Cytogenetic genotoxicity of amoxicillin

Amoxicillin (AMO), a drug used in the treatment of infections caused by susceptible bacteria, has been evaluated for its ability to induce genotoxicity in human peripheral blood lymphocytes. The potential genotoxic effects of AMO were investigated in vitro by the sister chromatid exchange (SCE), chromosomal aberration (CA), and micronucleus (MN) tests. The cells were treated with 400, 600, 800, and 1,000 μg/ml AMO in the presence and absence of a metabolic activator (S9 mix), respectively. In this study, AMO did not induce SCEs or CAs in human peripheral blood lymphocytes both in the presence and absence of the metabolic activator. AMO concentration‐dependently decreased the proliferation index (PI) in the absence of the metabolic activation for 24‐hr treatment period. Mitotic index (MI) was generally found to have been reduced when compared with the negative control but not with the solvent control in cultures treated with AMO for 24 hr. AMO did not decrease the PI and MI in the presence of the metabolic activator. Furthermore, AMO neither induced the formation of MN nor decreased the nuclear division index in human peripheral blood lymphocytes both in the presence and absence of the metabolic activator. According to the present results, we suggest that AMO does not pose genotoxic risk for patients who are under therapy against bacterial infections. Environ. Mol. Mutagen., 2010. © 2009 Wiley‐Liss, Inc.     

Cytogenetic damage in female Pakistani agricultural workers exposed to pesticides

Bhawalpur is a major cotton-growing area in Pakistan. Cotton picking in Pakistan is carried out by females and as a result of the intensive use of pesticides during the growing season these females are exposed to pesticide residues in the picking season. In the present study, peripheral blood was obtained from 69 cotton pickers and 69 unexposed females and used to assess the effect of pesticide exposure on genetic damage as well as on hepatic enzymes and serum cholinesterase. The subjects were of similar average age in workers and control groups (37.55 +/- 12.75 vs. 37.52 +/- 13.47, P > 0.05). Average exposure time of the picker females was 10.26 +/- 6.14 years. Subjects from the exposed group did not use any protective measures during their work activities. Levels of serum cholinesterase were lower, and levels of alkaline phosphatase, alanine aminotransferase, and aspartate aminotransferase were higher in the exposed workers as compared with the control group (P < 0.001). The exposed group exhibited significantly increased frequencies of binucleated cells with micronuclei (12.72 +/- 3.48 vs. 4.35 +/- 2.44, P < 0.001) and total number of micronuclei in binucleated lymphocytes (16.51 +/- 4.27 vs. 5.86 +/- 3.09, P < 0.001) in comparison with subjects of the control group. The binucleated cells with micronuclei frequency also seemed to increase with age in both the groups, however, the magnitude of increase was greater in exposed group than the control. Results from the present study indicate that occupational exposure to pesticide mixtures results in cytogenetic damage in exposed females.     

烟草和酒精诱导人淋巴细胞SCE和微核的研究

目的　研究烟草和酒精共同作用 ,观察其对人体是否存在有害的叠加效应。方法　采用人全血培养淋巴细胞染色体SCE和微核在酒精和烟草酒精提取物作用下的改变。结果　酒精组与对照组相比 ,SCE频率增高 (P <0 0 5 ) ,各烟草处理组SCE高于酒精组 ,与对照组相比更高 ,存在显著差异 (P <0 0 1)。微核在酒精组与对照组相比 ,显著提高。结果　酒精与烟草之间存在叠加效应。     

Cytogenetic biomonitoring on a group of petroleum refinery workers

Workers employed in petroleum refineries are exposed to a wide range of toxic compounds (benzene, polycyclic aromatic hydrocarbons, heavy metals, etc.) with known mutagenic and carcinogenic potential. In this study, we investigated by using the cytokinesis block micronucleus (CBMN) assay on human peripheral blood lymphocytes (PBL) whether general occupational exposure in petroleum refineries resulted in early biological effects, which would be indicative of adverse health effects in the long term. In this study, out of more 500 workers enrolled in the study, 79 male subjects (46 nonsmokers and 33 smokers), employed in two different Italian petroleum refineries, and a total of 50 male control subjects (34 nonsmokers and 16 smokers) were selected by using very strict selection criteria. The comparison of chromosome damage in PBL between exposed and control populations pointed out a significant increase of micronuclei in the exposed group, correlated with the length of employment. Results confirm that smoking is the principal confounding factor for the responses. In conclusion, our results are indicative of a potential genotoxic risk related to the complex occupational exposure in petroleum refineries, despite the measures adopted in the plants, and corroborate the need to increase safety measures to avoid exposure to chemical agents.     

