Sister Chromatid Exchange Analysis Research Articles

Alteration of sister chromatid exchange frequencies in gastric cancer and chronic atrophic gastritis patients with and without H. pylori infection.

To determine, by counting sister chromatid exchange (SCE) frequencies, whether genetic impairment and DNA damage have an effect on the pathogenesis of gastric cancer (GC). Analysis of SCE is a cytogenetic technique used to show DNA damage as a result of an exchange of DNA fragments between sister chromatids. We analyzed SCE frequency in 24 patients with GC, 26 patients with chronic atrophic gastritis (CAG), and 15 normal controls. The presence of H. pylori was confirmed by urease test, toluidine-blue stain and hematoxylin-eosin stain. SCE was significantly increased in H. pylori-negative GC patients, and in H. pylori-negative CAG patients compared with controls (7.41 +/- 1.36 and 6.92 +/- 1.20, respectively, vs 5.54 +/- 0.8, P < 0.001). There was no difference in the SCE frequency between H. pylori-negative GC patients and H. pylori-negative CAG patients (P > 0.05). On other hand, the SCE frequencies in H. pylori-positive GC patients were higher than those in H. pylori-positive CAG patients (9.20 +/- 0.94 vs 7.93 +/- 0.81, P < 0.01). Furthermore, H. pylori-positive GC patients had a higher SCE frequency than H. pylori-negative GC patients (9.20 +/- 0.94 vs 7.41 +/- 1.36, P < 0.001). Similarly, a significant difference was detected between H. pylori-positive CAG patients and H. pylori-negative CAG patients (7.93 +/- 0.81 vs 6.92 +/- 1.20, P < 0.05). We suggest the increased SCE in patients reflects a genomic instability that may be operative in gastric carcinogenesis.

The Effects of Etodolac, Nimesulid and Naproxen Sodium on the Frequency of Sister Chromatid Exchange after Enclused Third Molars Surgery

PurposeNon-steroidal anti-inflammatory drugs (NSAID) are frequently used in oral surgical procedures in dentistry. The evaluation of the frequency of sister chromatid exchange (SCE) is accepted as a reliable cytogenetic method to assess the genotoxic effects of environmental factors.Materials and MethodsIn this study, the genotoxic effects of various NSAIDs were assessed in 30 patients to who they were administered following encluosed third molar surgery using SCE analysis before and after the operation. The frequency of SCE was evaluated before the operation and after 3 days of etodolac, nimesulid and naproxen use.ResultsThere was no statistically significant difference in the frequency of SCE between the preoperative and postoperative states in patients given etodolac, nimesulid or naproxen sodium.ConclusionShort term use of selective and non-selective NSAIDs was not associated with a significant genotoxic effect that could be detected using the SCE method in peripheric lymphocytes.

Response of Human Lymphocytes In Vitro to Two Dietary Furans, Furfuryl Alcohol and 2-Furyl Methyl Ketone

The genotoxicity of furfuryl alcohol and 2-furyl methyl ketone was assessed in human lymphocytes in vitro using standard chromosome aberration analysis and sister chromatid exchange (SCE) analysis. Three doses ranging from 240 to 960 ppm which corresponded to in vivo doses were selected. Lymphocytes were cultured using TC-199 medium plus required supplements and initiated using blood drawn from healthy human donors who had not been exposed to any drugs or chemicals for a substantial period of time. After 24, 48, and 72 hr of compound exposure, slides were prepared and analyzed for variations in the mitotic index (MI) and chromosome aberrations. For SCE analysis, 10 μg of 5-bromodeoxy uridine was added 24 hr after culture initiation and slides were prepared and stained using the standard technique. Two hundred metaphase spreads were scored per dose per period to analyze chromosome aberrations, and 5000 cells were randomly scored to calculate the MI. Seventy-five metaphase spreads in second mitotic divisions were scored for each dose and period for SCE analysis. All data were subjected to stastical analysis. There was an insignificant induction of chromosome aberrations and mitodepression (MD), while SCEs showed an increased incidence with both compounds which was dose and period dependent.

