Mean Micronucleus Frequency Research Articles

Genotoxic effects of alcohol in human peripheral lymphocytes modulated by ADH1B and ALDH2 gene polymorphisms

Ethanol is almost totally broken down by oxidative metabolism in vivo. Ethanol per se is considered to be neither carcinogenic, mutagenic nor genotoxic. However, during the metabolic conversion of ethanol to acetaldehyde and acetate, the organism is exposed to both ethanol and acetaldehyde and therefore ethanol is suspected to be co-carcinogenic. The genetic polymorphisms of alcohol dehydrogenase-2 ( ADH1B) and acetaldehyde dehydrogenase-2 ( ALDH2) influence the metabolism of alcohol. The ADH1B*1/ *1 genotype encodes the low-activity form of ADH1B, and ALDH2*1/ *2 and ALDH2*2/ *2 genotype encode inactive ALDH2. The aim of this study was to test the hypothesis that polymorphisms of the ADH1B and ALDH2 genes are significantly associated with genotoxicity induced by alcohol drinking, measured using the cytokinesis-block micronucleus (CBMN) assay, an established biomarker of genome instability, in peripheral blood lymphocytes of 286 healthy Japanese men. There was a significant trend for the mean micronuclei (MN) frequency in habitual or moderate drinkers without a smoking habit to increase as the numbers of the *1 allele in ADH1B increased ( P = 0.039 or P = 0.029) and the *2 allele in ALDH2 increased ( P = 0.019 or P = 0.037). A logistic regression analysis showed that the number of subjects with MN frequency levels more than median value of MN (3.0) was significantly higher in the subjects with the ADH1B*1 allele as adjusted estimates (OR 2.08, 95% C.I. 1.24–3.48), when the OR for the subjects with the ADH1B*2/ *2 genotype was defined as 1.00. The number of subjects with MN frequency levels more than median value of MN was also significantly higher in the subjects with the ALDH2*2 allele as adjusted estimates (OR 1.79, 95% C.I. 1.04–3.11), when the OR for the subjects with the ALDH2*1/ *1 genotype was defined as 1.00. The results of this study have identified important novel associations between ADH1B/ ALDH2 polymorphisms and genotoxicity in alcohol drinkers.

Monitoring the genotoxic effects of radiosynovectomy with Re‐186 in paediatric age group undergoing therapy for haemophilic synovitis

The aim of this study was to investigate the genotoxic effect on the peripheral blood lymphocytes potentially induced by Re-186 in paediatric age group undergoing radiosynovectomy for haemophilic synovitis, by using chromosomal aberration analysis (CA) and the micronuclei (MN) assay for detecting chromosomal aberrations, as well as the sister chromatid exchanges (SCE) technique for assessing DNA damage. Cytogenetic analyses were evaluated in 20 boys (mean age: 13.8 +/- 2.7 years) before, and 2 and 90 days after radiosynovectomy from the peripheral lymphocytes of the patients. Joint retention and extra-articular spread of the radionuclides were evaluated by using quantitative gamma camera imaging. Imaging after radiosynovectomy revealed local lymph node visualization in 8 (40%) patients and hepatosplenic visualization in 3 (15%) patients due to extra-articular leakage of radioactive material. The mean frequency of chromosome aberrations (0.2 +/- 0.4/1000 cells) determined prior to the onset of therapy was not significantly increased in comparison with control values obtained 2 days (0.4 +/- 0.5/1000 cells) and 90 days (0.2 +/- 0.4/1000 cells) after therapy (P = 0.754 and P = 1.0). In the analysis of MN and SCE, when we compare the baseline levels, the mean MN and SCE frequencies were slightly higher in the control analyses performed 2 and 90 days after radiosynovectomy but there were no significant differences between baseline and control levels (chi(2) = 2.621, P = 0.270 and F = 0.573, P = 0.569, respectively). The major finding of this study with relatively small sample is that, radiosynovectomy with Re-186 does not seem to induce early genotoxic effects on the peripheral blood lymphocytes in paediatric age group.