Increased chromosome damage in pediatric heart catheterization patients after diagnostic fluoroscopy and cineangiography

Chromosome damage (CD) and sister chromatid exchange (SCE) levels were studied in lymphocytes from 30 pediatric heart catheterization patients receiving radiation during diagnostic fluoroscopy and cineangiography procedures. Forty-eight-hour CD and 72-hr SCE cultures were prepared from sequential samples taken from each patient: samples 1-3 via the catheter the same day (1) before exposure, (2) after fluoroscopy, and (3) after cineangiography; and sample 4 by venipuncture the next morning. Significant increases in CD (dicentrics, rings, and fragments), but not SCE, were observed. From a mean base level of 0.4% cells with CD, the CD levels increased 2-3-fold in samples 3 and 4 (p = .001). Rings only occurred in samples 3 and 4. While increased CD levels also correlated with increasing age, body surface area, and weight, partial correlations controlling for these factors clearly indicate that the CD effects are principally attributable to the radiological procedures (p = .001). Increased CD levels correlated with both the roentgen dose of cineangiography exposure (p = .002) and the volume of contrast medium (p = .000); however, partial correlations, controlling for either factor, indicate that the contrast medium was the principal factor (p = .006).     

Cytogenetic damage in female Chilean agricultural workers exposed to mixtures of pesticides

The VIII Region of Bio-Bio is a major fruit-growing area of Chile that makes intensive use of agricultural pesticides. The cytogenetic damage associated with exposure to mixtures of pesticides was evaluated by comparing peripheral blood lymphocyte micronucleus (MN) frequencies in a group of 64 female agricultural workers and 30 female controls. The exposed subjects worked during the spring and summer in thinning and pruning fruit trees and in harvesting and packing different fruits, such as raspberries, grapes, apples, and kiwis. They did not use any protective measures during their work activities. A significant increase in the frequency of binucleated cells with micronuclei (BNMN) was found in the exposed women as compared with the controls (36.94 +/- 14.47 vs. 9.93 +/- 6.17 BNMN/1000 BN cells; P < 0.001). The frequency of BNMN varied as a function of age in both the exposed and control groups, but no correlation was found between BNMN frequency and the duration of exposure. Also, smoking and other habits had no effect on MN frequency. Our study confirms that occupational exposure to pesticide mixtures results in cytogenetic damage.     

Cytogenetic effects of shale-derived oils in murine bone marrow

The cytogenetic effects of exposure of mice to shale-derived oils by either skin painting or intraperitoneal injection were examined. Skin painting with 40 mg crude oil every other day for five weeks had no effect on the frequency of chromosomal aberrations observed in the bone marrow. Three daily intraperitoneal injections of 0.5–2.0 ml/kg per day of a crude shale oil from an aboveground retort induced a dose-related increase in the frequency of aberrations observed. A hydrotreated sample of this oil produced a similar pattern of aberration induction, but at lower frequencies. Crude shale oil from a modified in situ retort induced the highest frequency of chromosomal aberrations at the 0.5 ml/kg dose; lower frequencies were induced at the 1.0 ml and 2.0 ml/kg doses. Of the three shale oils tested, only the crude oil from the aboveground retort induced increased frequencies of sister chromatid exchanges and only at doses that also induced structural aberrations. These studies indicate that structural aberration analysis is a more sensitive test than sister chromatid exchange analysis for the type of DNA damage induced by shale-derived oils in murine bone marrow cells.     

