Simple detection of chemical mutagens by the alkaline single-cell gel electrophoresis (Comet) assay in multiple mouse organs (liver, lung, spleen, kidney, and bone marrow)

Sasaki, Yu; Nishidate, Emi; Izumiyama, Fusako; Matsusaka, Naonori; Tsuda, Shuji

doi:10.1016/s1383-5718(97)00073-9

Cited by 109 publications

(36 citation statements)

References 16 publications

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“…Comet assay is a very sensitive technique for detecting individual eukaryote cells of multiple classes of DNA damage and incomplete excision repair sites associated with the elimination of DNA damage (Fairbairn et al, 1995). Sasaki et al (1997) used the comet assay to detect DNA damage by various mutagens and found that hepatic DNA strand breaks were increased by oral administration of B [a]P. In DNA strand breaks caused by dimethylarsinic acid, MT-I/II null mice were more susceptible than wild-type mice (Jia et al, 2004).…”

Section: Discussionmentioning

confidence: 99%

“…The liver was gently homogenized according to the method described by Sasaki et al (1997) with some moddone according to the method described by Singh et al quantify the average DNA strand breaks, all of them were (normal; score = 0); tail length < head length (score = 1); tail length < 2 × head length (score = 2); tail length < 3 × head length (score = 3). Comet scores were the calculated sum of 50 nuclei.…”

Section: Single Cell Gel Electrophoresis (Comet) Assaymentioning

confidence: 99%

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Protective role of metallothionein in benzo[a]pyrene-induced DNA damage

et al. 2009

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INTRODUCTION Benzo[a]pyrene (B[a]P) is one of the polycyclic aromatic hydrocarbons (PAH) referred to as a human carcinogen (IARC monographs, in preparation). B[a]P has beenshown to induce gene mutations, chromosomal aberrations and other types of genotoxic effects in in vitro and in vivo. Most humans are more or less constantly exposed to B[a]P. As a product of incomplete combustion, B[a]P is present in tobacco smoke, diesel engine exhaust, urban air, various types of processed foods, coal-tar, creosote, asphalt, and various occupational settings such as coal B[a]P is metabolized to 7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE) by cytochrome P450 (P450 or CYP). BPDE is the ultimate carcinogenic form of B[a]P and binds to the exocyclic nitrogen of deoxyguanosine in DNA. BPDE-DNA adducts in target tissues were closely related to development of cancer in animals and humans (Culp and Beland, 1994). In addition, oxidative DNA damage also plays an important role in the process of B[a]P carcinogenesis, because B[a]P produces quinone derivatives which easily generate reactive oxygen species (ROS) and oxidatively damage DNA (Penning et al., 1996). As a result of hydroxyl radical interaction with DNA, many types of oxidized a]P administration, although that of wild-type mice was only slightly changed. Because cytochrome P450 (CYP) plays a major role in the process of B[a]P metabolic activation, we attempted to reveal the effect of MT on metabolic activation of B[a]P. Although CYP1A activities were elevated in the livers of MT-I/II null mice and wild-type mice treated with B[a]P, it was not different between both strains of mice. In addition, a]P treatment, whereas MT was not detected in livers of MT-I/II null mice with or without B[a]P treatment. These results demonstrate that MT acts as an endogenous defensive factor against B[a]P-induced DNA damage.

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Section: Discussionmentioning

confidence: 99%

Section: Single Cell Gel Electrophoresis (Comet) Assaymentioning

confidence: 99%

Protective role of metallothionein in benzo[a]pyrene-induced DNA damage

et al. 2009

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“…To obtain nuclei, the homogenate was centrifuged at 700 g for 10 min at 08C, and the precipitate was resuspended in chilled homogenizing buffer at 0.5 g=mL and allowed to settle; precipitated clumps were then removed. Doses and times were selected based on the preliminary studies as well as literature-reported values (Sasaki et al, 1997a(Sasaki et al, , 1997bTsuda et al, 2000;Sekihashi et al, 2002).…”

Section: Treatment and Organ Preparationmentioning

confidence: 99%

“…The in vivo alkaline SCGE assay was conducted based on the method described by Sasaki et al (1997a) with some modifications. One hundred microliters of NMP agarose was quickly layered on conventional slides, the slides were covered with a cover slip, and then the slides were placed on ice to allow agarose to gel.…”

Section: Slide Preparation and Alkaline Scge Assaymentioning

confidence: 99%

Protective Effect of Crocus sativus Stigma Extract and Crocin (trans-crocin 4) on Methyl Methanesulfonate–Induced DNA Damage in Mice Organs

