The utility of epithelial-cell micronuclei in the assessment of intermittent exposures

Epithelial-cell micronuclei (MN) are potentially useful markers of occupational exposure to genotoxicants. With intermittent exposures, cells sampled either before or after a specific time interval, reflecting the time it takes for damaged cells to become available at the epithelial surface, are unlikely to be exposure-related. It may then be important to conduct an exposure-window analysis, with the goal of identifying the relevant exposures.We re-analysed individual exposure data from a previous study (Suruda et al. 1993) of MN formation in 22 male mortuary science students exposed to formaldehyde during a 90-day embalming class. We conducted an exposurewindow analysis and compared the results with those obtained with 90-day cumulative exposure. The window widths varied between 7 and 25 days, in 1 day increments, assuming a constant 7-day cell-cycle. We assessed the fit (likelihood-ratio test) of a linear regression model, regressing the change in buccal MN prevalence on formaldehyde exposure, using both asymptotic and non-asymptotic methods. Exposures defined from 7-15 to 7-18 days before specimen collection provided a slightly better fit than the 90-day cumulative exposure, with a doubling of the regression coefficient for the exposure effect (for the 7-16-days window LR = 5.32, p = 0.032, coefficient = 0.088 MN per 1000 cells per ppm-hr; 95% CI = 0.014, 0.16; for the 90-day cumulative exposure LR = 4.44, p = 0.048, coefficient = 0.045 MN per 1000 cells per ppm-hr, 95% CI = 0.0038, 0.086). Although hampered by the small number of subjects, these results reinforce the potential importance of exposure timing.     

Assessment of local genotoxic effects of formaldehyde in humans measured by the micronucleus test with exfoliated buccal mucosa cells

Volunteers (10 women, 11 men) were exposed to formaldehyde (FA) vapors for 4h per day over a period of 10 working days under strictly controlled conditions. Exposure varied randomly each day from constant 0.15 ppm up to 0.5 ppm with four peaks of 1.0 ppm for 15 min each (13.5 ppm h cumulative exposure over 10 working days). FA was masked on four days by co-exposure to ethyl acetate. During exposure, subjects had to perform bicycle exercises (about 80 W) three times for 15 min. Buccal smears were prepared 1 week before the start of the study (control 1), at the start of the study before the first exposure (control 2), at the end of the exposure period of 10 days and 7, 14 and 21 days thereafter. Two thousand cells per data point were analyzed for the presence of micronuclei (MN) and the frequency of MN per 1000 cells was determined on slides coded by an independent quality-assurance unit. No significant increase in the frequency of MN was measured at any time point after the end of the exposure. Twenty-one days after the end of the exposure MN frequencies were significantly lower in comparison with control 1. This study, which was performed under GLP-like conditions, clearly indicates that FA does not induce MN in buccal mucosa cells after peak exposures up to 1 ppm and a cumulative exposure of 13.5 ppm h over 2 weeks.     

Validation of micronuclei frequency in peripheral blood lymphocytes as early cancer risk biomarker in a nested case-control study

Aim of this work was to assess the predictive value of micronuclei (MN) frequency in peripheral blood lymphocytes (PBL) for the risk of cancer death in disease-free individuals. Blood samples from 1650 subjects selected from the general population of Pisa, Italy, were collected between June 1991 and November 1993. The follow-up until January 2005 recorded a total of 111 deaths (52 for cancer). MN frequency was assessed for 49 cancer cases and 101 matched controls. A significantly higher MN frequency was found in cancer cases (4.7+/-3.4 MN/1000 BN cells) versus controls (1.5+/-1.7; p<0.0001). Donors were stratified in two classes and multivariate logistic regression analysis confirmed that individuals with high MN frequency (>2.5 MN/1000 BN cells) had a significantly increased risk of cancer death (OR=10.7; 95% CI=4.6-24.9; p<0.0001) when compared to individuals with low MN frequency (相似文献   

Re-evaluation of a reported increased micronucleus frequency in lymphocytes of workers occupationally exposed to formaldehyde

