Application of Single-Cell Gel Electrophoresis (Comet) Assay for Assessing Levels of DNA Damage in Canine and Feline Leukocytes

Heaton, Paul R.; Ransley, Raymond; Charlton, C. J.; Mann, Sarah; Stevenson, Joy; Smith, Brigitte H. E.; Rawlings, John M.; Harper, E. Jean

doi:10.1093/jn/132.6.1598s

Cited by 60 publications

(38 citation statements)

References 31 publications

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“…It has been reported that freezing of lymphocytes does not increase DNA strand breaks as compared to those of freshly isolated lymphocytes, and that fresh and frozen lymphocytes responded almost identically to hydrogen peroxide (Duthie et al, 2002). Visual scoring system used in this study reported to be accurately corresponding to the data that are measured by computer image analysis in human lymphocytes (Collins, 2004), in equine lymphocyte (Marlin et al, 2004) and in canine and feline leukocytes (Heaton et al, 2002). The method used for visual scoring involves classifying comets into four categories on the basis of the perceived length of migration and relative proportion of DNA in the tail.…”

Section: Discussionmentioning

confidence: 56%

Oxidative stress status of highly prolific sows during gestation and lactation

Berchieri-Ronchi

Kim

Yan

et al. 2011

Animal

167

122

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Elevated oxidative stress is reported to be associated with pregnancy complications in highly prolific sows. Oxidative DNA damage and the antioxidant status were determined in blood samples collected during the course of gestation and lactation in multiparous sows. Blood samples were drawn from sows (n 5 5) on days 30, 60, 90 and 110 of gestation (G30, G60, G90 and G110, respectively), on day 3, 10 and 18 of lactation (L3, L10 and L18, respectively) and on day 5 of postweaning (W5). Lymphocytes were isolated from the fresh blood and cryopreserved in each time point. Lymphocyte DNA damage was analyzed by alkaline single-cell gel electrophoresis (comet assay) to determine the single-and double-strand brakes and endogenous antioxidant concentrations using an HPLC system with UV detection. The comet assay showed elevated (P , 0.05) DNA damage (between 38% and 47%) throughout the gestational and lactational periods than during early gestation (G30; 21%). Plasma retinol concentration was reduced (P , 0.05) at the end of gestation (G110) compared with G30. Plasma a-tocopherol concentrations also showed a similar trend as to retinol. This study indicates that there is an increased systemic oxidative stress during late gestation and lactation, which are not fully recovered until the weaning compared with the G30, and that antioxidant nutrients in circulation substantially reduced in the mother pig at G110.

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Section: Discussionmentioning

confidence: 56%

Oxidative stress status of highly prolific sows during gestation and lactation

Berchieri-Ronchi

Kim

Yan

et al. 2011

Animal

167

122

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“…The comet assay also as known single cell gel electrophoresis (SCGE) was performed and scored with slight modifications according to , Kizilian et al (1999), Tice et al (2000), Saleha Banu et al (2000), Heaton et al (2002), Das et al (2006) and Turkez (2011). The OxiSelect 96-Well Comet Assay kits (Cell Biolabs , San Diego, CA, USA) were used to perform this assay.…”

Section: Scge Assaymentioning

confidence: 99%

Antioxidative, anticancer and genotoxic properties of α-pinene on N2a neuroblastoma cells

2013

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α-Pinene, an organic monoterpene, is found in essential oils of pine and coniferous trees. To date, although various biological activities of α-pinene have been demonstrated, its neurotoxicity has never been explored. Therefore in this study, we aimed to describe in vitro antiproliferative and/or cytotoxic properties by 3-(4,5-dimetylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test, genotoxic damage potentials by single cell gel electrophoresis, and oxidative effects by total antioxidant capacity (TAC) and total oxidative stress (TOS) analysis of α-pinene. Statistical analysis of MTT assay results indicated significant (p < 0.05) decreases of the cell proliferation rates in healthy neurons treated with α-pinene at only 400 mg/L, while significant decreases were observed in N2a cells at 100, 200 and 400 mg/L. On the other hand, the mean values of the total scores of cells showing DNA damage were not found significantly different from the control values on both cells. In addition, our results indicated that 10 and 25 mg/L of α-pinene treatment caused increases of TAC levels in primary rat neurons without any alterations of its level in N2a cells. However, α-pinene treatments at higher doses led to increases of TOS levels in both cell types. Overall our results suggest that α-pinene is of a limited therapeutic use as an anticancer agent.

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“…Then the slides were washed for 5 min with 1 % acetic acid, twice with distilled water, and air-dried. DNA impairment was evaluated by a visual score on a qualitative scale, ranging from 0 (no DNA damage) to 4 (extensive DNA damage, based on the length of the comet tail and on the relative proportion of DNA in the comet head and tail [10]. DNA damage was calculated in units introduced by Collins et al [5].…”

Section: Dna Comet Assaymentioning

confidence: 99%

Correlation of the cytotoxic activity of four different alkaloids, from Chelidonium majus (greater celandine), with their DNA intercalating properties and ability to induce breaks in the DNA of NK/Ly murine lymphoma cells

Kaminskyy¹,

Lootsik

Stoika³

2006

Open Life Sciences

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Abstract:The purpose of this study was to examine the relationship between the DNA intercalating characteristics and the DNA damaging capacity of four alkaloids extracted from Chelidonium majus L, as well as their toxicity towards murine NK/Ly lymphoma cells. Chelerythrine, sanguinarine and coptisine were found to be intercalated into the DNA isolated from NK/Ly cells, meanwhile, chelidonine exhibited no affinity to DNA. Sanguinarine exhibited the greatest toxicity toward NK/Ly cells, and the toxicity of the other three decreased in descending order: chelerythrine, coptisine and chelidonine. Chelerythrine and sanguinarine caused DNA damage, illustrated by the formation of comets of the third class. Coptisine was less toxic than chelerythrine and sanguinarine, and affected the formation the same class of comets in higher concentration. The quantity of comets induced by chelidonine were negligible, a finding consistent with its inability to intercalate into DNA structure. The ability of four main alkaloids of Chelidonium majus L., to intercalate into DNA isolated from murine NK/Ly lymphoma cells, correlated with their ability to induce breaks in cellular DNA and with their toxic effect towards those cells.

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Application of Single-Cell Gel Electrophoresis (Comet) Assay for Assessing Levels of DNA Damage in Canine and Feline Leukocytes

Cited by 60 publications

References 31 publications

Oxidative stress status of highly prolific sows during gestation and lactation

Oxidative stress status of highly prolific sows during gestation and lactation

Antioxidative, anticancer and genotoxic properties of α-pinene on N2a neuroblastoma cells

Correlation of the cytotoxic activity of four different alkaloids, from Chelidonium majus (greater celandine), with their DNA intercalating properties and ability to induce breaks in the DNA of NK/Ly murine lymphoma cells

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