Analysis of Spontaneous and Streptonigrin-Induced Sister Chromatid Exchanges in Peripheral Lymphocytes of Aircrew Members of International Flights

In this work, we extend our previous studies concerning mutagen sensitivity in flight personnel from commercial airlines by analyzing the frequency of spontaneous and streptonigrin (SN)-induced sister chromatid exchanges (SCEs) in peripheral blood lymphocytes of 18 long-haul aircrew members from Argentina and of 18 control individuals. Statistical analysis revealed no significant differences between aircrew and controls in the background level of SCEs (p > 0.05), which suggests that chronic exposure to cosmic radiation and other occupational hazards does not affect SCEs frequency in peripheral lymphocytes of aircrews. The fact that almost no correlation was found between cumulative flight hours and the yield of spontaneous SCEs in aircrews adds further support to this assumption. Therefore, the background SCEs frequency cannot be use as a valid biomarker to determine the genotoxic effects of cosmic radiation or other occupational hazards exposure in aircrews. Following SN treatment, a significant increase in the mean frequency of SCEs was observed in the control group (p < 0.05) but not in the aircrew group (p > 0.05), suggesting that at the population level, aircrew are more resistant to the mutagenic effects of SN than controls. The reasons of this resistance remain to be determined. Since cosmic radiation had no effect on the background SCEs frequency and no relationship was found between cumulative flight hours and SCEs inducer effect by SN in aircrews, a direct effect of cosmic radiation on SN resistance should be discarded.

Application of cytogenetic endpoints and comet assay on human lymphocytes treated with atorvastatin in vitro

This study investigated the genotoxic potential of atorvastatin on human lymphocytes using comet assay, structural chromosome aberrations (CA) and sister-chromatid exchange (SCE) analysis. Lymphocyte cultures were treated with a single drug at a concentration of 30.21 ng/mL. For comet assay, cells exposed to atorvastatin for 24 h, 48 h and 72 h were embedded in agarose slides, lysed with alkaline lysis solution and exposed to an electric field. DNA migrated within the agarose and formed comets whose length depends on the amount of DNA damage. For analysis of structural CA, cells were grown on medium for 48 h and for SCE analysis for 72 h. Structural CA did not induce significant damage to the genome, although a higher CA frequency was observed in cells treated with atorvastatin for 3 h, 20 h and 48 h than in control samples. Results of the SCE analysis did show statistically significant differences in the mean SCE number between atorvastatin-exposed and control human lymphocytes and between different exposure times. Comet assay also showed increased DNA damage caused in atorvastatin-exposed human lymphocytes than in corresponding control cells for exposure times of 24 h, 48 h and 72 h for the tail length and for 72 h for the tail moment. Results obtained in this study point to the significance of biological indicators providing information on the primary genome damage after long-term exposure, which can help to establish drug therapeutic concentrations that do not put patients with high blood cholesterol to a greater treatment-related risk.

DNA Methyltransferases Modulate the Bystander Effect in Mouse Embryonic Stem Cells.