A replicate evaluation of increased micronucleus (MN) frequencies in peripheral lymphocytes of workers occupationally exposed to formaldehyde (FA) was undertaken to verify the observed effect and to determine scoring variability. May-Grünwald-Giemsa-stained slides were obtained from a previously performed cytokinesis-block micronucleus test (CBMNT) with 56 workers in anatomy and pathology laboratories and 85 controls. The first evaluation by one scorer (scorer 1) had led to a highly significant difference between workers and controls (3.96 vs 0.81 MN per 1000 cells). The slides were coded before re-evaluation and the code was broken after the complete re-evaluation of the study. A total of 1000 binucleated cells (BNC) were analysed per subject and the frequency of MN (in ‰) was determined. Slides were distributed equally and randomly between two scorers, so that the scorers had no knowledge of the exposure status. Scorer 2 (32 exposed, 36 controls) measured increased MN frequencies in exposed workers (9.88 vs 6.81). Statistical analysis with the two-sample Wilcoxon test indicated that this difference was not significant (p=0.17). Scorer 3 (20 exposed, 46 controls) obtained a similar result, but slightly higher values for the comparison of exposed and controls (19.0 vs 12.89; p=0.089). Combining the results of the two scorers (13.38 vs 10.22), a significant difference between exposed and controls (p=0.028) was obtained when the stratified Wilcoxon test with the scorers as strata was applied. Interestingly, the re-evaluation of the slides led to clearly higher MN frequencies for exposed and controls compared with the first evaluation. Bland-Altman plots indicated that the agreement between the measurements of the different scorers was very poor, as shown by mean differences of 5.9 between scorer 1 and scorer 2 and 13.0 between scorer 1 and scorer 3. Calculation of the intra-class correlation coefficient (ICC) revealed that all scorer comparisons in this study were far from acceptable for the reliability of this assay. Possible implications for the use of the CBMNT in human biomonitoring studies are discussed.     

Cytogenetic effects in rotogravure printers exposed to toluene (and benzene).

Comparing 21 rotogravure printers exposed to toluene (medians: time-weighted air level 150 mg/m3, blood toluene 1.6 mumole/l) and 21 unexposed controls (median blood toluene less than or equal to 0.01 mumole/l) there was a significant increase in the frequency of micronuclei (MN) in pokeweed mitogen (PWM)-stimulated peripheral blood lymphocytes in the printers, as compared to the controls (2.8% vs. 1.5%; p = 0.03; all p adjusted for age and smoking). The frequency of small MN (size ratio MN/main nucleus less than or equal to 0.03) in PWM-stimulated lymphocytes was associated with the exposure (1% vs. 0.3%; p = 0.05). Furthermore, among the exposed subjects there was an association between blood toluene and small MN (0.17% per mumole/l; p = 0.0005). Small MN in phytohemagglutinin (PHA) cultures displayed no association with any exposure parameter. However, in the printers, an estimated cumulative exposure index was weakly correlated with the frequency of total MN in PHA-stimulated cells (0.00003% per mg/m3 x year; p = 0.07). Among the printers, chromosomal breaks in PHA-stimulated cells were associated with the duration of earlier benzene exposure (0.03% per year; p = 0.01). The results of this study strongly indicate that toluene causes a clastogenic effect on the B-cells even at low exposure levels. Further, earlier benzene exposure seems to have caused chromosomal breaks in T-cells.     

Cytogenetic analysis of nasal mucosa cells and lymphocytes from high-level long-term formaldehyde exposed workers and low-level short-term exposed waiters

The evidence for genotoxic potential of formaldehyde (FA) in humans is insufficient and conflicting. We previously reported a higher frequency of micronuclei in nasal and oral exfoliative cells from students exposed to formaldehyde vapor for short-term. To further evaluate the genetic effects of long-term occupational exposure to FA and short-term exposure to FA of indoor sources, the frequencies of micronuclei (MN) in nasal mucosa cells, sister chromatid exchanges (SCEs) of peripheral lymphocytes, and the lymphocyte subsets were evaluated in 18 non-smoking workers (mean exposure duration was 8.6 years) in an FA factory and 16 non-smoking waiters exposed to FA for 12 weeks in a ballroom. A non-smoking student group without occupational exposure (n=23) to FA was used as control. The 8h time-weighted average (TWA) concentrations of formaldehyde was 0.985+/-0.286 mg/m3 with the ceiling exposure concentration of 1.694 mg/m3 in the workshop, and 0.107+/-0.067 mg/m3 in the ballroom (5 h TWA). Higher frequencies of micronuclei per thousand cells in nasal mucosa cells of workers versus control (2.70+/-1.50 versus 1.25+/-0.65, p<0.05) and higher frequency of SCEs in peripheral lymphocytes of workers group (8.24+/-0.89 versus 6.38+/-0.41, p<0.05) were observed. Increased frequency of micronuclei in nasal mucosa cells or SCE in peripheral lymphocytes was not found among waiters group. The results suggest that the genotoxic potential of high level FA exposure may have occupational risks in long-term exposure groups.     