Micronucleus frequency in women with genital Chlamydia Trachomatis infection before and after therapy

The main aim of the present study was to investigate the influence of infection with the intracellular bacterium Chlamydia trachomatis, and subsequent treatments with oral doxycycline or azithromycin on the frequency of micronuclei (MN) in peripheral blood lymphocytes of adult female patients receiving standard doses of these drugs. The frequency of micronuclei was measured in the lymphocytes of 38 newly diagnosed adult women with genital C. trachomatis infection. Samples were taken before and after the therapy, and from 50 healthy control females. The therapy was taken orally during 10 days at 2 × 100 mg per day, and then for another 10 days at 1 × 100 mg per day for doxycycline, and as a single dose of 1 g for azithromycin. Isolated lymphocytes from all subjects were cultured by use of the whole-blood method and blocked in metaphase with cytochalasin B (Cyt B). One thousand binucleate cells per subject were scored according to published criteria. The frequency of micronuclei was not significantly higher in samples of infected females before therapy, compared with the baseline frequency in healthy control females ( p > 0.05). In patients who received doxycycline, the micronucleus frequency after the end of therapy was significantly higher than before treatment ( p < 0.001). The mean frequency of micronuclei in females after the end of the therapy with azithromycin did not show an increase ( p > 0.05). The application of linear regression analysis showed that the difference in micronucleus frequency before and after therapy (effect of the antibiotics) was affected by the therapy type. Age and smoking did not affect micronucleus frequency in analyzed samples of patients ( p = 0.078, 0.579). We conclude that C. trachomatis infection does not induce micronuclei in peripheral blood lymphocytes of infected adult female patients. Therapy with doxycycline significantly increases the micronucleus frequency in lymphocytes of treated patients, but treatment with azithromycin does not induce micronuclei.

Micronuclei induction and chromosomal aberrations in Rattus norvegicus by chloroacetic acid and chlorobenzene

Chloroacetic acid (CAA) and chlorobenzene (CB) have been evaluated for in vivo mutagenic potential in Rattus norvegicus, employing the following criteria : (i) chromosomal aberrations (CAs) such as breaks, gaps, exchanges, rings, and multiple aberrations and (ii) micronuclei (MN) induction. Three sublethal doses, 0.008, 0.01, and 0.012 mg/g b. wt. of CAA and 0.75, 1.0, and 1.25 mg/g b. wt. of rat of CB were administered and the bone marrow cells evaluated in each of the three treated groups at 12, 24, and 48 h, respectively. Mean MN frequencies of 4.40±0.2 and 5.42±0.3, obtained respectively for CAA and CB. The higher induction of MN by CAA and CB was dose- and time-dependent. Most significant impact ( P<0.05) for either of the compounds was observed at 24 h post administration, when the recorded mean frequency of CAs was maximum for CAA (4.33±0.6) as well as for CB (4.66±0.5).

A polymorphism of the methionine synthase reductase gene increases chromosomal damage in peripheral lymphocytes in smokers

The cytogenetic effects of cigarette smoke has been evaluated as one of many potential confounders in a large number of biomonitoring studies of occupationally or environmentally exposed populations and control subjects. Despite the well-known presence of carcinogens in the cigarette smoke, the results in the scientific literature linking smoking habits to micronuclei (MN) frequency, one of the cytogenetic markers, are rather controversial. Here, we investigated the relationships among MN frequency, smoking habits and five folate metabolic enzyme gene polymorphisms ( MTHFR C677T and A1298C, MTR A2756G, MTRR A66G and TYMS 3′UTR) in 132 healthy Japanese men who were non-habitual drinkers. In never- and former-smokers, no statistically significant differences in the mean MN frequencies were observed according to the five folate metabolic enzyme gene polymorphisms. In current-smokers, however, subjects with the AA genotype for MTRR had a significantly higher mean MN frequency than the AG genotypes for MTRR ( p < 0.05). Furthermore, among subjects with the AA genotype for MTRR, current-smokers were found to have a significantly higher mean MN frequency than never- and former-smokers ( p < 0.05). To further characterize this association, we stratified the smoking status into five groups: non-smokers (never-smokers and former-smokers), 1–10 cigarettes/day, 11–20 cigarettes/day, 21–30 cigarettes/day and ≥31 cigarettes/day. There was an overall trend for the mean MN frequency in subjects with the MTRR AA genotype to increase as the number of cigarettes smoked per day increased ( p < 0.01, Jonckheere–Terpstra test). The results of our preliminary study suggest that the MTRR AA genotype acts to increase the MN frequency resulting from cigarette smoking. Therefore, studies on human genotoxicity based on cytogenetic markers of MN should take into account both the MTRR polymorphism and the potential confounding effect of smoking, although these preliminary findings need to be validated in larger populations because of the relatively small sample size.

