Okadaic acid: chromosomal non-disjunction analysis in human lymphocytes and study of aneugenic pathway in CHO-K1 cells.
Okadaic acid (OA) is the main marine toxin implicated in the diarrhetic shellfish poisoning (DSP) in humans after consumption of contaminated bivalve molluscs. We have previously shown that OA was an in vitro aneugenic compound that induced chromosome loss via micronuclei formation in CHO-K1 cells. The aims of this study were to investigate the chromosomal non-disjunction (ND) potential of OA in human lymphocytes and the pathways involved for aneuploidy in CHO-K1 cells. Firstly, we analysed the formation of micronuclei and the non-disjunction for chromosomes 1 and 17 in binucleated human lymphocytes cells with the cytokinesis-blocked micronucleus (CBMN) assay coupled to a fluorescent in situ hybridization (FISH) technique with centromere-specific DNA probes. We showed that OA statistically increased the frequency of micronucleated lymphocytes in the dose range from 20 to 35 nM. However, FISH analysis did not reveal any increase in the non-disjunction for both chromosomes whatever the concentration between 2.5 and 35 nM. However, a significant increase in ND for the chromosome 17 was found at 1 nM. Secondly, in CHO-K1 cells, we investigated the dose and time dependent effects of OA: (i) on cell cycle progression, (ii) on mitotic-phase arrest and (ii) on mitotic spindle and centrosome abnormalities. The results showed that OA induced a progressive accumulation of mitotic CHO-K1 cells in prometaphase, an induction of multipolar mitotic spindle with centrosome amplification and the formation of multinucleated cells. We concluded that OA did not induce chromosome non-disjunction but should more likely induced chromosome loss in human lymphocytes. Moreover, our results obtained in CHO-K1 suggest that OA induced aneuploidy by preventing the chromosome attachment to the mitotic spindle and by amplifying the centrosome. The mode of action of the toxin in relation to its inhibition of protein phosphatases 1 (PP1) and 2A (PP2A) and the mitosis process is discussed.
Hégarat
Ludovic Le
LL
AFSSA, Laboratoire d'Etudes et de Recherches sur les Médicaments Vétérinaires et les Désinfectants, Unité de Toxicologie Génétique des Contaminants Alimentaires, B.P. 90203, 35302 Fougères, France.
Orsière
Thierry
T
Botta
Alain
A
Fessard
Valérie
V
D024422:Aneugens / Q000633:toxicity*; D000782:Aneuploidy; D000818:Animals; D016466:CHO Cells; D002453:Cell Cycle / Q000187:drug effects; D002478:Cells, Cultured; D018385:Centrosome / Q000187:drug effects; D002877:Chromosomes, Human / Q000187:drug effects*; D002878:Chromosomes, Human, Pair 1 / Q000187:drug effects; D002886:Chromosomes, Human, Pair 17 / Q000187:drug effects; D003078:Colchicine / Q000633:toxicity; D006224:Cricetinae; D003412:Cricetulus; D004305:Dose-Response Relationship, Drug; D006801:Humans; D007700:Kinetics; D008214:Lymphocytes / Q000187:drug effects*; D048629:Micronuclei, Chromosome-Defective / Q000187:drug effects; D015162:Micronucleus Tests; D008954:Models, Biological; D009630:Nondisjunction, Genetic; D019319:Okadaic Acid / Q000633:toxicity*; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors; D049468:Prometaphase / Q000187:drug effects; D008941:Spindle Apparatus / Q000187:drug effects
D024422:Aneugens; D019319:Okadaic Acid; D010749:Phosphoprotein Phosphatases; D003078:Colchicine