Effect of smoking habit on the frequency of micronuclei in human lymphocytes: results from the Human MicroNucleus project

The effect of tobacco smoking on the frequency of micronuclei (MN) in human lymphocytes has been the object of many population studies. In most reports, the results were unexpectedly negative, and in many instances smokers had lower frequencies of MN than non-smokers. A pooled re-analysis of 24 databases from the HUMN international collaborative project has been performed with the aim of understanding the impact of smoking habits on MN frequency. The complete database included 5710 subjects, with 3501 non-smokers, 1409 current smokers, and 800 former smokers, among subjects in occupational and environmental surveys. The overall result of the re-analysis confirmed the small decrease of MN frequencies in current smokers (frequency ratio (FR) = 0.97, 95% confidence interval (CI) = 0.93-1.01) and in former smokers (FR = 0.96, 95% CI = 0.91-1.01), when compared to non-smokers. MN frequency was not influenced by the number of cigarettes smoked per day among subjects occupationally exposed to genotoxic agents, whereas a typical U-shaped curve is observed for non-exposed smokers, showing a significant increase of MN frequency in individuals smoking 30 cigarettes or more per day (FR = 1.59, 95% CI = 1.35-1.88). This analysis confirmed that smokers do not experience an overall increase in MN frequency, although when the interaction with occupational exposure is taken into account, heavy smokers were the only group showing a significant increase in genotoxic damage as measured by the micronucleus assay in lymphocytes. From these results some general recommendations for the design of biomonitoring studies involving smokers can be formulated. Quantitative data about smoking habit should always be collected because, in the absence of such data, the simple comparison of smokers versus non-smokers could be misleading. The sub-group of heavy smokers (> or =30 cigarettes per day) should be specifically evaluated whenever it is large enough to satisfy statistical requirements. The presence of an interaction between smoking habit and occupational exposure to genotoxic agents should be always tested.     

Ginseng reduces the micronuclei yield in lymphocytes after irradiation

To assess the effect of Chinese ginseng in modifying the radiation-induced micronuclei (MN) yield in human G(o) peripheral blood lymphocytes (PBL), we conducted the cytokinesis-blocked (CB) MN assay in blood samples obtained from healthy volunteers (n=4). Before (137)Cs ex vivo irradiation, mononuclear cell cultures from each sample were incubated 24 h with different concentrations (0-2000 microg ml(-1)) of crude water extract of ginseng dry root. We found that (1) at 0 Gy and without the presence of ginseng, MN yield (mean+/-S.E.M.) was 11.7+/-2.7 per 1000 binucleated (BN) cells. Different concentrations of ginseng crude water extract did not affect the MN yields and the proliferative activity of PBL; (2) after 1 and 2 Gy exposure, radiation alone sharply increased the MN yields, respectively, to 119.6+/-17.4 and 340.5+/-20.9 per 1000 BN cells. However, treatment with ginseng for 24 h before radiation exposure, resulted in a significant linear decline of MN yields as ginseng concentration increases. Compared to radiation alone, the extent to which ginseng water extract reduced the MN yields induced by 1 Gy exposure was 46.0% at 1500 microg ml(-1) and 61.5% at 2000 microg ml(-1), and with 2 Gy exposure, it was 38.6% at 1500 microg ml(-1) and 46.5% at 2000 microg ml(-1); (3) MN data suggested a tendency for overdispersion relative to the Poisson model; and (4) over the different levels of ginseng concentrations, the trend in micronucleated BN index was as similar as that of the MN yields. These results indicated that (1) ginseng crude water extract exerts no apparent cytogentic effect on human PBL at concentrations up to 2000 microg ml(-1) as evaluated by the CBMN assay; and (2) the protection of ginseng water extract against (137)Cs-induced MN in human PBL is concentration-dependence. Therefore, our findings indicated that ginseng may have therapeutic value as a possible radioprotector for normal tissue during radiotherapy of cancer patients.     