Gene–environmental interactions between alcohol-drinking behavior and ALDH2 and CYP2E1 polymorphisms and their impact on micronuclei frequency in human lymphocytes

Ethanol is converted to acetaldehyde by alcohol dehydrogenase (ADH), cytochrome p4502E1 (CYP2E1) and catalase. This metabolite is then detoxified by aldehyde dehydrogenase 2 (ALDH2), a key enzyme in the elimination of acetaldehyde, via further oxidation to acetic acid. The toxic effects of acetaldehyde are well documented and may be partially mediated by genotoxic damage. In the present study, we investigated the effects of alcohol-drinking behavior and genetic polymorphisms in two different genes (ALDH2 and CYP2E1) on the micronuclei (MN) frequency in 248 healthy Japanese men. Genotyping was performed by PCR–RFLP analysis. The ALDH2 variant (deficient type) was significantly associated with an increased MN frequency in subjects drinking more than three times/wk, while habitual drinkers with wild-type CYP2E1 also had a significantly increased MN frequency. Furthermore, when the subjects were divided into eight groups according to their drinking frequency and genotypes of ALDH2 and CYP2E1, we found that habitual drinkers with homozygous CYP2E1 *1/ *1 and heterozygous ALDH2 *1/ *2 or homozygous ALDH2 *2/ *2 showed the highest mean MN frequency. In the present study, we found clear associations among ALDH2 and CYP2E1 gene polymorphisms, alcohol-drinking behavior and genotoxic effects in a healthy Japanese population. Therefore, analysis of the polymorphisms of alcohol-metabolizing enzymes may lead to elucidation of the mechanism(s) for individual susceptibilities to the toxicity of ethanol metabolites.

A Molecular Epidemiology Case Control Study on Pleural Malignant Mesothelioma

Pleural malignant mesothelioma is an uncommon neoplasm usually associated with asbestos exposure. The increasing incidence of malignant mesothelioma cases involving individuals with low levels of asbestos exposure suggests a complex carcinogenetic process with the involvement of other cofactors. Cytogenetic studies revealed the complexity of the genetic changes involved in this neoplasm reflecting the accumulation of genomic damage. One of the most used methodologies for assessing genomic damage is the cytokinesis-blocked micronucleus test applied in peripheral blood lymphocytes (PBL). This approach allows the detection of chromosomal alterations expressed in binucleated cells after nuclear division in vitro. This marker could provide a tool for assessing genetically determined constitutional differences in chromosomal instability. A biomonitoring study was carried out to evaluate the micronuclei frequency in PBLs of patients with pleural malignant mesothelioma with respect to lung cancer, healthy, and risk controls as a marker of cancer susceptibility in correlation with the presence of SV40. A significant increased micronuclei frequency was observed in patients with malignant mesothelioma in comparison with all the other groups, the mean micronuclei frequency was double in patients with malignant mesothelioma compared with healthy controls, risk controls, and patients with lung adenocarcinoma (median 11.4 binucleated cells with micronuclei/1,000 binucleated cells versus 6.2, 6.1, and 5.1, respectively). Our data indicate that human T lymphocyte samples carry DNA sequences coding for SV40 large T antigen at low prevalence, both in cancer cases and controls. Evidence of cytogenetic damage revealed as micronuclei frequency in mesothelioma cancer patients could be related to exogenous and endogenous cofactors besides asbestos exposure.

