title: Marine toxin okadaic acid induces aneuploidy in CHO-K1 cells in presence of rat liver postmitochondrial fraction, revealed by cytokinesis-block micronucleus assay coupled to FISH.
abstract: Okadaic acid (OA), a major polyether toxin involved in diarrhetic shellfish poisoning (DSP), is a potent tumor promoter in rodent skin and glandular stomach and a specific inhibitor of the serine/threonine protein phosphatases PP1 and PP2A. A previous study, which used the cytokinesis-block micronucleus (CBMN) assay in CHO-K1 cells, showed that OA induced chromosome damage in the presence of a rat liver metabolic activation system (S9). To support OA biotransformation by S9, the same test system was performed, and DNA damage induced by OA was measured with and without metabolic activation as well as in the presence of heat-inactivated S9 fraction. The results showed that only in the presence of active S9 did OA significantly increased the frequency of micronucleated binucleated (MNBN) cells. After a 4-h treatment a 2- to 5-fold increase of MNBN cells was observed at 30 nM and at 50 nM of OA. However, without S9 or in the presence of heat-inactivated S9, OA did not induce any chromosome damage. We concluded that OA can be metabolically activated in vitro into metabolites that are more genotoxic. The CBMN assay coupled with fluorescence in situ hybridization (FISH) using a DNA probe for centromere detection was performed to discriminate between clastogenic (chromosome breakage) and aneugenic (chromosome loss) effects. FISH analysis showed that OA metabolites increased in a dose-dependent manner in centromere positive micronuclei (CEN+): 60% of CEN+ at 30 nM and 75% of CEN+ at 50 nM of OA. The uptake of OA into CHO-K1 cells and the biotransformation of the toxin are discussed.
authors.affiliation: AFSSA, Laboratoire d'Etudes et de Recherches sur les Médicaments Vétérinaires et les Désinfectants, Unité de Toxicologie Alimentaire, B.P. 90203, 35302 Fougères cedex, France.
authors.forename: Ludovic
authors.initials: L
authors.lastname: Le Hégarat
authors.forename: Valérie
authors.initials: V
authors.lastname: Fessard
authors.forename: Jean Michel
authors.initials: JM
authors.lastname: Poul
authors.forename: Sylviane
authors.initials: S
authors.lastname: Dragacci
authors.forename: Pascal
authors.initials: P
authors.lastname: Sanders
mesh_terms: D000704:Analysis of Variance; D000782:Aneuploidy; D000818:Animals; D016466:CHO Cells; D002455:Cell Division / Q000187:drug effects; D019457:Chromosome Breakage; D002872:Chromosome Deletion; D006224:Cricetinae; D003412:Cricetulus; D004249:DNA Damage / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D017404:In Situ Hybridization, Fluorescence; D008099:Liver / Q000737:chemistry; D048629:Micronuclei, Chromosome-Defective / Q000187:drug effects*; D015162:Micronucleus Tests; D019319:Okadaic Acid / Q000493:pharmacokinetics / Q000633:toxicity*; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D019319:Okadaic Acid