Cells exposed to radiation or other genotoxic agents can induce DNA damage and other stress responses in non-irradiated cells that are either cultured with the irradiated cells or have been exposed to culture medium from irradiated cells. This is called the bystander effect. In a previous study we found that the descendents of bystander cells exposed to Mitomycin C (MMC) are themselves capable of inducing homologous recombination in un-exposed cells. This suggests that MMC induces persistent and transmissible changes in expression in bystander cells. Bystander effects are likely caused by epigenetic mechanisms rather than “classic” mutations, i.e. changes in DNA sequence. One of the epigenetic mechanisms cells employ for changing expression is DNA methylation in which DNA methyltransferases (DNMTs) add a methyl group to the 5 carbon of cytosine. In this study we asked if ionizing radiation can induce transmissible DNA damage in bystander cells by examining if bystander cells exposed to irradiated cells were themselves able to induce damage in naive cells. Furthermore, we asked if this was dependent on DNMT activity in the irradiated cells. We irradiated wild-type (WT) and DNMT triple knockout (DNMT TKO) mouse embryonic stem cells (ESCs) and after two weeks of continuous culture, we collected conditioned medium (CM). CM was then added to cultures of naive WT ESCs (primary bystanders). Three weeks later, CM was collected from the primary bystanders and added to naïve WT cells (secondary bystanders). We assessed DNA damage by evaluating strand breaks using the alkaline Comet assay and sister chromatid exchange (SCE) analysis. As expected, we found that medium from cells irradiated with 5 Gy induced modest damage in bystander cells. The median Olive tail moment was 2.8 in bystander cells exposed to conditioned medium from irradiated cells compared to 1.0 in control bystander cells (p < 0.0001). Homologous recombination was 0.15 chromatid exchanges per chromosome compared to 0.092 in control bystanders (p < 0.0001). We also observed an increase in strand breaks in secondary bystanders of a similar magnitude to that found in primary bystanders, indicating that radiation-induced bystanders are themselves able to induce damage. In contrast to WT cells, the irradiated DNMT TKO cells did not induce strand breaks in bystander cells, as measured by the Comet assay, but did induce HR. Surprisingly, we also observed that un-irradiated DNMT TKO cells induce DNA damage in bystanders, and furthermore that the magnitude of the effect is similar to that induced by irradiated WT cells. These data suggest that methyltransferases have a complex role in bystander effects. Bystander effects may be mediated by free radicals. To see if the DNMT TKO cells had changes in antioxidant levels, glutathione (GSH) and glutathione peroxidase (GPX) activity were determined. There was no significant change in GSH levels between WT and DNMT TKO cells. However, DNMT TKO cells had significantly higher levels of GPX activity (275.4 + 19.8 mU/mg protein) compared to control cells (122.0 + 16.4 mU/mg, p= 0.0001). Taken together, these results show that radiation-induced bystander cells can themselves induce damage in un-irradiated cells and suggest that cells lacking DNA methylation activity can induce bystander effects.

Assessment of the genotoxicity of imidacloprid and metalaxyl in cultured human lymphocytes and rat bone-marrow

Imidacloprid and metalaxyl are two pesticides that are widely used in agriculture, either separately, or in combination. These agents were studied for their possible genotoxic effects with respect to the following cytogenetic end-points: (1) in vitro micronucleus (MN) formation and sister-chromatid exchange (SCE) induction in human lymphocytes and (2) in vivo micronucleus induction in polychromatic erythrocytes (PCEs) of the rat bone-marrow. The results of the MN analysis indicate that MN frequencies after treatment with both pesticides, separately or as a mixture, do not significantly differ from those in the controls except after treatment with metalaxyl alone at 50 μg/ml ( p < 0.05). The results of the SCE analysis show that SCE frequencies after treatment with imidacloprid do not differ significantly from those in the controls. A statistically significant increase ( p < 0.05) in SCE frequency resulted from treatments with metalaxyl at 5, 10 and 100 μg/ml and with the combination of imidacloprid and metalaxyl at 100 and 200 μg/ml. Finally, the in vivo micronucleus assay with rat bone-marrow polychromatic erythrocytes showed a statistically significant effect upon separate treatments with imidacloprid and metalaxyl at doses of 300 mg/kg body weight (b.w.) ( p < 0.01) or upon combined treatment with 200 mg/Kg b.w. ( p < 0.001) and 400 mg/kg b.w. ( p < 0.05).

The chlorophenoxy herbicide dicamba and its commercial formulation banvel ® induce genotoxicity and cytotoxicity in Chinese hamster ovary (CHO) cells

The sister chromatid exchange (SCE) frequency, the cell-cycle progression analysis, and the single cell gel electrophoresis technique (SCGE, comet assay) were employed as genetic end-points to investigate the geno- and citotoxicity exerted by dicamba and one of its commercial formulation banvel ® (dicamba 57.71%) on Chinese hamster ovary (CHO) cells. Log-phase cells were treated with 1.0–500.0 μg/ml of the herbicides and harvested 24 h later for SCE and cell-cycle progression analyses. All concentrations assessed of both test compounds induced higher SCE frequencies over control values. SCEs increased in a non-dose-dependent manner neither for the pure compound ( r = 0.48; P > 0.05) nor for the commercial formulation ( r = 0.58, P > 0.05). For the 200.0 μg/ml and 500.0 μg/ml dicamba doses and the 500.0 μg/ml banvel ® dose, a significant delay in the cell-cycle progression was found. A regression test showed that the proliferation rate index decreased as a function of either the concentration of dicamba ( r = −0.98, P < 0.05) or banvel ® ( r = −0.88, P < 0.01) titrated into cultures in the 1.0–500.0 μg/ml dose-range. SCGE performed on CHO cells after a 90 min pulse-treatment of dicamba and banvel ® within a 50.0–500.0 μg/ml dose-range revealed a clear increase in dicamba-induced DNA damage as an enhancement of the proportion of slightly damaged and damaged cells for all concentrations used ( P < 0.01); concomitantly, a decrease of undamaged cells was found over control values ( P < 0.01). In banvel ®-treated cells, a similar overall result was registered. Dicamba induced a significant increase both in comet length and width over control values ( P < 0.01) regardless of its concentration whereas banvel ® induced the same effect only within 100.0–500.0 μg/ml dose range ( P < 0.01). As detected by three highly sensitive bioassays, the present results clearly showed the capability of dicamba and banvel ® to induce DNA and cellular damage on CHO cells.