Cytogenetic biomonitoring in children with chronic tonsillitis: Micronucleus frequency in exfoliated buccal epithelium cells

To investigate the possible harmful cytogenetic effects associated with chronic tonsillitis by analyzing the micronucleus frequency and other nuclear abnormalities in exfoliated buccal epithelial cells. The study consisted of 20 children with chronic tonsillitis, and 20 control subjects with similar age and sex. The ages ranged between 5 and 12 years old (mean age: 7.5). The patients were diagnosed as having chronic tonsillitis on the basis of history, throat culture and clinical examinations. Buccal cell samples were collected with a wooden spatula. The samples were then applied to clean microscope slides. Smears were air dried and fixed in methanol:acetic acid. Then slides were stained by the Feulgen reaction technique. Three slides were prepared for each subject and 1000 cells were evaluated per slide to determine the frequencies of micronucleus and other nuclear abnormalities (binucleats, karyorrhexis and karyolysis). Statistically, Mann-Whitney U-test was used to analyze and compare the data. The mean micronucleus frequencies in patient and control groups were 5.29+/-1.67 and 1.58+/-0.33, respectively. In the patient group, mean binucleus, karyorrhexis and karyolysis frequencies were 3.13+/-1.2, 2.04+/-0.64, and 1.74+/-0.47, respectively. However, in the control group, mean binucleus, karyorrhexis and karyolysis frequencies were 1.43+/-0.47, 1.26+/-0.45, and 0.88+/-0.27, respectively. The mean frequencies of all parameters in the patient group were higher than the control values, and the difference was found to be statistically significant (p<0.001). Our results revealed that children with chronic tonsillitis could be under risk of significant cytogenetic damage.

Comparison of genomic damage caused by 5-nitrofurantoin in young and adult mice using the in vivo micronucleus assay

The antibiotic 5-nitrofurantoin (5-NF) has been used widely for the treatment of urosepsis in children during the last 20 years. Recent experimentation suggests the need for reevaluating its genotoxic potential. Because of possible differences in the metabolism and clearance of 5-NF in young and adult animals, we conducted a study to determine whether micronuclei caused by 5-NF were age-related. The in vivo micronucleus (MN) assay was performed on 3- and 8-week-old mice given single intraperitoneal injections of 5, 10, and 50 mg/kg 5-NF. Blood samples from the tail vein were taken before injection (baseline) and at 48, 96, 168, and 336 hr (2 weeks) after the treatment. One thousand reticulocytes were analyzed for micronuclei from each animal. Compared to similar baseline values for young and adult mice, 5-NF caused a significant increase in MN frequency in both age groups. The mean MN frequency in the young animals was higher than in the adult animals for each dose and sampling time. MN frequencies remained significantly elevated in young animals even 2 weeks after exposure to 5-NF. The results of the study confirm the genotoxic potential of 5-NF in young and adult animals, and indicate that young animals are more sensitive to the genotoxic effects of 5-NF than adult mice and that the response in young mice persists for a significantly longer time. These findings may be related to poorly developed mechanisms of xenobiotic detoxification and renal elimination in young animals.

An automated scoring procedure for the micronucleus test by image analysis.

The micronucleus assay (MNT) in human lymphocytes is frequently used to assess chromosomal damage as a consequence of environmental mutagen exposure, to assess the effect of mutagens or to search for reduced DNA repair capacity after a mutagenic challenge. We have established an automated scoring procedure for the cytokinesis blocked MNT based on computerized image analysis (Metasystems Metafer 4 version 2.12). To evaluate the results we used the reproducibility of counts, established a dose-response curve for gamma-irradiation and used the ability of the system to differentiate between breast cancer patients and controls as a biological reference, a difference which we had observed before by visual counting. Blood cultures were irradiated with gamma-rays (2 Gy) at the beginning and treated with cytochalasin B during the last 24 h. The slides were stained with Giemsa for visual counting and with DAPI for automated analysis. Our test sample consisted of 73 persons (27 with breast cancer and 26 female and 20 male controls). A comparison between visual counting (controls, mean MN frequency 313) and automated counting (mean MN frequency 106) in slides from the same culture revealed a large drop for the automated counts. However, the automated counts were as reproducible as the visual counts [coefficient of variation (CV) on the sample approximately 20%; CV on repeated counts of the same slides approximately 5%] and both counts were highly correlated. Furthermore, the discrimination between cases and controls improved for automated counting of slides from the same cultures [visual odds rato (OR) < or = 4.0, P = 0.009; automated OR > 16, P < 0.0001], with a strong dependence on the set of parameters used. This improvement was confirmed in a validation sample of an additional 21 controls and 20 cases (OR = 11, P = 0.0018) performed as a prospective or diagnostic test.

