scholarly journals In Vivo Alkaline Comet Assay and Enzyme-modified Alkaline Comet Assay for Measuring DNA Strand Breaks and Oxidative DNA Damage in Rat Liver

10.3791/53833 ◽  
2016 ◽  
Author(s):  
Wei Ding ◽  
Michelle E. Bishop ◽  
Lascelles E. Lyn-Cook ◽  
Kelly J. Davis ◽  
Mugimane G. Manjanatha
Keyword(s):  
Dna Damage ◽  
Comet Assay ◽  
Rat Liver ◽  
Oxidative Dna Damage ◽  
Dna Strand Breaks ◽  
Alkaline Comet Assay ◽  
Strand Breaks ◽  
Dna Strand

scholarly journals Cylindrospermopsin-Microcystin-LR Combinations May Induce Genotoxic and Histopathological Damage in Rats

Toxins ◽  
2020 ◽  
Vol 12 (6) ◽  
pp. 348 ◽  
Author(s):  
Leticia Díez-Quijada ◽  
Concepción Medrano-Padial ◽  
María Llana-Ruiz-Cabello ◽  
Giorgiana M. Cătunescu ◽  
Rosario Moyano ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Dna Damage ◽  
Oxidative Dna Damage ◽  
Fatty Degeneration ◽  
Dna Strand Breaks ◽  
Histopathological Study ◽  
Strand Breaks ◽  
Dna Strand

Cylindrospermopsin (CYN) and microcystins (MC) are cyanotoxins that can occur simultaneously in contaminated water and food. CYN/MC-LR mixtures previously investigated in vitro showed an induction of micronucleus (MN) formation only in the presence of the metabolic fraction S9. When this is the case, the European Food Safety Authority recommends a follow up to in vivo testing. Thus, rats were orally exposed to 7.5 + 75, 23.7 + 237, and 75 + 750 μg CYN/MC-LR/kg body weight (b.w.). The MN test in bone marrow was performed, and the standard and modified comet assays were carried out to measure DNA strand breaks or oxidative DNA damage in stomach, liver, and blood cells. The results revealed an increase in MN formation in bone marrow, at all the assayed doses. However, no DNA strand breaks nor oxidative DNA damage were induced, as shown in the comet assays. The histopathological study indicated alterations only in the highest dose group. Liver was the target organ showing fatty degeneration and necrotic hepatocytes in centrilobular areas, as well as a light mononuclear inflammatory periportal infiltrate. Additionally, the stomach had flaking epithelium and mild necrosis of epithelial cells. Therefore, the combined exposure to cyanotoxins may induce genotoxic and histopathological damage in vivo.

scholarly journals Repair of DNA Strand Breaks by the Overlapping Functions of Lesion-Specific and Non-Lesion-Specific DNA 3′ Phosphatases

2001 ◽  
Vol 21 (21) ◽  
pp. 7191-7198 ◽  
Author(s):  
John R. Vance ◽  
Thomas E. Wilson
Keyword(s):  
Dna Damage ◽  
Oxidative Dna Damage ◽  
Synthetic Lethality ◽  
Dna Strand Breaks ◽  
Triple Mutant ◽  
Phosphatase Domain ◽  
Strand Breaks ◽  
Alternative Pathways ◽  
Processing Enzymes ◽  
Dna Strand

ABSTRACT In Saccharomyces cerevisiae, the apurinic/apyrimidinic (AP) endonucleases Apn1 and Apn2 act as alternative pathways for the removal of various 3′-terminal blocking lesions from DNA strand breaks and in the repair of abasic sites, which both result from oxidative DNA damage. Here we demonstrate that Tpp1, a homologue of the 3′ phosphatase domain of polynucleotide kinase, is a third member of this group of redundant 3′ processing enzymes. Unlike Apn1 and Apn2, Tpp1 is specific for the removal of 3′ phosphates at strand breaks and does not possess more general 3′ phosphodiesterase, exonuclease, or AP endonuclease activities. Deletion ofTPP1 in an apn1 apn2 mutant background dramatically increased the sensitivity of the double mutant to DNA damage caused by H2O2 and bleomycin but not to damage caused by methyl methanesulfonate. The triple mutant was also deficient in the repair of 3′ phosphate lesions left by Tdp1-mediated cleavage of camptothecin-stabilized Top1-DNA covalent complexes. Finally, the tpp1 apn1 apn2 triple mutation displayed synthetic lethality in combination with rad52, possibly implicating postreplication repair in the removal of unrepaired 3′-terminal lesions resulting from endogenous damage. Taken together, these results demonstrate a clear role for the lesion-specific enzyme, Tpp1, in the repair of a subset of DNA strand breaks.