Genotoxic biomonitoring study of population residing in pesticide contaminated regions in Göksu Delta: Micronucleus, chromosomal aberrations and sister chromatid exchanges

Pesticides are widely used throughout the world in agriculture to protect crops and in public health to control diseases. Nevertheless, exposure to pesticides represents a potential risk to humans. This paper describes a study of possible genetic damage in the people living in regions contaminated with complex mixture of pesticides in Göksu Delta. In this study, used methods were chromosomal aberration (CA), sister chromatid exchange analysis (SCE) in the peripheral blood lymphocytes, and micronucleus (MN) assay in the buccal epithelial cells. In the present investigation, 32 affected subjects consist of 16 smoking and 16 non-smokings and an equal number of control subjects were assessed for genome damage. Micronucleus (MN), Broken egg (BE), Karyorrhexis (KR), Karyolysis (KL) and Binucleus (BN) frequencies were higher in affected subjects than in controls. Smoking had a statistically significant effect on the Micronucleus, Karyorrhexis and Binucleus frequencies for both the control and the exposed group. Also smoking and exposure affected the frequency of sister chromatid exchange and chromosomal aberrations compared with control groups.

Detection of genotoxicity in the marine environment: A preliminary feasibility study using primary mussel tissue culture

The purpose of this study was to evaluate the feasibility and potential usefulness of primary cultures of somatic tissues from adult mussel by means of sister chromatid exchange induction (SCE). This research is an initial pilot study carried out with mussel mantle tissue using seawater artificially contaminated with cadmium and polluted seawater from the port of Arcachon. With regard to cadmium concentration, mean SCE numbers showed a progressive increase from 1.07 +/- 0.18 per diploid cell in controls (i.e. cultures without contaminant) to 2.91 +/- 0.42 per diploid cell for the highest concentration, 10 - 4 M. With regard to the medium prepared with seawater from the port of Arcachon, the mean SCE number reached a value of 5.85+/- 0.85 per diploid cell. The analysis of SCEs induced by cadmium showed DNA responses even at the lowest concentration (i.e. 10 - 7 M). The study demonstrates the feasibility of the sister chromatid exchange (SCE) approach based upon primary mussel tissue culture, for the genotoxicity testing of contaminated seawater. Highlights from this procedure are (1) the presence of an active cell proliferation, (2) the use whole-water samples, (3) the possibility of culturing without serum, (4) the absence of cell dissociation before culturing and (5) a cellular proliferation which can be obtained in cultures carried out in a medium containing seawater whose salinity is comprise between 28 and 35 per thousand.

Irinotecan Toxicity to Human Blood Cells in vitro: Relationship between Various Biomarkers