Evidence of association of the CYP2E1 genetic polymorphism with micronuclei frequency in human peripheral blood

Micronuclei (MN) are used as one of the cytogenetic biomarkers, and intra- and inter-individual variations in this frequency have been reported in human blood lymphocytes. Polymorphisms in a few metabolic enzyme genes seem to account for a proportion of this variability, but the impacts of specific genetic variants on the MN frequency have not yet been clarified. Here, we investigated the relationship between the MN frequency and several gene polymorphisms in 90 healthy Japanese men. The subjects with the CYP2E1 ∗3 variant allele had a statistically lower mean MN frequency than subjects with the CYP2E1 ∗1/ ∗1 wild type. Furthermore, the adjusted odds ratio (OR) of the CYP2E1 ∗3 variant with higher MN frequency levels was also significantly lower and calculated to be 0.25 (95% CI 0.07–0.83), when the OR for the subjects with the CYP2E1 ∗1/ ∗1 wild type was defined as 1.00. These data suggest that the CYP2E1 ∗3 polymorphism may have the potential to influence the baseline frequency of MN.

Effects of ALDH2 gene polymorphisms and alcohol-drinking behavior on micronuclei frequency in non-smokers

Alcohol abuse is a serious health problem, leading to life-threatening damage to most of the important organ systems. Genotoxic damage is used as an early effect indicator in the surveillance of human exposure to genotoxic substances. Intra- and inter-individual variations of baseline frequencies of micronuclei (MN) in peripheral blood lymphocytes of human populations have been reported previously. Polymorphisms in a few metabolic enzyme genes seem to account for a proportion of this variability, but the impact of specific genetic variants on MN frequencies has not yet been clarified. In 42 healthy Japanese non-smoking men, we investigated the relationship between the MN frequency levels and genetic polymorphisms in three different genes: aldehyde dehydrogenase 2 (ALDH2), X-ray repair cross-complementing group 1 (XRCC1) and excision repair cross-complementing group 2 (ERCC2). Genotyping was performed by PCR–RFLP analysis. The ALDH2 variant (deficient-type) was significantly associated with increased MN frequency levels in subjects with drinking more than three times per week, whereas the XRCC1 and ERCC2 variants seemed to be unrelated to the MN frequency. The ALDH2-deficient habitual drinkers had an average MN frequency of 5.88±0.58 (±S.E.) compared with 3.20±0.80 in the ALDH2-proficient habitual drinkers ( P<0.05). The ALDH2-proficient non-habitual drinkers had the lowest MN frequency (1.56±0.41). Furthermore, subjects with highest levels of mean MN frequency, who consumed more than 100 g of alcohol per week and more than three times per week, had A2 genotype of ALDH2. A significant odds ratio (12.25, P<0.05) for the MN frequency levels above the 50th percentile value was observed for the ALDH2-deficient individuals versus the ALDH2-proficient individuals after adjustment for several confounders. These results strongly suggest that human early genotoxic effect studies based on the cytogenetic markers of MN should take into account both the individual ALDH2 polymorphism and the potential confounding effect of the drinking behavior.

Assessment of cytogenetic damage in lymphocytes and in exfoliated nasal cells of dental laboratory technicians exposed to chromium, cobalt, and nickel

