Comet assay on mouse oocytes: an improved technique to evaluate genotoxic risk on female germ cells  

Oleh:

Berthelot-Ricou, Anais ; Perrin, Jeanne ; Giorgio, Carole Di ; Meo, Michel De ; Botta, Alain ; Courbiere, Blandine

Jenis: Article from Journal - ilmiah internasional
Dalam koleksi: Fertility and Sterility (keterangan: ada di ClinicalKey) vol. 95 no. 04 (Mar. 2011), page 1452-1457.
Topik: GENOTOXIC; Mice; oocytes; comet assay; germ cells; genotoxicity; DNA damages

Ketersediaan

Perpustakaan FK Nomor Panggil: F02.K.2011.03 Non-tandon: 1 (dapat dipinjam: 0) Tandon: tidak ada

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Isi artikel

Objective To develop and validate an efficient comet assay on mouse oocytes without depellucidation. Design In vitro experiments using a murine model. Setting Biogenotoxicology research laboratory in Aix-Marseille II University, France. Animal(s) CD1 prepubescent female mice. Intervention(s) DNA lesions in oocytes were evaluated by the alkaline comet assay. After oocyte retrieval, we first studied the effect of zona pellucida (ZP) on comet morphology. For this study, we applied the comet assay to mature oocytes with and without ZP after exposure to simulated sunlight irradiation (SSI) compared with negative controls. Next, nondepellucidated mouse oocytes were exposed to three well-known genotoxic agents (SSI, methylmethanesulfonate [MMS], and hydrogen peroxide [H2O2]) and compared with negative controls. Images of oocytes were analyzed with Komet software. Main Outcome Measure(s) DNA damages were quantified and expressed as olive tail moment (OTM), defined as the product of the tail length and the fraction of total DNA in the tail. OTM?2 were calculated from OTM; they corresponded to the degrees of freedom (n) of each OTM distribution obtained from at least 50 oocytes. OTM?2 is an indicator of DNA lesions. The test was considered positive and statistically significant when OTM?2 increased in oocytes compared with the medium-only control cells. Result(s) There was no difference in comet aspect between oocyte groups with and without ZP. The three genotoxic agents significantly increased DNA damages as compared with the control groups. The OTM?2 values were (mean ± SD): 2.1 ± 0.07, 7.73 ± 0.35, 3.35 ± 0.15, and 12.4 ± 0.51 in control, SSI, MMS, and H2O2 groups, respectively. Conclusion(s) Comet assay on non depellucidated mouse oocytes is a rapid and easy test. This assay would be useful to assess the genotoxicity on female germ cells of chemicals, drugs, or environmental pollutants and the efficiency of antioxidant molecules.

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