The genotoxic effect of potassium metabisulfite using chromosome aberration, sister chromatid exchange, micronucleus tests in human lymphocytes and chromosome aberration test in bone marrow cells of rats

Potassium metabisulfite (PMB) is used as an antimicrobial substance in many kinds of foods. In the present study, the effects of PMB on chromosome aberrations (CAs), sister chromatid exchanges (SCEs), and micronucleus (MN) formation in human lymphocytes and as well as its effect on CAs in bone marrow cells of rats were investigated. The human lymphocytes were treated with 25, 50, 100, and 200 microg/ml of PMB for 24 and 48 hr. PMB was also intraperitoneally (ip) injected to the rats as a single dose of 150, 300, and 600 mg/kg body weight (b.w.) for 12 and 24 hr before sacrifice. PMB induced abnormalities such as structural and numerical (total) CAs, SCEs, and MN formations in a dose dependent manner in the lymphocytes of the 24- and 48-hr treatment periods. In addition, PMB showed a cytotoxic effect by decreasing the replication index (RI), mitotic index (MI) and nuclear division index (NDI) in a dose dependent manner in human lymphocytes. The compound induced CA as well and decreased the MI in bone marrow cells of rats. It might be concluded that PMB had a high genotoxic and cytotoxic risk.     

Follow-up biological and genotoxicological monitoring of acrylonitrile- and dimethylformamide-exposed viscose rayon plant workers

In order to investigate the genotoxic effects of occupational acrylonitrile (ACN) and dimethylformamide (DMF) exposures, clinical serum and urine parameters and genotoxicological endpoints such as chromosome aberration (CA), sister chromatid exchange (SCE), high frequency SCE (HFC), cell cycle kinetics, and UV-induced unscheduled DNA synthesis (UDS) were followed up three times during a 20-month period in peripheral blood lymphocytes (PBL) of 26 workers (13 maintainers and 13 fiber producers) occupationally exposed to ANC and/or DMF in a viscose rayon plant, 26 matched control subjects, and six industrial controls (all males). Six of the 26 exposed subjects were hospitalized because of liver dysfunction that had developed due to inhalative DMF exposure. The rate of smoking was estimated on the basis of serum thiocyanate (SCN) levels. Average peak air ACN and DMF concentrations were over the maximum concentration limits at the time of both investigations. Urine ACN and monomethyl-formamide (MMF) excretions of the exposed subjects were almost doubled after work shifts. An increase in lymphocyte count (in months 0 and 7), and severe alterations in the liver function were observed in the exposed subjects. In PBLs the proliferative rate index (PRI) was already increased in month 0 compared with the controls. In each study, significant increases in CA and SCE frequencies, as well as increases in UDS were found in PBLs of the exposed subjects. The frequencies of chromatid breaks and acentric fragments further increased in month 7 and remained constantly elevated in month 20. Increased yields of both chromatid and chromosome-type exchange aberrations first appeared in month 20, when HFCs were 2.72 times more frequent in fiber producers than in maintainers. The role of some important biological confounding factors (age, white blood cell count, and hematocrit) and lifestyle confounding factors (smoking and drinking habits) were subjected to an analysis of variance during the second study. Increased CA, SCE, and UDS were found both in control and exposed smokers when current smoking was established on the basis of the serum SCN levels. The cytogenetic data suggest that occupational exposures to ACN and DMF induce considerable genotoxic consequences and may increase the cancer risk in the exposed human populations. Environ. Mol. Mutagen. 31:301–310, 1998 © 1998 Wiley-Liss, Inc.     

Induction of polyploidy in human lymphocytes in vitro by excess adenine,but not by adenosine

It is known that high levels of DNA precursors can be both clastogenic and mutagenic in cultured cell lines and in vivo. The purpose of the present study was to examine at an observational level the cytogenetic effects of adenine and adenosine in primary human cell cultures. Human peripheral blood lymphocytes from four donors were cultured and treated with a range of concentrations of adenine and adenosine. Although no increase in sister chromatid exchange (SCE) frequency was observed with either compound, there was a statistically significant, dose-related increase in the proportion of polyploid cells in cultures treated with adenine, but not in those treated with adenosine. Some of the polyploid metaphases found after adenine treatment contained diplochromosomes, suggesting that endoreduplication might have been involved in polyploid formation in these cells. It is concluded that a high level of adenine can cause genetic changes in human lymphocytes by interfering with mitosis, perhaps by disturbing the balance of DNA precursor pools. © 1995 Wiley-liss, Inc.     