Genotoxic risk assessment of pathology and anatomy laboratory workers exposed to formaldehyde by use of personal air sampling and analysis of DNA damage in peripheral lymphocytes

A study was conducted to evaluate the genotoxic effect of occupational exposure to formaldehyde on pathology and anatomy laboratory workers. The level of exposure to formaldehyde was determined by use of passive air-monitoring badges clipped near the breathing zone of 59 workers for a total sampling time of 15min or 8h. To estimate DNA damage, a chemiluminescence microplate assay was performed on 57 workers before and after a 1-day exposure. Assessment of chromosomal damage was carried out by use of the cytokinesis-blocked micronucleus assay (CBMN) in peripheral lymphocytes of 59 exposed subjects in comparison with 37 controls matched for gender, age, and smoking habits. The CBMN assay was combined with fluorescent in situ hybridization with a pan-centromeric DNA probe in 18 exposed subjects and 18 control subjects randomized from the initial populations. Mean concentrations of formaldehyde were 2.0 (range <0.1-20.4ppm) and 0.1ppm (range <0.1-0.7ppm) for the sampling times of 15min and 8h, respectively. No increase in DNA damage was detected in lymphocytes after a one-workday exposure. However, the frequency of binucleated micronucleated cells was significantly higher in pathologists/anatomists than in controls (16.9 per thousand+/-9.3 versus 11.1 per thousand+/-6.0, P=0.001). The frequency of centromeric micronuclei was higher in exposed subjects than in controls (17.3 per thousand+/-11.5 versus 10.3 per thousand+/-7.1) but the difference was not significant. The frequency of monocentromeric micronuclei was significantly higher in exposed subjects than in controls (11.0 per thousand+/-6.2 versus 3.1 per thousand+/-2.4, P<0.001), while that of the acentromeric micronuclei was similar in exposed subjects and controls (3.7 per thousand+/-4.2 and 4.1 per thousand+/-2.7, respectively). The enhanced chromosomal damage (particularly chromosome loss) in peripheral lymphocytes of pathologists/anatomists emphasizes the need to develop safety programs.     

Induction of micronuclei in wild-type and p53(+/-) transgenic mice by inhaled bromodichloromethane

Bromodichloromethane (BDCM) is commonly present in trace amounts in drinking water as a disinfection by-product. BDCM has been shown to be carcinogenic in mice and rats when given by gavage at relatively high doses. Genotoxic activity as well as induced regenerative cell proliferation may contribute to the carcinogenic potential of BDCM. The purpose of the current studies was to evaluate the ability of BDCM to induce micronuclei (MN) in bone marrow and blood of wild-type and p53(+/-) mice on the C57BL/6 and FVB/N genetic backgrounds using the inhalation route of exposure. Toxicity studies were being conducted in this laboratory with inhaled BDCM to select doses for longer-term cancer bioassays using wild-type and p53(+/-) transgenic mice on different genetic backgrounds. Bone marrow samples from these experiments were evaluated for the induction of MN after 1 and 3 weeks of exposure. Accumulation of MN in the peripheral blood was also evaluated at the 13-week time point of a cancer study with the p53(+/-) mice. For the 1-week time point, male C57BL/6 wild-type and p53(+/-) mice and FVB/N wild-type and p53(+/-) mice were exposed daily for 6h per day for 7 consecutive days to atmospheric BDCM concentrations of 0, 1, 10, 30, 100, or 150 ppm. In a second experiment, mice were exposed daily for 6h per day for 3 weeks to atmospheric BDCM concentrations of 0, 0.5, 1, 3, 10, or 30 ppm. Resulting levels of polychromatic erythrocytes (PCE) containing MN were assessed in the bone marrow. For all of the 1- and 3-week exposure groups, the only statistically significant increase in the percentage of bone marrow PCE cells containing MN was in the 1-week 100 ppm BDCM exposure group in the FVB/N wild-type mice (control 0.26% versus exposed 1.16%). C57BL/6 p53(+/-) mice and FVB/N p53(+/-) mice were exposed daily for 6 h per day for 13 weeks to atmospheric BDCM concentrations of 0, 0.5, 3, 10, or 15 ppm. MN were quantified in samples of peripheral blood. Statistically significant increases in the percentage of peripheral blood NCE cells containing MN were seen at the highest BDCM exposure group of 15 ppm in both the C57BL/6 p53(+/-) strain (control 0.36% versus exposed 0.67%) and the FVB/N p53(+/-) strain (control 0.36% versus exposed 0.86%). These data indicate weak induction of MN by BDCM, but only at high atmospheric concentrations relative to normal environmental exposures and with extended periods of exposure. Although comparisons are difficult because responses were negative or marginal, the p53 genotype or the genetic background did not appear to substantially alter susceptibility to the genotoxic effects of BDCM.     