Toxic effects of the antineoplastic drug irinotecan on human blood cells at concentrations of 9.0 microg/ml and 4.6 microg/ml were evaluated in vitro. Using the alkaline and neutral comet assay significantly increased levels of primary DNA damage in lymphocytes were detected. The induction of apoptosis/necrosis, as determined by a fluorescent assay, was also notably increased. Cytogenetic outcomes of the treatment were assessed by the analysis of structural chromosome aberrations and fluorescence in situ hybridization. A significantly higher incidence of chromatid breaks and complex quadriradials was observed. Painted chromosomes 1, 2 and 4 were equally involved in translocations, but only the chromosome 1 was involved in the formation of quadriradials. Sister chromatid exchange analysis was performed in parallel with the analysis of lymphocyte proliferation kinetics. The higher concentration of irinotecan caused almost seven-time increase, while the lower one caused a five-time increase of the basal sister chromatid exchange frequency, accompanied with significant lowering of the lymphocyte proliferation index. Using the cytokinesis-block micronucleus assay, a dose-dependent increase in micronucleus frequency along with the formation of nuclear buds and nucleoplasmic bridges was noticed. Inhibitory effects of irinotecan on enzyme acetylcholinesterase (AChE) were studied in erythrocytes. An IC(50) value of 5.0 x 10(-7) was established. Irinotecan was found to be strong inhibitor of the acetylcholine hydrolysis and to cause a continuous decrease of catalytic activity of AChE. The results obtained on a single donor may contribute to the understanding of irinotecan toxicity, but further in vitro and in vivo studies are essential in order to clarify remaining issues, especially on possible inter-individual variability in genotoxic responses to the drug.

Elevated sister chromatid exchange frequencies in New Zealand Vietnam War veterans

From July 1965 until November 1971, New Zealand Defence Force Personnel fought in the Vietnam War. During this time more than 76,500,000 litres of phenoxylic herbicides were sprayed over parts of Southern Vietnam and Laos, the most common being known as ‘Agent Orange’. The current study aimed to ascertain whether or not New Zealand Vietnam War veterans show evidence of genetic disturbance arising as a consequence of their now confirmed exposure to these defoliants. A sample group of 24 New Zealand Vietnam War veterans and 23 control volunteers were compared using an SCE (sister chromatid exchange) analysis. The results from the SCE study show a highly significant difference (P < 0.001) between the mean of the experimental group (11.05) and the mean of a matched control group (8.18). The experimental group also has an exceptionally high proportion of HFCs (cells with high SCE frequencies) above the 95th percentile compared to the controls (11.0 and 0.07%, respectively). We conclude that the New Zealand Vietnam War veterans studied here were exposed to a clastogenic substance(s) which continues to exert an observable genetic effect today, and suggest that this is attributable to their service in Vietnam.

Genotoxic Evaluation of Furfuryl Alcohol and 2-Furyl Methyl Ketone by Sister Chromatid Exchange (SCE) Analysis

Genotoxicity of furfuryl alcohol and 2-furyl methyl ketone is evaluated by sister chromatid exchange analysis (SCEs) in mouse bone marrow system using standard technique. Three doses of the compounds ranging from 1000-4000 ppm were given by oral gauvage to experimental animals for one day in single dose series and for five consecutive days in multiple dose series to simulate a human situation. 5-Bromodeoxyuridine (5-Brdu) was intraperitoneally injected at hourly intervals for 6 hr at a concentration of 0.04mg/g body weight. In both single and multiple dose series the animals were exposed to 5-Brdu for 26 hr and were sacrificed after 12, 24 and 48 hr following last feeding. Control animals received distilled water and same concentration of Brdu. Bone marrow slides were prepared and stained as per standard procedure. 100 metaphase spreads in second mitotic division were scored from each dose and period and subjected to statistical analysis. There was significant induction of SCEs in both single and multiple dose series with both the compounds. It was found to be dose dependant and the period dependancy showed a decline after 24 hr.

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.

Conditional deletion of Nbs1 in murine cells reveals its role in branching repair pathways of DNA double-strand breaks

NBS1 forms a complex with MRE11 and RAD50 (MRN) that is proposed to act on the upstream of two repair pathways of DNA double-strand break (DSB), homologous repair (HR) and non-homologous end joining (NHEJ). However, the function of Nbs1 in these processes has not fully been elucidated in mammals due to the lethal phenotype of cells and mice lacking Nbs1. Here, we have constructed mouse Nbs1-null embryonic fibroblasts and embryonic stem cells, through the Cre-loxP and sequential gene targeting techniques. We show that cells lacking Nbs1 display reduced HR of the single DSB in chromosomally integrated substrate, affecting both homology-directed repair (HDR) and single-stranded annealing pathways, and, surprisingly, increased NHEJ-mediated sequence deletion. Moreover, focus formation at DSBs and chromatin recruitment of the Nbs1 partners Rad50 and Mre11 as well as Rad51 and Brca1 are attenuated in these cells, whereas the NHEJ molecule Ku70 binding to chromatin is not affected. These data provide a novel insight into the function of MRN in the branching of DSB repair pathways.