Sister chromatid exchange with and without in vitro mutagen induction in cultured skin fibroblasts from patients with adenomatosis of the colon and rectum

Fibroblast cell strains were obtained from skin biopsies taken from patients with adenomatosis of the colon and rectum (ACR), and their relatives. A total of 50 different fibroblast strains were tested for their frequencies of sister chromatid exchange (SCE) in vitro. These strains included nine from patients with the Gardner syndrome, 21 from patients with non-Gardner ACR, and 20 cell strains from healthy relatives who were not at an increased risk for ACR. In 23 strains, the SCE frequencies after in vitro exposure to N-methyl-N'-nitro-N-nitroso-guanidine (MNNG) were also determined. Both with and without MNNG induction, SCE values in the Gardner strains were found to be significantly higher than in the control strains (p less than 0.02 and p less than 0.03, respectively). Non-Gardner ACR strains differed only slightly from controls, thus making the difference between the control group and the pooled Gardner + non-Gardner ACR group not significant. In all groups, there was a significant increase in SCE after MNNG exposure, and those strains which had low SCE values spontaneously, also tended to have relatively moderate SCE values after MNNG induction. There was no significant difference between the ratios of SCE values with and without MNNG exposure in the different groups.     

Cytogenetic biomonitoring in traffic police workers: Micronucleus test in peripheral blood lymphocytes

Atmospheric pollution represents a relevant environmental hazard which has been associated with considerable excess mortality, morbidity, and increased rates of respiratory diseases in humans. To date, more than 3,000 environmental chemical compounds have been identified in the ambient atmosphere, including a variety of mutagenic and/or carcinogenic agents, such as polycyclic aromatic hydrocarbons (PAHs), aromatic amines, and heterocyclic compounds. Positive associations between cytogenetic markers and airborne levels of PAHs have been reported by experimental and human studies. Traffic has been implicated as the major determinant for the concentration of PAHs and, therefore, for the genotoxic activity of urban air. A biomonitoring study has been conducted in 82 Italian traffic police workers exposed to air pollutants and 34 control subjects (matched by age, gender, and smoking habits) not exposed to traffic pollutants. The aim of this study was to assess the cytogenetic effects, such as micronucleus frequency in peripheral blood lymphocytes, and to estimate the association with individual exposure to PAH. Statistical analysis of the frequency of micronuclei in binucleated cells showed higher mean levels in referent subjects (4.03%) than in traffic police officers (3.73%). Smoking showed no effect on the frequency of micronuclei. The study failed to detect any association between micronucleus frequency and individual level of benzo(a)pyrene, considered a marker of exposure to PAHs. These findings indicate that exposure to urban air pollutants does not result in increased levels of micronuclei in peripheral white blood cells. Environ. Mol. Mutagen. 30:396–402, 1997 © 1997 Wiley-Liss, Inc.     

Confirmation of the chromosome damaging effects of lamivudine in in vitro human peripheral blood lymphocytes

The aim of this study was to investigate genotoxic effects of lamivudine (an analogue of cytidine) using in vitro sister chromatid exchange (SCE), chromosome aberration (CA), and micronucleus (MN) tests in human peripheral lymphocytes. The cells were treated with 75, 100, 125, and 150 microg/ml concentrations of lamivudine (roughly 30-60 times higher than plasma levels achieved in patients receiving this drug) for two (24- and 48-hr) treatment periods. Lamivudine induced SCEs at the highest concentration (150 microg/ml) in the 24-hr treatment, and at 125 and 150 microg/ml in the 48-hr treatment, when compared to the solvent control. During both treatment periods, structural chromosome aberrations were significantly increased at 100, 125, and 150 microg/ml lamivudine concentrations. However, the increases of SCEs (22%) and CAs (50%) were weak. In addition, lamivudine reduced both the proliferation index (PI) and the mitotic index (MI) significantly at all concentrations for the two treatment periods. The MI was reduced by lamivudine in a dose-dependent manner during both the 24- and 48-hr treatment periods. In contrast, the PI was reduced by lamivudine only during the 48-hr treatment period. A weak but significant increase in MN formation was observed following lamivudine treatment at 100, 125, and 150 microg/ml for 48 hr, but no significant increase in micronuclei were observed following 24-hr treatment. In conclusion, lamivudine has a weak genotoxic effect at elevated doses on human peripheral lymphocytes.     

