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A novel approach using DNA-repair-deficient chicken DT40 cell lines for screening and characterizing the genotoxicity of environmental contaminants.

Ji K, Kogame T, Choi K, Wang X, Lee J, Taniguchi Y, Takeda S - Environ. Health Perspect. (2009)

Bottom Line: We aimed to establish a sensitive and fast screening method that is also capable of characterizing mechanisms of genotoxicity.We also characterized DNA lesions induced by NaAsO(2).Genotoxicity of given stressors was successfully screened based on a comparison of proliferation kinetics between wild-type and DNA-repair-deficient mutants in 48 hr.

View Article: PubMed Central - PubMed

Affiliation: School of Public Health, Seoul National University, Seoul, Korea.

ABSTRACT

Background: Many bacterial or mammalian cell-based test systems, such as the Ames test, chromosomal aberration assays, or gene mutation assays, are commonly used in developed countries to detect the genotoxicity of industrial chemicals. However, the specificity is generally limited and the sensitivity is not sufficiently high. In addition, most assays cannot provide information on mechanisms of genotoxicity of a given chemical.

Objectives: We aimed to establish a sensitive and fast screening method that is also capable of characterizing mechanisms of genotoxicity.

Methods: We developed a novel bioassay employing gene-disrupted clones of the chicken DT40 B-lymphocyte line, which are designed to be deficient in several specific DNA repair pathways. Genotoxic chemicals can delay cellular proliferation in DNA-repair-deficient clones more significantly than in wild-type cells by interfering with DNA replication, thereby inducing DNA damage. In addition, we verified the validity of this assay by analyzing the genotoxicity of gamma-rays, ultraviolet (UV) light, and sodium metaarsenite (NaAsO(2)). We also characterized DNA lesions induced by NaAsO(2).

Results: Genotoxicity of given stressors was successfully screened based on a comparison of proliferation kinetics between wild-type and DNA-repair-deficient mutants in 48 hr. We also found that NaAsO(2) apparently induces at least two types of damage: chromosomal breaks and UV photoproduct-like DNA lesions.

Conclusion: This bioassay is a reliable and sensitive screening tool for environmental mutagens as well as for further characterizing the nature of detected genotoxicity.

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Related in: MedlinePlus

Protocol for treatment of cells deficient for DSB repair (A) and response of these cells to NaAsO2 with or without NAC (B).
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f1-ehp-117-1737: Protocol for treatment of cells deficient for DSB repair (A) and response of these cells to NaAsO2 with or without NAC (B).

Mentions: To evaluate the involvement of reactive oxygen species (ROS) in DNA damage induction by NaAsO2, we pretreated DT40 cells with 1 mM N-acetyl-L-cysteine (NAC; Sigma), an ROS scavenger, 2 hr before NaAsO2 treatment (Figure 1A). Minimum concentration and treatment times were determined by preliminary experiments using 15 μM NaAsO2.


A novel approach using DNA-repair-deficient chicken DT40 cell lines for screening and characterizing the genotoxicity of environmental contaminants.

Ji K, Kogame T, Choi K, Wang X, Lee J, Taniguchi Y, Takeda S - Environ. Health Perspect. (2009)

Protocol for treatment of cells deficient for DSB repair (A) and response of these cells to NaAsO2 with or without NAC (B).
© Copyright Policy - public-domain
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2801191&req=5

f1-ehp-117-1737: Protocol for treatment of cells deficient for DSB repair (A) and response of these cells to NaAsO2 with or without NAC (B).
Mentions: To evaluate the involvement of reactive oxygen species (ROS) in DNA damage induction by NaAsO2, we pretreated DT40 cells with 1 mM N-acetyl-L-cysteine (NAC; Sigma), an ROS scavenger, 2 hr before NaAsO2 treatment (Figure 1A). Minimum concentration and treatment times were determined by preliminary experiments using 15 μM NaAsO2.

Bottom Line: We aimed to establish a sensitive and fast screening method that is also capable of characterizing mechanisms of genotoxicity.We also characterized DNA lesions induced by NaAsO(2).Genotoxicity of given stressors was successfully screened based on a comparison of proliferation kinetics between wild-type and DNA-repair-deficient mutants in 48 hr.

View Article: PubMed Central - PubMed

Affiliation: School of Public Health, Seoul National University, Seoul, Korea.

ABSTRACT

Background: Many bacterial or mammalian cell-based test systems, such as the Ames test, chromosomal aberration assays, or gene mutation assays, are commonly used in developed countries to detect the genotoxicity of industrial chemicals. However, the specificity is generally limited and the sensitivity is not sufficiently high. In addition, most assays cannot provide information on mechanisms of genotoxicity of a given chemical.

Objectives: We aimed to establish a sensitive and fast screening method that is also capable of characterizing mechanisms of genotoxicity.

Methods: We developed a novel bioassay employing gene-disrupted clones of the chicken DT40 B-lymphocyte line, which are designed to be deficient in several specific DNA repair pathways. Genotoxic chemicals can delay cellular proliferation in DNA-repair-deficient clones more significantly than in wild-type cells by interfering with DNA replication, thereby inducing DNA damage. In addition, we verified the validity of this assay by analyzing the genotoxicity of gamma-rays, ultraviolet (UV) light, and sodium metaarsenite (NaAsO(2)). We also characterized DNA lesions induced by NaAsO(2).

Results: Genotoxicity of given stressors was successfully screened based on a comparison of proliferation kinetics between wild-type and DNA-repair-deficient mutants in 48 hr. We also found that NaAsO(2) apparently induces at least two types of damage: chromosomal breaks and UV photoproduct-like DNA lesions.

Conclusion: This bioassay is a reliable and sensitive screening tool for environmental mutagens as well as for further characterizing the nature of detected genotoxicity.

Show MeSH
Related in: MedlinePlus