Aneugenic potential of okadaic acid revealed by the micronucleus assay combined with the FISH technique in CHO-K1 cells
Aneugenic potential of okadaic acid revealed by the micronucleus assay combined with the FISH technique in CHO-K1 cells
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DOI：
10.1093/mutage/18.3.293
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发表时间：
2003-05-01
期刊：
影响因子：
2.7
通讯作者：
Dragacci, S
中科院分区：
文献类型：
--
作者：
Le Hegarat, L;Puech, L;Dragacci, S
冈田酸（Okadaic acid，OA）是引起人类贝类中毒的主要毒素，在啮齿类动物的皮肤和胃中具有很强的肿瘤促进作用，并能抑制丝氨酸/苏氨酸蛋白磷酸酶，特别是PP 1和PP 2A。关于OA潜在遗传毒性的研究为数不多，但结果相互矛盾。为了评价OA诱导DNA损伤的能力，在CHO-K1细胞系中进行了微核阻断试验。用20（类似于5倍）和30 nM（类似于10倍）OA处理24 h后，获得了统计学显著的微核诱导，但没有强烈的细胞毒性。然后，为了区分OA的致染色体断裂或非整倍体效应，使用（TTAGGG）（n）DNA探针检测着丝粒，结合荧光原位杂交（FISH）进行微核试验。荧光原位杂交分析表明OA主要诱导着丝粒阳性微核（20 nM和30 nM OA诱导率分别为68.9%和77.0%）。因此，OA可以被认为是非整倍体的。使用相同的试验，在有和无代谢活化的情况下处理4 h后研究OA的生物转化。结果表明，OA的反应性代谢产物产生的遗传毒性潜力显着增加。有丝分裂过程中所涉及的不同成分和OA抑制蛋白磷酸酶之间的关系进行了讨论。
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 (similar to5-fold) and 30 nM (similar to10-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.