Does exhaustive exercise result in oxidative stress and associated DNA damage in the chub (Leuciscus cephalus)?
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Abstract DNA strand breaks [as determined by the conventional and formamidopyrimidine glycosylase (FPG)‐modified Comet assay] and antioxidant defense status [as indicated by superoxide dismutase (SOD) activity and reduced glutathione (GSH) concentration] were evaluated in healthy adult chub ( Leuciscus cephalus ) after exhaustive exercise [swimming to their critical swimming speed ( U crit ), twice in succession with a 40 min rest period between] vs. confined (unexercised) control fish. The conventional Comet assay revealed significantly higher DNA strand breaks in all the tissues (blood, liver, and gill), with the highest increase over background evident in the epithelial gill cells of swum fish compared to the controls. Moreover, when the FPG‐modified Comet assay was conducted to reveal specific oxidative lesions, the gill cells of exercised fish sustained the highest level of oxidative DNA damage in comparison to the control. Data on tissue antioxidant defense mechanism were less conclusive, with no significant differences in the tissue levels of SOD or GSH. This suggests that either the degree of oxidative stress was not great enough to evoke a response in terms of defense mechanisms or the timescale of antioxidant defense response was somewhat different from the time between the application of stress and subsequent tissue sampling. From the swimming data, U crit was significantly lower on the second assessment compared to the first (repeat ratio: 0.76), suggesting that the fish were exercised to a level which was not sustainable. Overall, these findings support the theory that acute extreme exercise could result in oxidative stress and associated DNA damage in fish. These observations suggest that fish living in fast flowing and polluted rivers are at increased risk of DNA damage. Environ. Mol. Mutagen., 2006. © 2006 Wiley‐Liss, Inc.Keywords:
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Abstract Background: Smoking has been shown to have a positive effect on DNA damage in almost all the cells of the body. Quantitative analysis of this damage will help in assessing the etiopathogenesis of various nicotine induced damage to the body. Comet assay has been an emerging tool in this regard and hence was applied by us to estimate the severity of DNA damage in smokers. Aims & Objectives: To evaluate the DNA genotoxicity in peripheral blood lymphocytes in smokers and their comparison with non smokers & assess the quantitative damage. Materials and methods: 30 smokers & 20 non smokers were recruited & their peripheral blood was taken for the comet assay to look for Olive moment & Tail moment to quantitatively assess the DNA damage due to cigarette smoking. Results: In our study there was no significant difference in the analysis of DNA damage (with regard to tail moment & olive moment) in smokers versus non smokers (P value: more than 0.05). Conclusions: Though smoking is known to cause DNA damage, we did not find significant differences between the two groups probably due to other multifactorial etiologies for genotoxicity.
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Currently, humans and other living things can have many genotoxic damages due to reasons such as chemicals, drugs, unfavorable environmental effects. Breaks in the DNA structure due to genotoxic damage can cause mutations, changes in chromosome structure and cancer. Comet assay, also known as a single cell gel electrophoresis (SCGE), is a sensitive, rapid and cheap method used to measure DNA damage and repair at the individual cell level. With this method, cells are called this way because of they appear as comets under a microscope. DNA damage and repair can be detected in each cell type that can be obtained as single cell suspension, with comet assay. In addition to the direct determination of DNA damage in a single cell, it is possible to determine whether all cells in a population suffer from the same amount of damage. Furthermore, this assay may help to predict the heterogeneous response of cells during any treatment, and the tumor response in radiotherapy and chemotherapy treatment protocols. Especially in human biomonitoring studies, it is also used in the investigation of DNA damage in people who exposed to various occupational or accident-related environmental or workplace-related agents. In this review, a general overview of the comet assay and its currently applications to determine the genotoxicity of environmental factors and chemicals are discussed. Key words : DNA damage, DNA repair, genotoxicity, comet assay, single cell gel electrophoresis. DOI : 10.7176/JSTR/5-5-02
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In order to determine background levels of DNA strand breaks, we examined 80 healthy individuals by comet assay considering age, sex, and smoking as confounding factors. Only age was found to have a significant effect on basal levels. One thousand cells of each donor were graded by eye into 5 categories according to the amount of DNA in the tail: classification group A (no damage) <5%, B (low damage) 5-20%, C (medium damage) 20-40%, D (high damage) 40-95%, and group E (total damage) >95%. The interpretation of the comet assay was modified to achieve a tail factor, which represents the DNA damage of 1000 scored cells as a single number, without the need of an image analysis software package. Hydrogen peroxide and bleomycin used for in vitro exposure of lymphocytes, produced clear dose-related responses in the comet assay. Our data encourage the application of the used classification model for a sensitive and fast quantification of DNA damage. Results in this study are in agreement with most of the earlier investigations.