Detection of DNA strand breaks by comet assay in sputum leucocytes of bitumen-exposed workers: A pilot study

2010 ◽  
Vol 29 (9) ◽  
pp. 721-729 ◽  
Author(s):  
B. Marczynski ◽  
M. Raulf-Heimsoth ◽  
B. Pesch ◽  
B. Kendzia ◽  
HU Käfferlein ◽  
...  
Keyword(s):  
Dna Damage ◽  
Comet Assay ◽  
Rank Correlation ◽  
Dna Strand Breaks ◽  
Strand Breaks ◽  
Blood Lymphocytes ◽  
Exposed Workers ◽  
Before And After ◽  
Dna Strand ◽  
Post Shift

DNA strand breaks were determined in leucocytes of induced sputum (IS) and compared with DNA strand breaks in blood lymphocytes from 42 bitumen-exposed workers pre and post shift. Comet assay results were expressed in arbitrary units based on visual scoring (sputum leucocytes) and Olive tail moment (OTM, blood lymphocytes). DNA damage in IS leucocytes was overall high but did not change during shift. Level of DNA strand breaks in IS samples correlated with total cell count and neutrophil content (Spearman rank correlation coefficient rs = 0.47, p = 0.001, rs= 0.48, p = 0.001, respectively) and with IL-8 concentration before and after shift (rs = 0.31, P = 0.048, and rs = 0.43, P = 0.005). DNA damage in IS was not associated with DNA strand breaks in blood lymphocytes (rs = —0.04, p = 0.802 before shift, rs = 0.27, p = 0.088 after shift). A higher level of DNA strand breaks was measured in blood lymphocytes before shift (median OTM 1.7 before and 1.3 after shift, p = 0.023). A strong correlation was found between the number of neutrophils and IL-8 concentration in IS before and after shift (rs = 0.77 and rs= 0.75, p < 0.001). This study showed an association between genotoxic and inflammatory effects in the lower airways and compared simultaneously DNA strand breaks in IS and blood of bitumen-exposed workers.

Use of comet assay for the risk assessment of oil- and chemical-industry workers

2014 ◽  
Vol 155 (47) ◽  
pp. 1872-1875 ◽  
Author(s):  
János Megyesi ◽  
Anna Biró ◽  
László Wigmond ◽  
Jenő Major ◽  
Anna Tompa
Keyword(s):  
Dna Damage ◽  
Occupational Exposure ◽  
Comet Assay ◽  
Oxidative Dna Damage ◽  
Microscopic Method ◽  
Monitoring Method ◽  
Strand Breaks ◽  
Cell Counts ◽  
Polycyclic Aromatic ◽  
Dna Strand

Introduction: The comet assay is a fluorescent microscopic method that is able to detect DNA strand-breaks even in non-proliferative cells in samples with low cell counts. Aim: The aim of the authors was to measure genotoxic DNA damage and assess oxidative DNA damage caused by occupational exposure in groups exposed to benzene, polycyclic aromatic carbohydrates and styrene at the workplace in order to clarify whether the comet assay can be used as an effect marker tool in genotoxicology monitoring. Method: In addition to the basic steps of the comet assay, one sample was treated with formamido-pirimidine-DNA-glycolase restriction-enzyme that measures oxidative DNA damage. Results: An increase was observed in tail moments in each group of untreated and Fpg-treated samples compared to the control. Conclusions: It can be concluded that occupational exposure can be detected with the method. The comet assay may prove to be an excellent effect marker and a supplementary technique for monitoring the presence or absence of genotoxic effects. Orv. Hetil., 2014, 155(47), 1872–1875.

scholarly journals DNA damage in human whole blood caused by radiopharmaceuticals evaluated by the comet assay

