Analysis of glutathione and vitamin C effects on the benzenetriol-induced DNA damage in isolated human lymphocytes.

The alkaline single-cell gel electrophoresis (or Comet) assay was applied to evaluate the eventual DNA damage induced by the triphenolic metabolite of benzene, 1,2,4-benzenetriol (BT), in isolated human lymphocytes. Prior to BT treatment, ranging from 5 to 50 microM, a supplementation with glutathione (GSH, 350 microg/ml) was carried out to assess whether GSH may have a modulating effect on the Comet response. The effect of a fixed dose of BT was also evaluated in the presence of the exogenous antioxidant vitamin C (40 and 200 microM). Additionally, we investigated whether the polymorphism of glutathione S-transferase T1 (GSTT1) gene may affect the individual level of BT-induced DNA damage in vitro. For all donors included in the present study, BT produced a significant dose-response relationship. No clear effect of GSH preincubation was seen on the BT-induced response. On the contrary, a significant reduction of DNA damage was observed in the presence of vitamin C (at least at 200 microM). Although our data suggest some individual differences according to the GSTT1 genotype in the outcome of the Comet assay, a large number of individuals should be studied in further investigations to obtain reliable conclusions.

Identification of a single nucleotide polymorphism in the TATA box of the CYP2A6 gene: impairment of its promoter activity.

Human cytochrome P450 2A6 (CYP2A6) constitutes the major nicotine oxidase, and large interindividual differences are seen in the levels of this enzyme, to a great extent caused by the distribution of several different polymorphic gene variants mainly located in the open reading frame (ORF). In the present study, we report a common polymorphism located in the 5' flanking region of CYP2A6 affecting its expression. DHPLC analysis and complete sequence of the open reading frame of the gene from a Turkish individual revealed a -48T > G substitution disrupting the TATA box. Using dynamic allele-specific hybridization (DASH), genotyping of this novel variant (named CYP2A6*9) was carried out in 116 Swedish, 132 Turkish, and 102 Chinese subjects, and the allele frequencies were found to be 5.2, 7.2, and 15.7%, respectively. The significance of the polymorphism was investigated by the construction of luciferase reporter plasmids containing 135 or 500 bp of the 5'-upstream region of the gene transfected into human hepatoma B16A2 cells. The constructs carrying the -48T > G mutation were only expressed at about 50% of the wild-type alleles. It is concluded that the CYP2A6*9 allele might be one of the most common CYP2A6 variants in Caucasians that alters the levels of enzyme expression.

Evaluation of DNA damage by the Comet assay in shoe workers exposed to toluene and other organic solvents.

The alkaline single-cell gel electrophoresis (or Comet) assay was applied to evaluate DNA damage in cryopreserved peripheral blood mononuclear leukocytes from 34 female shoe workers exposed to organic solvents and a group of 19 non-exposed women. We also investigated whether the polymorphisms of glutathione S-transferase M1 (GSTM1) and T1 (GSTT1) genes affect individual level of DNA damage possibly induced by the solvent exposure. Chemical measurements of workplace air in the two factories studied showed that the workers were exposed to acetone, gasoline, and toluene in both factories and to ethylacetate and diisocyanate in one factory. In the exposed workers, the average level of blood hemoglobin was lower and that of urinary hippuric acid higher than in the non-exposed individuals. However, the occupational exposure to organic solvents did not affect the Comet values. Neither did age, smoking, or the GSTM1 genotype have any effect on the outcome of this assay. The low prevalence of the GSTT1-null genotype precluded conclusions on the influence of GSTT1 polymorphism.

Examination of various biomarkers measuring genotoxic endpoints from Barcelona airport personnel.

Three different biomarkers: sister-chromatid exchanges (SCE), micronuclei (MN), and the Comet assay, were used to evaluate different kinds of genetic damage in peripheral blood lymphocytes from 34 male workers at Barcelona airport, exposed to low levels of hydrocarbons and jet fuel derivatives. The control group consisted of 11 unexposed men. We also investigated the ras p21 protein levels in plasma, in order to evaluate whether the ras gene could serve as a suitable potential marker of carcinogenic pollution in occupationally exposed cohorts. SCE and MN analyses failed to detect any statistically significant increase in the airport workers when compared with the controls, and in fact, the frequency of binucleated cells with MN in the exposed group was significantly lower than that obtained in the control. However, slight but significant differences in the mean comet length and genetic damage index were observed between the exposed and control groups when using the Comet assay. There were no statistically significant differences between both groups in p21 plasma levels. Smoking was shown to affect significantly both SCE and high frequency cells (HFC) in the exposed group.

SCE analysis in peripheral blood lymphocytes of a group of filling station attendants.

Petroleum derivatives constitute a complex mixture of chemicals which contain known genotoxicants. Thus, chronic occupational exposure to such derivatives may be considered to possess genotoxic risk. Service station attendants are workers exposed to petroleum derivatives and, as consequence, biomonitoring studies designed to evaluate the genetic risk are required. Here we present the data on sister chromatid exchange (SCE) frequencies, proliferation rate index (PRI) and cells with high frequency of SCE (HFC) from a group of 42 attendant workers. In addition, data from a group of 45 unexposed (controls) are presented. The results obtained indicate that there are no significant differences in SCE values between the exposed workers when compared to the non-exposed individuals. The only difference is found when smoking habit is considered, the smokers having significant increases in the SCE frequency. There is a slight increase in the HFC frequency of the occupationally exposed group but the difference did not attain statistical significance.

No increase in micronuclei frequency in cultured blood lymphocytes from a group of filling station attendants.

Service station attendants are workers that are definitely exposed to petroleum derivatives. Taking into account that this exposure has been considered to possess genotoxic risk, here we present data on the biomonitoring of a group of 50 service station workers and 43 controls. Micronuclei (MN) from peripheral blood lymphocytes has been considered as the genetic endpoint to be studied and, in addition, data on the concentration of aromatic hydrocarbons at the workplace, urinary metabolites and differential white blood cell count have also been analysed. The results obtained indicate no significant differences between petrol station attendants and controls, when the effects of petrol exposure were investigated by differential white blood cell count and analysis of MN frequencies in phytohaemagglutinin-stimulated lymphocytes. Regarding the urinary metabolites, a significant increase in the phenol level was found in the exposed workers.