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The aim of this study was to investigate the association between DNA damage and blood lead levels in individuals occupationally exposed to lead. To evaluate this association, 61 workers exposed to lead were monitored in terms of DNA damage in blood lymphocytes. The levels of DNA damage were measured according to 3 comet assay parameters, including tail intensity (TI), tail moment (TM), and DNA tail (DNAt). A statistically significant positive correlation was found between the lead levels and TI, TM, and DNAt (p <.01). Smoking had independent effects on DNA damage. A statistically significant difference was observed between smokers and nonsmokers in regards to DNA damage parameters (p <.05). In addition, the lead and DNA damage levels in smokers were found to be significantly higher than the levels observed in nonsmoking workers (p <.05). Our results show that exposure to lead induces genotoxic effects in peripheral lymphocytes, as measured by comet assays.
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JS-118 is an extensively used insecticide in China. The present study investigated the genotoxic effect of JS-118 on whole blood at 24, 48, 72 and 96 h by using alkaline comet assay. Male Kunming mice were given 6.25, 12.5, 25, 50 and 100 mg/kg BW of JS-118 intraperitoneally. A statistically significant increase in all comet parameters indicating DNA damage was observed at 24 h post-treatment ( p < 0.05). A clear concentration-dependent increase of DNA damage was revealed as evident by the OTM (arbitrary units), tail length (µm) and tail DNA (%). From 48 h post-treatment, a gradual decrease in mean comet parameters was noted. By 96 h of post-treatment, the mean comet tail length reached control levels indicating repair of damaged DNA. This study on mice showed different DNA damage depending on the concentration of JS-118 and the period of treatment. The present study provided further information of the potential risk of the genetic damage caused by JS-118.
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To assess the possible applicability of comet assay in the evaluation of DNA damage caused by ionizing radiation. The alkaline comet assay or single-cell gel electrophoresis has been used as a standard method for measuring and analyzing DNA damage.Peripheral blood samples were collected from papillary thyroid cancer patients who received 131I by oral administration. Blood samples were taken just before the treatment, on the first day of treatment, and 1 week posttreatment. To determine the radiation-induced DNA damage, alkaline comet assay was performed.It was found that significantly high levels of DNA damage occurred in first day samples when compared to control samples according to tail moment measurements. Also, a decrease in the level of damage was observed in the 1-week samples.Our observations and data confirmed that treatment with 131I for papilloma thyroid cancer can cause DNA damage in circulating lymphocytes, and the comet assay seemed suitable to assess the effect of radioactive iodine for the patients.
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Radon-induced biological effects have been studied mainly through epidemiological investigations, and well-controlled in vitro and in vivo experiments. To provide data explaining radon exposure-induced harmful effects in natural environment, exposure assessment under these conditions is needed. The objective of the study was to examine the level of genetic damage assessed with biomarkers of DNA single- and double-strand breaks (SSBs and DSBs) in peripheral blood mononuclear cells obtained from individuals continuously exposed to Rn in homes. Naturally elevated Rn concentrations in homes can be found in the South of Poland, in Kowary city. Measurements of expression of phosphorylated histone γH2AX was used as a marker of DNA double strand breaks. To detect DNA single and double-strand breaks and alkali labile sites, the alkaline comet assay was used. Oxidative damage of DNA was evaluated by formamidopyrimidyne (FPG)-modified comet assay. The blood was collected from 94 volunteers living in Kowary. Subjects were grouped according to their status of living in radon concentration ≥100 Bq/m3 (n = 67), and <100 Bq/m3 (n = 27). The statistically significant differences in levels of DNA damage in peripheral lymphocytes assessed with comet assay were found to be associated with levels of radon exposure in indoor air (p = 0.034). DNA damage in the comet assay was significantly correlated with DNA damage assessed with γH2AX staining. Results of the present study indicate the suitability of alkaline comet assay for the detection of DNA damage in peripheral blood lymphocytes of people environmentally exposed to radon.
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Abstract Rickshaw pullers (RPs) engage in strenuous physical activity and are exposed to the air pollutants found in urban environments. Air pollutants and the reactive oxygen species generated by the physical activity both potentially can damage DNA. In the present study, the Comet assay, a sensitive tool for measuring DNA damage in single cells, was used to study genomic DNA damage in lymphocytes of Indian RPs. The study evaluated DNA damage in 118 healthy male volunteers, including 63 RPs whose work demanded high levels of physical activity for 7–9 hr/day, and 55 controls matched for age, habits, socio‐economic status, and exposure to air pollution. A significant increase was found for the mean Olive tail moment (arbitrary units) among the RPs (4.13 ± 0.11; P < 0.001) in comparison with the controls (3.21 ± 0.10). Likewise, comet tail length (μm) (RPs: 58.98 ± 1.01 vs. controls: 52.38 ± 1.24) and tail DNA (%) (RPs: 13.52 ± 0.31 vs. controls: 10.04 ± 0.24) were also significantly higher for RPs compared with those of their matched controls (both, P < 0.001). To our knowledge, this is the first demonstration that physical activity due to occupation can produce DNA damage in peripheral lymphocytes. Environ. Mol. Mutagen., 2006. © 2005 Wiley‐Liss, Inc.
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