Mutagenesis ◽  
2019 ◽  
Vol 34 (3) ◽  
pp. 239-244 ◽  
Author(s):  
Heinz H Schmeiser ◽  
Karl-Rudolf Muehlbauer ◽  
Walter Mier ◽  
Ann-Christin Baranski ◽  
Oliver Neels ◽  
...  
Keyword(s):  
Dna Damage ◽  
Comet Assay ◽  
Dna Strand Breaks ◽  
Strand Breaks ◽  
Human Whole Blood ◽  
Time Points ◽  
Dna Migration ◽  
Activity Concentrations ◽  
Positron Emitters ◽  
Dna Strand

Abstract Radiopharmaceuticals used for diagnosis or therapy induce DNA strand breaks, which may be detectable by single-cell gel electrophoresis (called comet assay). Blood was taken from patients before and at different time points after treatment with radiopharmaceuticals; blood cells were investigated by the comet assay using the percentage of DNA in the tail as the critical parameter. Whereas [225Ac]Ac-prostate-specific membrane antigen (PSMA)-617 alpha therapy showed no difference relative to the blood sample taken before treatment, beta therapy with [177Lu]Lu-PSMA-617 3 h post-injection revealed a small but significant increase in DNA strand breaks. In blood of patients who underwent positron emission tomography (PET) with either [18F]2-fluor-2-deoxy-D-glucose (FDG) or [68Ga]Ga-PSMA-11, an increase of DNA migration determined by the comet assay was not found when analysed at different time points (2–70 min) after intravenous tracer injection. Human whole blood was incubated with the targeted clinically relevant therapeutic radiopharmaceuticals [225Ac]Ac-PSMA-617, [177Lu]Lu-PSMA-617 and [90Y]Y-DOTA(0)-Phe(1)-Tyr(3)-octreotide (DOTA-TOC) at different activity concentrations (kBq/ml) for 5 days and then analysed by the comet assay. DNA damage increased with higher concentrations of all radiolabeled compounds tested. [177Lu]Lu-PSMA-617 caused higher blood cell radiotoxicity than equal activity concentrations of [90Y]Y-DOTA-TOC. Likewise, whole human blood was exposed to the positron emitters [18F]FDG and [68Ga]Ga-PSMA-11 in vitro for 24 h with activity concentrations ranging between 5 and 40 MBq/ml. The same activity concentration dependent elevated DNA migration was observed for both compounds although decay energies are different. This study demonstrated that the amount of DNA damage detected by the comet assay in whole human blood is similar among different positron emitters and divergent by a factor of 200 between alpha particles and beta radiation.

scholarly journals Assessment of genotoxicity and depuration of anthracene in the juvenile coastal fish Trachinotus carolinus using the comet assay

2013 ◽  
Vol 61 (4) ◽  
pp. 215-222 ◽  
Author(s):  
Fabio Matsu Hasue ◽  
Maria José de Arruda Campos Rocha Passos ◽  
Thaís da Cruz Alves dos Santos ◽  
Arthur José da Silva Rocha ◽  
Caroline Patrício Vignardi ◽  
...  
Keyword(s):  
Dna Damage ◽  
Comet Assay ◽  
Aquatic Organisms ◽  
Dna Strand Breaks ◽  
Time Dependent ◽  
Clean Water ◽  
Oxygen Species ◽  
Coastal Fish ◽  
Strand Breaks ◽  
Dna Strand

In the environment, anthracene is characterized as being persistent, bioaccumulative and toxic to aquatic organisms. Biotransformation of xenobiotic substances, such as anthracene, produces reactive oxygen species that may induce DNA strand breaks. The aim of the present study was to evaluate the DNA damage in juvenile T. carolinus exposed to different concentrations (8, 16 and 32 µg.L-1) of anthracene for 24 h in the dark then subsequently allowed to depurate in clean water for different periods of time (48, 96 or 144 h) using the comet assay. Our results show that anthracene is genotoxic to T. carolinus and that DNA damage was dose- and depuration/time- dependent. Anthracenegenotoxicity was observed in all experimental concentrations. Depuration seemed to be more efficient in fish exposed to thelowest anthracene concentration and maintained in clean water for 96 h.