Hosseinzadeh

Abootorabi

Sadeghnia

2008

DNA and Cell Biology

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This study was designed to examine the effect of aqueous extract of Crocus sativus stigmas (CSE) and crocin (trans-crocin 4) on methyl methanesulfonate (MMS)-induced DNA damage in multiple mice organs using comet assay. Adult male NMRI mice in different groups were treated with either physiological saline (10 mL=Kg, intraperitoneal [ip]), CSE (80 mg=Kg, ip), crocin (400 mg=Kg, ip), MMS (120 mg=Kg, ip), and CSE (5, 20, and 80 mg=Kg, ip) 45 min prior to MMS administration or crocin (50, 200, and 400 mg=Kg, ip) 45 min prior to MMS administration. Mice were scarified about 3 h after each different treatment, and the alkaline comet assay was used to evaluate the effect of these compounds on DNA damage in different mice organs. The percent of DNA in the comet tail (% tail DNA) was measured. A significant increase in the % tail DNA was seen in nuclei of different organs of MMS-treated mice. In control groups, no significant difference was found in the % tail DNA between CSE-or crocin-pretreated and saline-pretreated mice. The MMS-induced DNA damage in CSEpretreated mice (80 mg=Kg) was decreased between 2.67-fold (kidney) and 4.48-fold (lung) compared to those of MMS-treated animals alone ( p < 0.001). This suppression of DNA damage by CSE was found to be depended on the dose, which pretreatment with CSE (5 mg=Kg) only reduced DNA damage by 6.97%, 6.57%, 7.27%, and 9.90% in liver, lung, kidney, and spleen, respectively ( p > 0.05 as compared with MMS-treated group). In the same way, crocin also significantly decreased DNA damage by MMS (between 4.69-fold for liver and 6.55-fold for spleen, 400 mg=Kg), in a dose-dependent manner. These data indicate that there is a genoprotective property in CSE and crocin, as revealed by the comet assay, in vivo.

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“…The former is very important so as to avoid pseudonegative results and the comet assay can detecta wide variety of genotoxic compounds both in vitro [35,36] and in vivo [3,[37][38][39][40]. The micronucleus test (MN test) is a standard procedure that can detect structural chromosome aberrations derived from initial damage in the S phase and/or numerical chromosome aberrations due to an eugenic effects in the M phase [40,41].…”

Section: Power Of the Comet Assay To Detect Low Level Genotoxicitymentioning

confidence: 99%

The Power of the Comet Assay to Detect Low Level Genotoxicity and DNA Repair Factors Affecting its Power

Sasaki

SatomiKawaguchi

Nakamura

2017

MOJT

Self Cite

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The comet assay is considered to be a rapid and sensitive procedure for quantitating DNA damage in mammalian cells. In this article, to interpret its outcomes adequately, its power to detect low level genotoxicity and factors affecting its power were reviewed. Although the development of initial lesions into alkali-labile sites and/or SSBs through repairing events is an important factor to support its detecting power, their repair reduce its power to detect SSBs as an initial DNA lesion. The acellular comet assay, in which slides with gel are prepared from untreated cells are exposed after lysis to test agents. Thus, detection of SSBs as initial lesions but not alkali-labile sites generated from DNA lesion such as alkylated bases and the power to detect low level SSBs as initial lesions is lower in the standard than in the acellular assay. The acellular comet assay would be practically used to detect SSBs as initial lesions. The inhibitors of re-synthesis andincisionsteps of the excision repair enhance and suppress comet-tail formation, respectively. Although the incision step of repairing process is necessary to support the sensitivity of the comet assay, its re-synthesis step reduces the power to detect low level genotoxicity. Although this assay can detect a wide variety of genotoxoic compounds and its power to detect genotoxicity were identical to that of the MN test, its power to detect low levels of genotoxic potential is inferior to that of the MN test. However, its power of to detect a low level of genotoxicity can be elevated to a level higher than that of the MN test by using DNA resynthesis inhibitors, such as araC and HU. In conclusion, the outcomes of comet assay should be interpretedappropriately in consideration of the action of DNA repair.

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Simple detection of chemical mutagens by the alkaline single-cell gel electrophoresis (Comet) assay in multiple mouse organs (liver, lung, spleen, kidney, and bone marrow)

Cited by 109 publications

References 16 publications

Protective role of metallothionein in benzo[a]pyrene-induced DNA damage

Protective role of metallothionein in benzo[a]pyrene-induced DNA damage

Protective Effect of Crocus sativus Stigma Extract and Crocin (trans-crocin 4) on Methyl Methanesulfonate–Induced DNA Damage in Mice Organs

The Power of the Comet Assay to Detect Low Level Genotoxicity and DNA Repair Factors Affecting its Power

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