Cytogenetic analysis of buccal cells from shoe-workers and pathology and anatomy laboratory workers exposed to n-hexane, toluene, methyl ethyl ketone and formaldehyde.

People employed in the shoe manufacture and repair industry are at an increased risk for cancer, the strongest evidence being for nasal cancer and leukaemia. A possible causal role for formaldehyde is likely for cancer of the buccal cavity and nasopharynx. Exfoliated buccal cells are good source of tissue for monitoring human exposure to inhaled and ingested occupational and environmental genotoxicants. To assess the cytogenetic damage related to occupational exposure to airborne chemicals during shoe-making and the processes in pathology and anatomy laboratories, the micronuclei (MN) count per 3000 cells was measured in buccal smears from shoe-workers (group I, n = 22) exposed to mainly n-hexane, toluene and methyl ethyl ketone (MEK) and from anatomy and pathology staff (group II, n = 28) exposed to formaldehyde (FA). Eighteen male university staff were used as controls. The mean time-weighted average (TWA) concentrations of n-hexane, toluene and MEK in 10 small shoe workshops were 58.07 p.p.m., 26.62 p.p.m. and 11.39 p.p.m., respectively. The measured air concentrations of FA in the breathing zone of the anatomy and pathology laboratory workers were between 2 and 4 p.p.m. Levels of 2,5-hexadione (2,5-HD) and hippuric acid (HA), metabolic markers of n-hexane and toluene exposure, respectively, were significantly higher in the urine of workers in group I than in control subjects (p < 0.001 and p < 0.01, respectively). The mean (+/- SD) MN (0/00) [corrected] frequencies in buccal mucosa cells from workers in group I, group II and controls were 0.62 +/- 0.45%, 0.71 +/- 0.56% and 0.33 +/- 0.30%, respectively (p < 0.05 and p < 0.05 compared with controls for group I and group II, respectively). The effects of smoking, age and duration of exposure on the frequency of micronucleated buccal cells from workers in all three groups studied were also evaluated. Overall, the results suggest that occupational exposure to organic solvents, mainly n-hexane, toluene, MEK and FA, may cause cytogenetic damage in buccal cells and that use of exfoliated buccal cells seems to be appropriate to measure exposure to organic solvents.     

A combination of the micronucleus assay and a FISH technique for evaluation of the genotoxicity of 1,2,4-benzenetriol

The cytokinesis-block micronucleus (CBMN) assay has emerged as one of the preferred methods for assessing chromosome damage. Micronuclei (MN) are small, extranuclear bodies that are formed in mitosis from acentric chromosomal fragments or chromosomes that are not included in each daughter nucleus. Thus, MN contain either chromosomal fragments or whole chromosomes. The CBMN assay, together with a fluorescence in situ hybridization (FISH) technique using specific centromeric probes for chromosomes 7 and 8, were employed in mitogen-stimulated human lymphocytes pretreated with the benzene metabolite, 1,2,4-benzenetriol (BT). Treatment of human lymphocytes resulted in the induction of MN in a dose-dependent manner. The frequency of MN in control lymphocytes was 4.5 per 1000 binucleated (BN) cells and this increased to 9.5, 14, 28 and 40 per 1000 BN cells at 10, 25, 50 and 100 microM BT, respectively. The frequency of aneuploidy 7 and 8 in BN cells also increased at each concentration. Aneuploidy 8 was more frequent than aneuploidy 7, suggesting that chromosome 8 is more sensitive to aneuploidy induction by BT. The frequency of MN containing centromere positive signals for chromosomes 7 and 8 increased with the concentration of BT. The frequency of MN with centromere positive signals was higher for chromosome 8 than for chromosome 7, also suggesting a greater sensitivity of chromosome 8 to this agent. These results suggest that combined application of the CBMN assay with a FISH technique, using chromosome-specific centromeric probes, would allow the detection of aneuploidy in human lymphocytes and identify the mechanistic origin of MN induced by a clastogen or aneugen.     