Acute hypoxia and hypoxic exercise induce DNA strand breaks and oxidative DNA damage in humans

2001 ◽  
Vol 15 (7) ◽  
pp. 1181-1186 ◽  
Author(s):  
PETER MØLLER ◽  
STEFFEN LOFT ◽  
CARSTEN LUNDBY ◽  
NIELS VIDIENDAL OLSEN
Keyword(s):  
Dna Damage ◽  
Oxidative Dna Damage ◽  
Acute Hypoxia ◽  
Dna Strand Breaks ◽  
Strand Breaks ◽  
Dna Strand ◽  
Hypoxic Exercise

scholarly journals Diallyl Sulfide Attenuation of Carcinogenesis in Mammary Epithelial Cells through the Inhibition of ROS Formation, and DNA Strand Breaks

Biomolecules ◽  
2021 ◽  
Vol 11 (9) ◽  
pp. 1313
Author(s):  
Selina F. Darling-Reed ◽  
Yasmeen Nkrumah-Elie ◽  
Dominique T. Ferguson ◽  
Hernan Flores-Rozas ◽  
Patricia Mendonca ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Proliferation ◽  
Dna Damage ◽  
Oxidative Dna Damage ◽  
Dna Strand Breaks ◽  
Diallyl Sulfide ◽  
Extracellular Medium ◽  
Strand Breaks ◽  
Ros Formation ◽  
Dna Strand

Garlic has long been used medicinally for many diseases, including cancer. One of the active garlic components is diallyl sulfide (DAS), which prevents carcinogenesis and reduces the incidence rate of several cancers. In this study, non-cancerous MCF-10A cells were used as a model to investigate the effect of DAS on Benzo (a)pyrene (BaP)-induced cellular carcinogenesis. The cells were evaluated based on changes in proliferation, cell cycle arrest, the formation of peroxides, 8-hydroxy-2-deoxyguanosine (8-OHdG) levels, the generation of DNA strand breaks, and DNA Polymerase β (Pol β) expression. The results obtained indicate that when co-treated with BaP, DAS inhibited BaP-induced cell proliferation (p < 0.05) to levels similar to the negative control. BaP treatment results in a two-fold increase in the accumulation of cells in the G2/M-phase of the cell cycle, which is restored to baseline levels, similar to untreated cells and vehicle-treated cells, when pretreated with 6 μM and 60 μM DAS, respectively. Co-treatment with DAS (60 μM and 600 μM) inhibited BaP-induced reactive oxygen species (ROS) formation by 132% and 133%, respectively, as determined by the accumulation of H2O2 in the extracellular medium and an increase in 8-OHdG levels of treated cells. All DAS concentrations inhibited BaP-induced DNA strand breaks through co-treatment and pre-treatment methods at all time points evaluated. Co-Treatment with 60 μM DAS increased DNA Pol β expression in response to BaP-induced lipid peroxidation and oxidative DNA damage. These results indicate that DAS effectively inhibited BaP-induced cell proliferation, cell cycle transitions, ROS, and DNA damage in an MCF-10A cell line. These results provide more experimental evidence for garlic’s antitumor abilities and corroborate many epidemiological studies regarding the association between the increased intake of garlic and the reduced risk of several types of cancer.

Detection of topoisomerase inhibitor-induced DNA strand breaks and apoptosis by the alkaline comet assay

2002 ◽  
Vol 520 (1-2) ◽  
pp. 47-56 ◽  
Author(s):  
Thierry Godard ◽  
Edwige Deslandes ◽  
François Sichel ◽  
Jean-Michel Poul ◽  
Pascal Gauduchon
Keyword(s):  
Comet Assay ◽  
Dna Strand Breaks ◽  
Alkaline Comet Assay ◽  
Strand Breaks ◽  
Topoisomerase Inhibitor ◽  
Dna Strand

Ascorbate potentiates DNA damage by 1-methyl-1-nitrosourea in vivo and generates DNA strand breaks in vitro

1987 ◽  
Vol 8 (11) ◽  
pp. 1657-1662 ◽  
Author(s):  
Paul V. Woolley ◽  
Shailendra Kumar ◽  
Peter Fitzgerald ◽  
Robert T. Simpson
Keyword(s):  
Dna Damage ◽  
Dna Strand Breaks ◽  
Strand Breaks ◽  
Dna Strand
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