Occupational exposure in airport personnel: Characterization and evaluation of genotoxic and oxidative effects

Airport personnel can be exposed to several polycyclic aromatic hydrocarbons (PAHs) from jet fuel vapours, jet fuel combustion products and diesel exhaust. The aim of this study was to characterize the exposure and to evaluate genotoxic and oxidative effects in airport personnel ( n = 41) in comparison with a selected control group ( n = 31). Environmental monitoring of exposure was carried out analysing 23 PAHs on air samples collected from airport apron, airport building and terminal/office area during 5 working days. The urinary 1-hydroxy-pyrene (1-OHP) following 5 working days, was used as biomarker of exposure. Genotoxic effects and early direct-oxidative DNA damage were evaluated by micronucleus (MN) and Fpg-modified comet assay on lymphocytes and exfoliated buccal cells, and by chromosomal aberrations (CA) and sister chromatid exchange (SCE) analyses. For comet assay, tail moment (the product of comet relative tail intensity and length) values from Fpg-enzyme treated cells (TMenz) and from untreated cells (TM) were used as parameters of oxidative and direct DNA damage, respectively. We found 27,703 μg/m 3 total PAHs in airport apron, 17,275 μg/m 3 in airport building and 9,494 μg/m 3 in terminal/office area. Urinary OH-pyrene did not show differences between exposed and controls. The exposed group showed a higher mean value of SCE frequency in respect to controls (4.6 versus 3.8) and an increase (1.3-fold) of total structural CA in particular breaks (up to 2.0-fold) and fragments (0.32% versus 0.00%), whereas there were no differences of MN frequency in both cellular types. Comet assay evidenced in the exposed group a higher value in respect to controls of mean TM and TMenz in both exfoliated buccal cells (TM 118.87 versus 68.20, p = 0.001; TMenz 146.11 versus 78.32, p < 0.001) and lymphocytes (TM 43.01 versus 36.01, p = 0.136; TMenz 55.86 versus 43.98, p = 0.003). An oxidative DNA damage was found, for exfoliated buccal cells in the 9.7% and for lymphocytes in the 14.6% of exposed in respect to the absence in controls. Our findings furnish a useful contribution to the characterization of civil airport exposure and suggest the use of comet assay on exfoliated buccal cells to assess the occupational exposure to mixtures of inhalable pollutants at low doses since these cells represent the target tissue for this exposure and are obtained by non-invasive procedure.

The alteration of sister chromatid exchange frequencies in Behçet's disease with and without HLA‐B51

The analysis of sister chromatid exchange (SCE) is a cytogenetic technique used to show DNA damage as a result of an exchange of DNA fragments between sister chromatids. It is known that there is an increased SCE frequency in Behçet's disease (BD). To investigate whether human leucocyte antigen (HLA)-B51-positive patients with Behçet's disease exhibit higher SCE frequencies than those without HLA-B51. Lymphocytes from 75 patients (38 women, 37 men) and from 50 controls (28 women, 22 men) were cultured in darkness for 72 h in the presence of bromodeoxyuridine. Metaphase chromosomes were stained with a fluorescence plus Giemsa technique after a standard harvest procedure. For HLA-B51 typing, DNA was extracted from ethylenediaminetetraacetic acid blood samples and HLA-B5 allele genotyping was performed by the polymerase chain reaction (PCR)-sequence specific primer method. Thirty-nine of 75 patients with BD (52%) and 15 of 50 controls (30%) were found HLA-B51-positive. The SCE frequencies in HLA-B51-positive patients were higher than in HLA-B51-negative ones (P < 0.001), whereas no difference was detected in the control group. This study revealed that there was a significant association between elevated SCE frequencies and existence of HLA-B51 patients with BD.