Protective effect of zinc on cadmium-induced micronuclei in V79 cells

The induction of micronuclei (MN) in mitotically active cells has been widely used and promoted as a biological marker of exposure to environmental toxins. In our study the effect of zinc on cadmium genotoxicity was investigated in V 79 cells. The results indicate that cadmium chloride exposure for 24 h increased micronucleus frequency and the percentage of binucleated cells in dose-dependent manner. At the highest concentration of cadmium (50 microM Cd) 23 MN were found in 1000 cells. The protective effect of zinc on cadmium genotoxicity was investigated at lower concentrations (5-25 microM CdCl2). At 50 microM Cd, the number of MN increased significantly (16 MN).     

A comparative biomonitoring study of populations residing in regions with low and high risk of lung cancer using the chromosome aberration and the micronucleus tests

Chromosome aberration (CA) and micronucleus (MN) tests were performed in peripheral blood lymphocytes from people residing in two districts of Chiang Mai, Thailand, a high-risk area, Saraphi (n=107), where the lung cancer incidence is three-fold higher than in a low-risk area, Chom Thong (n=118). The percentage of cells with CAs was significantly lower in the Saraphi population than in the Chom Thong population (0.47+/-0.91 versus 1.04+/-1.18, P=0.0001) as was the percentage of CAs (0.49+/-0.91 versus 1.08+/-1.21, P<0.0001) and the mitotic indices (1.25+/-0.44 versus 1.33+/-0.33, P=0.025). The frequency of MN in binucleated (BN) cells, however, was significantly higher in the Saraphi population (12.01+/-3.57 versus 9.99+/-3.11, P<0.0001) as was the percentage of BN cells with MN (1.14+/-0.31 versus 0.93+/-0.23, P<0.0001). There was no difference in the nuclear division indices (1.49+/-0.07 versus 1.47+/-0.11, P=0.1759) between the two populations. With regard to the effect of confounding factors, it was found that cigarette smoking influenced both CA and MN frequencies, and that the chewing of fermented tea leaves or betel nuts affected CA and sex affected MN frequencies. An increasing of CA and MN frequencies were seen in smokers and chewers over non-smokers and non-chewers, with CA frequencies being higher in Chom Thong smokers and chewers and MN frequency being higher in Saraphi smokers. However, pesticide exposure and alcohol consumption had no impact on CA and MN frequencies. Due to the conflicting results obtained in the two tests, we cannot make a clear statement regarding the potential effects of the environmental exposures in the two study populations.     

Analysis of erythrocyte glycophorin-A variants by flow cytometry in lung disease patients detects the effect of tobacco smoke.

The glycophoryn A (GPA) assay evaluates somatic in vivo mutations. It is considered a cumulative biodosimeter for genotoxic exposures and is under evaluation in cancer risk assessment. GPA, a polymorphic membrane protein of the erythrocytes, determines the MN blood groups. The NO and NN variant frequencies (VF) may be detected in MN subjects (about 50% of the population) by flow cytometry using two differently labelled antibodies. We explored if GPA NO and NN VF might be relevant to the assessment of individual lung cancer risk and susceptibility, in a small population with a high prevalence of heavy tobacco smokers: 8 lung cancer patients and 16 subjects with non-malignant lung diseases associated with increased risk of lung cancer. There was a wide interindividual variability and complete overlap between non-neoplastic and neoplastic patients. A significant positive correlation was seen with smoking duration in NO VF (p = 0.04, age-adjusted). Current smokers (n = 12) displayed higher NO values than never (n = 1) or ex-smokers (n = 11), 36.3 +/- 18.2 and 21.0 +/- 13.2, respectively (p < 0.01). No association was shown with occupational exposure. The present exploratory study suggests that assessment of individual lung cancer risk and susceptibility by the GPA assay does not seem to be feasible. The assay appears to provide a biomarker of longterm exposure to tobacco smoke.     

