Aneugenic potential of okadaic acid revealed by the micronucleus assay combined with the FISH technique in CHO-K1 cells.
Okadaic acid (OA) is a major toxin involved in diarrhetic shellfish poisoning in humans and has been shown to be both a potent tumor promoter in rodent skin and stomach and an inhibitor of serine/threonine protein phosphatases, specifically PP1 and PP2A. The research on the genotoxic potential of OA amounts to only a few studies, which give conflicting results. In order to evaluate the ability of OA to induce DNA damage, the cytokinesis-block micronucleus assay was performed in the CHO-K1 cell line. A statistically significant induction of micronuclei without strong cytotoxicity was obtained after a 24 h treatment with 20 (approximately 5-fold) and 30 nM (approximately 10-fold) OA. Then, in order to discriminate between a clastogenic or aneugenic effect of OA, the micronucleus assay was carried out in combination with fluorescence in situ hybridization (FISH) using a (TTAGGG)(n) DNA probe for centromere detection. FISH analysis showed that OA mainly induced centromere-positive micronuclei (68.9% induction with 20 nM OA and 77.0% with 30 nM). Therefore, OA can be considered aneugenic. Using the same assay, biotransformation of OA was studied after a 4 h treatment with and without metabolic activation. The results show that reactive metabolites of OA were generated with a significant increase in genotoxic potential. The relationship between the different components involved in the mitotic process and OA inhibition of protein phosphatase is also discussed.
Le Hegarat
Ludovic
L
AFSSA, Laboratoire d'Etudes et de Recherches sur les Médicaments Vétérinaires et les Désinfectants, Unité de Toxicologie Alimentaire, BP 90203, F-35302 Fougères, France.
Puech
Lilian
L
Fessard
Valérie
V
Poul
Jean Michel
JM
Dragacci
Sylviane
S
D024422:Aneugens / Q000494:pharmacology*; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D005821:Genetic Techniques; D017404:In Situ Hybridization, Fluorescence; D048629:Micronuclei, Chromosome-Defective / Q000378:metabolism*; D015162:Micronucleus Tests; D019319:Okadaic Acid / Q000494:pharmacology*
D024422:Aneugens; D019319:Okadaic Acid