microsomal activation. High levels of aromatic DNA adducts were induced in cultured mouse C3H1OT1/2 cells after in vitro exposure to Hamilton Harbour sediment extract. In two separate carcinogenicity experiments involving a sac fry microinjection assay with rainbow trout (Oncorhynchus mykiss), Hamilton Harbour sediment extract induced hepatocellular carcinomas in fish. No hepatic neoplasms were observed in fish that had been treated with sediment extract from Oakville Creek, or with extract from a reference sediment. The significance of these results is discussed in relation to the distribution of neoplasms in feral fish within western Lake Ontario.

32P‐assay of DNA adducts in white blood cells and placentas of pregnant women: Lack of residential wood combustion‐related adducts but presence of tissue‐specific endogenous adducts

Residential wood combustion (RWC), which has been increasingly used as a heating source, is of health concern because emissions from RWC contain carcinogenic polycyclic aromatic hydrocarbons (PAH). To assess health risk, the possible formation of PAH-DNA adducts in white blood cells (WBC) and placentas of nonsmoking women exposed to RWC smoke during pregnancy was measured by a nuclease P1-enhanced 32P-postlabeling assay having a sensitivity limit of one lesion per 10(9-10) DNA nucleotides. DNA samples isolated from 12 exposed specimens (8 WBC, 4 placentas) and 13 unexposed control specimens (8 WBC, 5 placentas) were hydrolyzed to mononucleotides, which were then 32P-labeled and separated by high-resolution thin-layer chromatography. Comparison of autoradiograms of exposed DNA samples with those of controls failed to show exposure-related adducts. All placental DNA maps exhibited one major (47 +/- 10%) and 12 minor extra spots, however, that were not seen on WBC-DNA maps. These derivatives corresponded to an average of 12 (+/- 6) modifications in 10(9) nucleotides. Similarly, WBC DNA showed four spots that were absent in placental DNA and were not quantified because of their low levels. Neither placental nor WBC DNA adducts coincided chromatographically with the product formed by the reaction of benzo(a)pyrene diol epoxide I with N2 of guanine in DNA. Although these results suggest that RWC smoke does not elicit detectable levels of aromatic DNA adducts in humans, lack of evaluation of RWC exposure levels leaves some uncertainty in this conclusion. The results, however, clearly show that placental and WBC DNA contain covalent modifications that are unrelated to RWC exposure. These DNA derivatives may be caused by ingestion and/or inhalation of, or skin contact with, low levels of environmental genotoxicants or may arise from endogenous electrophiles of as yet unknown origin.