Study of the cytogenetic effects of occupational exposure to pesticides on sanitation workers in Belo Horizonte, Brazil

Sanitation workers handling pesticides in the control of disease vectors constitute an occupationally exposed population to genotoxic substances. The aim of the present study was to investigate the relation between the occupational exposure to various pesticides and the presence of cytogenetic damage. Fifty-nine men were selected (29 sanitation workers and 30 control individuals) with ages varying between 18-57 years who lived and worked in the same area in Belo Horizonte (Brazil). The following parameters were determined for all individuals using the cytokinesis-block micronucleus (MN) assay in peripheral blood lymphocytes: MN/1000 binucleated cells (BC), BC with MN (BCMN)/1000 BC, nucleoplasmic bridges (NB)/1000 BC, apoptotic and necrotic cells/500 cells and nuclear division index. The analysis of covariance showed significantly higher (p < 0.05) mean frequencies of MN (15.81 +/- 1.31 vs 4.71 +/- 0.42), BCMN (15.10 +/- 1.22 vs 4.62 +/- 0.44), NB (4.59 +/- 0.76 vs 1.00 +/- 0.34), and necrotic cells (12.07 +/- 1.45 vs 5.17 +/- 0.70) in the exposed group when compared to the control group. There was no significant difference in the apoptotic cell frequency between the two groups, while the nuclear division index was significantly lower (1.49 +/- 0.02 vs 1.61 +/- 0.02) in the control group. Neither the time of exposure nor the smoking or alcohol drinking habit influenced the cytogenetic parameters examined. According to these results, occupational exposure to pesticides induced genotoxic and cytotoxic effects in sanitation workers.     

Thyroid cancer risk in areas of Ukraine and Belarus affected by the Chernobyl accident

The purpose of the present study was to analyze the thyroid cancer incidence risk after the Chernobyl accident and its degree of dependence on time and age. Data were analyzed for 1034 settlements in Ukraine and Belarus, in which more than 10 measurements of the (131)I content in human thyroids had been performed in May/June 1986. Thyroid doses due to the Chernobyl accident were assessed for the birth years 1968-1985 and related to thyroid cancers that were surgically removed during the period 1990-2001. The central estimate for the linear coefficient of the EAR dose response was 2.66 (95% CI: 2.19; 3.13) cases per 10(4) PY-Gy; for the quadratic coefficient, it was -0.145 (95% CI: -0.171; -0.119) cases per 10(4) PY-Gy(2). The EAR was found to be higher for females than for males by a factor of 1.4. It decreased with age at exposure and increased with age attained. The central estimate for the linear coefficient of the ERR dose response was 18.9 (95% CI: 11.1; 26.7) Gy(-1); for the quadratic coefficient, it was -1.03 (95% CI: -1.46; -0.60) Gy(-2). The ERR was found to be smaller for females than for males by a factor of 3.8 and decreased strongly with age at exposure. Both EAR and ERR were higher in the Belarusian settlements than in the Ukrainian settlements. In contrast to ERR, EAR increases with time after exposure. At the end of the observation period, excess risk estimates were found to be close to those observed in a major pooled analysis of seven studies of childhood thyroid cancer after external exposures.     

Analysis of micronuclei, histopathological changes and cell proliferation in nasal epithelium cells of rats after exposure to formaldehyde by inhalation

The frequencies of micronuclei (MN), histopathological changes and cell proliferation were determined in the nasal epithelium of male Fischer-344 rats after exposure to formaldehyde (FA) by whole-body inhalation for four weeks (6h/day, 5 days/week). Groups of 12 rats each were exposed to the target concentrations of 0, 0.5, 1, 2, 6, 10 and 15ppm. The micronucleus test (MNT) was performed with nasal epithelial cells prepared from six animals per group. Two thousand cells per animal were analysed for the presence of MN. The other six rats per group were subcutaneously implanted with osmotic pumps containing 5-bromo-2'-deoxyuridine (BrdUrd), three days prior to necropsy. Paraffin sections were made from the nasal cavity (four levels) of these animals for histopathology and cell-proliferation measurements. The frequency of cells with MN was not increased in any of the groups. However, there was also no induction of MN in nasal cells of rats exposed to a single dose of cyclophosphamide (CP, 20mg/kg) by gavage and analysed 3, 7, 14 or 28 days after the treatment. In contrast, nasal epithelial cells from rats exposed to 10 or 15ppm FA vapour showed clear site-specific pathological changes (focal epithelial degeneration, inflammation and squamous metaplasia) in a decreasing gradient (anterior to posterior). Analysis of slides after anti-BrdUrd antibody staining clearly indicated increased cell proliferation (unit length labelling indices, ULLI) after exposure to 6ppm and higher. No treatment-related effects were measured after exposure to 0.5, 1 and 2ppm. When comparing the cell-proliferation rate of normal epithelium with that of directly adjacent metaplastic epithelium, no consistent pattern was found: depending on the location, cell proliferation of normal epithelia was either higher or lower. Our results support previous findings demonstrating local cytotoxic effects in the nose of rats after inhalation of FA. However, induction of MN in the nasal epithelium as an indicator of a mutagenic effect was not seen. Because only limited experience exists for the MNT with rat nasal epithelial cells, this result has to be interpreted with great care. The contribution of mutagenicity to FA's carcinogenicity in rat nasal epithelium remains unclear.     