Dental laboratory technicians may be exposed to metal alloys that are used in the production of crowns, bridges and removable partial dentures. These alloys consist of 35–65% cobalt, 20–30% chromium, 0–30% nickel, and small amounts of molybdenum, silica, beryllium, boron and carbon. The aim of this study was to assess whether dental technicians are occupationally exposed to chromium, cobalt and nickel, by analyzing urinary excretion levels of these metals and to investigate the genotoxic effects of occupational exposure associated with dental prostheses production operations by analyzing cytokinesis-blocked micronucleus (CB-MN) frequencies in peripheral lymphocytes and micronucleus (MN) frequencies in exfoliated nasal cells from 27 dental laboratory technicians and 15 control subjects. The differences in the urinary excretion of metals between technicians and controls were statistically significant. The mean (±S.D.) CB-MN frequencies (‰) in peripheral lymphocytes were 4.00 (±2.98) among the dental technicians and 1.40 (±1.30) among the controls, a statistically significant difference ( P<0.005). The mean (±S.D.) MN frequencies (‰) in nasal cells were 3.50 (±1.80) among the dental technicians and 1.19 (±0.53) among the controls, which was also a statistically significant difference ( P<0.005). There was a significant correlation between duration of exposure and MN frequencies in lymphocytes ( r=0.642, P<0.01), but not in nasal cells of technicians. Our data reveal that in vivo exposure to chromium, nickel and cobalt metals is evident and that this occupational exposure may contribute to the observed genotoxic damage in two types of cells, e.g. lymphocytes and exfoliated nasal cells. However, it cannot be determined which compound(s) are responsible for the genotoxic damage observed in this study.

Micronucleus Assay in Patients with Lung Cancer, Chronic Obstructive Pulmonary Disease and Non-Smoker Control

During the last decades, cytogenetic markers in peripheral blood lymphocytes have been used to assess exposure to carcinogenic or mutagenic agents. This study was conducted with the aim of assessing the use of micronucleus [MN] assay as a marker in lung cancer and whether this method could predict cancer risk in patients with chronic obstructive pulmonary disease [COPD] as it is known that lung cancer is far more common in patients with COPD than in those with normal airflow. The study included 96 male subjects; 40 patients with lung cancer, 36 patients with COPD and 20 control subjects. Twenty five percent of cases with lung cancer and 86.1% of cases with COPD had MN frequency ranging from < 5-<15 MN/500 cytokinesis blocked [CB] binucleated cells compared to 100% in the control group. Forty percent of cases with lung cancer and 13.9% of cases with COPD had 15-20 MN/500 CB binucleated cells. Thirty five percent of cases with lung cancer had 20-25 MN/500 CB binucleated cells. These differences are statistically significant [X2= 75.2, p < 0.000]. In the lung cancer group, the mean MN/500 CB binucleated cells was 18.4±4.2 [range:12-27 MN/500 CB binucleated cells], in the group with COPD, the mean MN/500 CB binucleated cells was 8.3+4.1 [range:1-17 MN/500 CB binucleated cells] with 8 patients [22.2%] having MN in the cancer range, i.e. 12-17. The mean MN/500 CB binucleated cells of the control group was 5.4±2.5 [range:2-12 MN/500 CB binucleated cells]. These differences are statistically significant [least significant difference test]. All cases with lung cancer/and 63.9% of cases with COPD had mean MN frequency greater than the upper 95% CI of the mean MN frequency for the control group. Applying logistic regression analysis for the effect of risk factors revealed that lung cancer patients who were current or ex smokers had significantly higher MN frequencies than patients with COPD. The elevated levels of micronuclei in patients with lung cancer implies that micronucleus assay could be used as a method to detect cancer. Patients suffering from COPD who have elevated levels of micronuclei need to be followed up closely.

Cytogenetic analysis of buccal cells from shoeworkers 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 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.

Micronuclei in the peripheral blood and bone marrow cells of rats exposed to 2450 MHz radiofrequency radiation

Purpose : To determine the incidence of micronuclei in peripheral blood and bone marrow cells of rats exposed continuously for 24h to 2450 MHz continuous wave radiofrequency radiation (RFR) at an average whole-body specific absorption rate (SAR) of 12W/kg. Materials and methods : Eight adult male Sprague-Dawley rats were exposed to 2450 MHz RFR in circularly polarized waveguides. Eight sham-exposed rats were kept in similar waveguides without the transmission of RFR. Four rats were treated with mitomycin-C (MMC) and used as positive controls. All rats were necropsied 24h after the end of RFR and sham exposures, and after the 24h treatment with MMC. Peripheral blood and bone marrow smears were examined to determine the frequency of micronuclei (MN) in polychromatic erythrocytes (PCE). Results : The results indicated that the incidence of MN/2000 PCE were not significantly different between RFR- and sham-exposed rats. The group mean frequencies of MN in the peripheral blood were 2.3 ±0.7 in RFR-exposed rats and 2.1 ±0.6 in sham-exposed rats. In bone marrow cells, the average MN incidence was 3.8 ±1.0 in RFR-exposed rats and 3.4 ±0.7 in sham-exposed rats. The corresponding values in positive control rats treated with MMC were 23.5 ±4.7 in the peripheral blood and 33.8 ±7.4 in bone marrow cells. Conclusion : There was no evidence for the induction of MN in peripheral blood and bone marrow cells of rats exposed for 24h to 2450 MHz continuous wave RFR at a whole body average SAR of 12 W/kg.