Genotoxic repercussion of high-intensity radiation (x-rays) on hospital radiographers

Recent technological advances in the medical field have increased the plausibility of exposing humans to high-intensity wavelength radiations like x-rays and gamma rays while diagnosing or treating specific medical maladies. These radiations induce nucleotide changes and chromosomal alterations in the exposed population, intentionally or accidentally. A radiological investigation is regularly used in identifying the disease, especially by the technicians working in intensive care units. The current study observes the genetic damages like chromosomal abnormalities (CA) in clinicians who are occupationally exposed to high-intensity radiations (x-rays) at their workplaces using universal cytogenetic tools like micronucleus assay (MN), sister chromatid exchange and comet assay. The study was conducted between 100 exposed practitioners from the abdominal scanning, chest scanning, cranial and orthopedic or bone scanning department and age-matched healthy controls. We observed a slightly higher rate of MN and CA (p < .05) in orthopedic and chest department practitioners than in other departments concerning increasing age and duration of exposure at work. Our results emphasize taking extra precautionary measures in clinical and hospital radiation laboratories to protect the practitioners.     

On the importance of DNA strand breaks as the first event to initiate sister chromatid exchange (SCE): Experiments with restriction endonucleaseBglI

The exact molecular mechanism of sister chromatid exchange (SCE) is still unknown, despite the many reports dealing with this cytogenetic end point published in the last 40 years. One point to be investigated is the nature of the original lesion(s) in DNA leading to the production of SCE. Whereas, for chromosomal aberrations, the importance of DNA double-strand breaks has been well established, there is still controversy about the relative importance of strand breaks and base modifications for triggering the process of SCE formation. In the present paper, we have taken advantage of the ability of the restriction endonucleaseBglI to induce SCE and have exploited the fact that preincubation with 2,3-butanedione results in the loss ofBglI ability to cut DNA, while it is still able to recognize its sequence in DNA and bind to it, to see whether this alone is enough to initiate SCE formation, or if a physical DNA double-strand break is required. Our results seem to support the necessity of DNA breaks for SCE production.accepted for publication by J. S. (Pat) Heslop-Harrison     

Cytogenetic effects of short- and long-term exposure of chick embryos to the phenoxyherbicide 2,4-D

Before incubation, chick embryos were treated with the herbicide 2,4-dichlorophenoxy acetic acid (2,4-D) by injecting onto the inner shell membrane solutions of 0, 0.5, 1, 2, or 4 mg 2,4-D. A commercial formulation containing 37% 2,4-D iso-octyl ester as active ingredient and pure 2,4-D were tested. Sister chromatid exchange (SCE) and cell cycle kinetics were examined at days 4, 7, and 10 from 22 to 30 embryos per group. After 4 days of exposure to commercial 2,4-D, a small (P < 0.05) dose-related increase of SCE was seen for the 4-mg group. An enhanced SCE response upon long-term exposure to 2,4-D was apparent. After 10 days of exposure, SCE frequencies for the 2- and 4-mg commercial 2,4-D, and 4-mg pure 2,4-D groups were significantly higher than for the controls. A significant slowing of cell cycle at concentrations at and above 1 mg was seen. Also observed was a slight, not statistically significant proliferative effect at the lowest dose of 0.5 mg/embryo. Consistent with the results from other test systems, the present findings indicate that 2,4-D has a low to moderate genotoxic activity.     

Failure to demonstrate an effect of in vivo diagnostic ultrasound on sister chromatid exchange frequency in amniotic fluid cells

Antepartum use of diagnostic ultrasound has markedly reduced radiation exposure of the fetus. Previous investigations have documented the safety of ultrasound, but concern persists regarding its long-term effects. As new methods become available to study possible subtle effects of ultrasound, it is important to reevaluate this technique continually because of its universal use in obstetrics and elsewhere. We report results of in vivo studies of effect of diagnostic ultrasound on the sister chromatid exchange (SCE) frequency in amniotic fluid cells. SCE is a cytogenetic phenomenon believed to be a sensitive indicator of environmental perturbations and chromosome stability. In amniotic fluid cells from six pregnancies without ultrasound exposure and in 34 pregnancies that received varying amount of ultrasound immediately before amniocentesis, there was no difference in SCE frequency in exposed versus nonexposed cells. These data, which appear to confirm again the safety of ultrasound, are reassuring to both patients and clinicians.     