Micronuclei in peripheral lymphocytes and exfoliated urothelial cells of workers exposed to 4,4′-methylenebis-(2-chloroaniline) (MOCA)

4,4′-Methylenebis-(2-chloroaniline) (MOCA) is used in the manufacture of polyurethane. The IARC classifies MOCA as a probable human carcinogen. Suggested changes to guidelines for health surveillance of MOCA-exposed workers in Australia include a reduction in acceptable levels of urinary MOCA to below 15 μmol/mol creatinine. Twelve male workers aged 24 and 42 years were recruited into this study from four work locations where MOCA is used. Exfoliated urothelial cells from prework urine samples on a midweek work day were assessed for micronucleus (MN) frequencies. Postwork urine samples were analysed for total MOCA. Blood samples collected on the same day were cultured for 96 h and cytochalasin-B-blocked cells were scored for MN. Eighteen male control subjects (23–59 years) provided corresponding urine and blood samples. Median urinary MOCA concentrations were 6.5 μmol/mol creatinine (range 0.4–48.6 μmol/mol creatinine) in postwork samples of MOCA-exposed workers. MOCA was not detected in urine of control workers. Mean MN frequencies were higher in urothelial cells and lymphocytes of MOCA workers (14.27±0.56 and 13.25±0.48 MN/1000 cells) than in controls (6.90±0.18 and 9.24±0.29 MN/1000 cells). The mean number of micronucleate cells was also higher in both tissues of exposed workers (9.69±0.32 and 8.54±0.14 MN cells/1000 cells) than in controls (5.18±0.11 and 5.93±0.13 MN cells/1000). There was no correlation between postwork urinary MOCA concentrations and MN frequencies in either tissue. This study suggests that exposures to MOCA in South Australia are similar to those of a decade ago and are at levels similar to those currently acceptable in Australia. These are associated with genotoxic effects in urothelial cells and peripheral blood lymphocytes. It may be prudent to reduce MOCA exposures in line with proposed guidance values.     

Micronucleus frequency in human umbilical cord lymphocytes

The human fetus is exposed to a variety of environmental agents and drugs which cross the placenta and can induce DNA damage. Micronucleus (MN) determination is a suitable and sensitive method for measuring DNA damage and since umbilical cord blood is obtained without any risk for the newborn, we measured the frequency of MN in cells from cord blood in four groups of healthy newborns (NB): 35 NB whose mothers lived in two urban cities (groups I and II); 16 NB from an agricultural area (group III); and 15 NB of mothers with high-risk pregnancy (group IV). MN were also evaluated in the mothers of NB from group I (n=17) and group III (n=14). Acetylcholinesterase (AChE) concentration was measured in groups I and III. The average frequency of binucleated cells with MN was 3.7+/-1.4 in 1000 cells in mothers and 1+/-0.9 in 1000 cells in NB from urban areas; and 4.5+/-2.4 in 1000 cells in mothers and 2+/-1.5 in 1000 cells in NB from the agricultural area. The correlation between the frequency of MN in mothers and NB was significant (r=0.61, p<0.01). AChE levels of samples obtained both from group III mothers and from newborns were similar to those of group I. The Wilcoxon's rank-sum test was applied to measure differences in MN frequency; NB of group I were used as control group. A significant (p<0.01) higher frequency of MN (4+/-2) was found only in lymphocytes from NB from high-risk pregnancies. Data indicate that MN evaluation in umbilical cord samples might be useful in the identification of transplacental mutagens.