Detection of micronuclei in haemocytes of zebra mussel and great ramshorn snail exposed to pentachlorophenol

The frequency of micronuclei (MN) induced by pentachlorophenol (PCP) in haemocytes of zebra mussel, Dreissena polymorpha Pall. and great ramshorn snail, Planorbarius corneus L. was determined over a 14 days of exposure (sampling after 4, 7 and 14 days) under laboratory conditions. PCP doses for zebra mussel ranged from 10 to 150 μg/l, and for ramshorn snail from 10 to 450 μg/l. Micronuclei were detected after bisbenzimide fluorescent staining. Positive responses were observed in both species. The mean MN frequencies in treated mussels ranged between 0.69 and 7.50‰, and between 2.07 and 13.80‰ in treated snails. The spontaneous MN levels in mussels averaged from 0.5 to 2.75‰, and in snails from 1.56 to 2.00‰. Our results suggest that haemolymph of both species represent an appropriate test tissue in environmental genotoxicity assessment.

Monitoring of genotoxic damage in smokeless tobacco ((Maraş powder) consumers using micronucleated exfoliated oral cells

Maraş powder is a kind of smokeless tobacco used in the south-eastern region of Turkey. The present study was carried out to assess possible DNA damage in exfoliated oral cells of Maraş powder users by analysing the frequencies of micronuclei (MN), which is a simple and reliable biomarker for genotoxic damage and to screen for the detection of site-specific differences in the frequencies of MN. The mean (±SD) MN frequency in the inner lip mucosa site was 1.27(±0.55) % for Maraş powder users and 0.88(±0.47) % for non-smoking control subjects (p < 0.05) and 0.82(±0.40) % for the buccal site of Maraş powder users (p < 0.01). There was no significant site-specific difference between the inner lip site and the buccal mucosa site 0.73(±0.43) % in the MN frequency of non-smoking control subjects (p > 0.05). There was no significant effect of daily consumption of Maraş powder, and duration of usage on MN frequencies. The present study suggests that the oral use of smokeless tobacco represents a genotoxic hazard and also that use of MN from a single site may be misleading as a marker of genotoxic exposure. Habitual use of Maraş powder should be taken into account and could be considered unsafe.

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.

The micronucleus assay in exfoliated buccal cells: application to occupational exposure to polycyclic aromatic hydrocarbons

Many polycyclic aromatic hydrocarbons (PAHs) have been identified as cancer-inducing chemicals for animals and/or humans. Also, there is sufficient evidence that exposures in the occupational settings are carcinogenic or probably carcinogenic to human. Engine exhaust and used engine oils are major PAH sources in engine repair workshops and traffic. Analysis of micronucleus (MN) in exfoliated buccal cells is a sensitive method for monitoring genetic damage in human populations. In our study, we used three different occupational groups (Group 1; engine repair workers, Group 2; taxi drivers, Group 3; traffic police) and two controls (Control I for Group 1 and Control II for Group 2 and Group 3) for the exposed groups. We analysed MN frequencies in exfoliated buccal cells and compared the exposed groups (Group 1; n=34, Group 2; n=17, Group 3; n=15) and subjects not occupationally exposed to PAH (Control I; n=28, Control II; n=20). The mean (±S.D.) MN (%) frequencies in exfoliated buccal cells from Group 1 and Control I were 0.07±0.05 and 0.05±0.04, respectively ( p>0.05; Table 2). The mean (±S.D.) MN (%) frequencies in exfoliated buccal cells from Group 2, 3 and Control II were 0.12±0.05, 0.10±0.05 and 0.03±0.03, respectively ( p<0.0001, p<0.05; Table 2) Smokers and nonsmokers do not differ with respect to the incidence of MN in all groups.