Factors underlying variation in spontaneous and clastogen-induced sister chromatid exchanges and chromosome breakage frequencies.

The latent "factors" influencing spontaneous and clastogen-induced genetic damage, measured by rates of sister chromatid exchange (SCE) and chromosome breakage (CB), were investigated in a small sample of 20 unrelated, healthy individuals. The covariation of spontaneous and clastogen-induced (bleomycin [BLM], streptonigrin [SN], mitomycin-C [MMC], 4-nitroquinoline-1-oxide [4NQO]) SCEs and CBs was analyzed by maximum-likelihood factor analysis. A single-factor model resulted in large standardized regression coefficients of measured variables on the factor for spontaneous and BLM- and SN-induced SCE frequencies, and a modest regression coefficient for MMC-induced SCEs. A two-factor model, after varimax rotation, yielded one factor strongly associated with spontaneous and BLM- and SN-induced SCE frequencies, and a second factor associated with spontaneous and BLM- and SN-induced CBs. A bootstrap analysis of this data set indicated the statistical significance of one regression coefficient (i.e., P less than or equal to 0.05) and borderline significance (0.07 less than or equal to P less than or equal to 0.11) of three other regression coefficients on the first factor, to be interpreted as an effector of SCE frequencies. However, for the second factor, none of the bootstrapped regression coefficients was significant (P greater than 0.22). Due to the modest sample utilized in this study, the validity of this model should be further explored using additional, larger data sets.     

Cytogenetic and chemical detection of human exposure to polyhalogenated aromatic hydrocarbons

Peripheral lymphocytes from Taiwanese women (n = 35) exposed to polychlorinated aromatic hydrocarbons and from matched controls (n = 24) were assessed for the levels of sister chromatid exchanges (SCEs) after a 72-hour incubation of whole blood in the presence or absence of alpha-naphthoflavone (ANF) and for chromosome aberrations after 48 hours of incubation. Serum levels of polychlorinated biphenyl (PCB) congeners were measured for all individuals, and serum levels of several polychlorinated dibenzofurans (PCDFs) were measured for 12 exposed individuals by gas chromatography-mass spectometry. Blood concentrations of total PCBs in the exposed population averaged approximately 15 ppb, whereas mean PCDF values were 14 ppt. Major PCB congeners detected were 2,2' 4,4', 5,5'-hexa CB and 2,2'3,4,4',5-hexa CB. PCDFs detected were primarily 1,2,3,4,7,8,-hexachlorodibenzofuran (10.8 ppt) and 2,3,4,7,8-pentachlorodibenzofuran (2.7 ppt). Average SCE frequencies were 7.61 for controls and 7.30 for exposed individuals when assays were conducted in the absence of ANF, whereas respective values were 8.85 and 10.75 in the presence of ANF. Differences in the level of ANF-induced SCEs between the two populations were highly significant (P less than .001). Moreover, the ANF-induced SCEs were highly correlated with the serum concentrations of total PCBs and of several PCB congeners (P less than .001). Increases in ANF-induced SCEs appeared to be linear up to a PCB concentration of approximately 30 ppb. Chromosome aberration frequencies were similar in control and exposed populations. These studies demonstrate that in vivo exposure to PCBs and PCDFs result in an enhanced sensitivity of lymphocytes to the SCE-causing actions of ANF.     

Preparation of mouse bone marrow primary cultures for sister chromatid exchange and chromosomal aberration studies

Summary A procedure for preparing and culturing mouse bone marrow cells for cytogenetic studies is described. Animals are killed by cervical dislocation, the bone marrow is flushed from femora and tibia with Ham's F12 medium into centrifuge tubes. Bone marrow cells are collected by low-speed centrifugation (285 ×g). One million cells are suspended in a 30-ml Falcon flask with complete medium containing 3.45 ml Ham's F12 medium, 1 ml fetal bovine serum, 0.05 ml penicillin-streptomycin, and 0.5 ml whole uterus extract from pregnant mice. For sister chromatid exchange analysis, 20 µM of 5-bromo-2 -deoxyuridine is also added to the culture medium for 24 to 28 h. The cell cycle is approximately 12 to 14 h, in culture. This culture system can also be used for chromosomal aberration studies.