Sister chromatid exchange analysis in smelting plant workers exposed to arsenic

There are many studies documenting the genotoxic effects of environmental exposure to arsenic. Nevertheless, few data are available on the genotoxic risks of occupational arsenic exposure. In the present study, we have evaluated whether or not occupational exposure to arsenic in a copper smelting plant results in a significant increase in the frequency of sister chromatid exchange (SCE). SCE frequencies, proliferation rate index (PRI), and high frequency cells (HFCs) were evaluated in peripheral blood lymphocytes from a group of 105 arsenic-exposed workers from a Chilean smelting plant (exposed group). Similar assays were conducted on a group of 55 workers employed at the same mine but involved in administrative jobs (internal control), and on 48 workers of another mine, with no significant levels of arsenic (external control). Small but significant increases in SCE frequency were observed in the arsenic-exposed workers compared with the external control group (6.28+/-0.09 vs. 5.84+/-0.14 SCE/cell; P<0.01). Also, significantly higher frequencies of HFCs were observed in the exposed group (2.21%+/-0.20%) than in either the external control group (1.20+/-0.23; P=0.002) or the internal control group (1.30+/-0.24; P=0.008). However, there was no relationship between arsenic levels in the urine of the subjects and SCE or HFC frequency. The results of the study indicate that copper smelting results in slightly increased levels of DNA damage. However, our data were not consistent with arsenic exposure being the cause of the increase.

Cell cycle kinetics and cytogenetic changes in human lymphocytes exposed to oestradiol in vitro

Metabolic conversion of oestrogen phenolic groups may create conditions of oxidative stress accompanied by damage of cellular macromolecules including DNA. The aim of this investigation was to evaluate the cell cycle kinetics and possible cytogenetic changes in cultured human peripheral blood lymphocytes exposed to seven experimental concentrations of 17β-oestradiol (range 10-10 M to 10-4 M). Cell cycle kinetics was analyzed on metaphase spreads prepared for a standard analysis of sister-chromatid exchanges (SCEs) stained by fluorescent-plus-Giemsa (FPG) technique. Cytogenetic changes were monitored by analysis of chromosome damage (gaps and breaks), structural and numerical aberrations. On the basis of the obtained results it can be concluded that oestradiol has no significant influence on cell cycle kinetics and mitotic index of cultured human lymphocytes. However, at estradiol concentration of 7×10-6 M, and at higher concentrations used in this experiment, there was a significant increase of gaps, breaks and aneuploidies. On the other hand, oestradiol treatment has not changed the frequency of polyploid cells. Therefore, it can be concluded that high concentrations of oestradiol pose some genetic risk detectable at cytogenetic level.

Sister Chromatid Exchange and Oxidative DNA Damage in Paving Workers exposed to PAHs

Paving workers are exposed during road paving to several polycyclic aromatic hydrocarbons (PAHs) contained in asphalt fumes. In this study early genotoxic and oxidative effects of exposure to bitumen fumes were evaluated in 19 paving workers and 22 controls. Environmental and biological monitoring of exposure was carried out, measuring, on personal air samples from exposed workers collected during three working days, the concentration of 14 PAHs and urinary OH-pyrene at the end of each of the three working days. Genotoxic effect was evaluated analysing sister chromatid exchange (SCE) frequency and direct-oxidative DNA damage by formamido-pyrimidine-glycosylase (Fpg)-modified comet assay on lymphocytes. Tail moment values from Fpg-enzyme treated cells (TMenz) and from untreated cells (TM) were used as parameters of direct and oxidative DNA damage, respectively. For each subject, the TMenz/TM ratio >2.0 was used to indicate the presence of oxidative damage. DNA damage was also evaluated analysing comet percentage. Personal air samples showed low level of total PAHs (2.843 microg m(-3)) with prevalence of 2-3 ring PAHs (2.693 microg m(-3)). Urinary OH-pyrene after work-shift of the three working days was significantly higher than that found at the beginning of the working week. SCE analysis did not show any difference between two groups while an oxidative DNA damage was found in 37% of exposed with respect to the absence in controls. Comet percentage was significantly higher (P = 0.000 ANOVA) in the exposed than in controls. The results demonstrate the high sensitivity of comet assay to assess early oxidative effects induced by exposure to bitumen fumes at low doses and confirm the suitability of urinary OH-pyrene as a biomarker of PAH exposure. In conclusion the study suggests the use of Fpg-modified comet test as a biomarker of early genotoxic effects and that of urinary OH-pyrene as a biomarker of PAH exposure to furnish indications in terms of characterization, prevention and management of risk in occupational exposure to mixtures of potentially carcinogenic substances.