Clare M Gillian, Lorenzon Giocondo, Akhurst Leslie C, Marzin Daniel, van Delft Joost, Montero Regina, Botta Alain, Bertens Arma, Cinelli Serena, Thybaud Véronique, Lorge Elisabeth
Department of Genetic Toxicology, Safety Assessment, Astra Charnwood, Loughborough, Leics, UK.
Mutat Res. 2006 Aug 4;607(1):37-60. doi: 10.1016/j.mrgentox.2006.04.001. Epub 2006 Jun 12.
This study on the in vitro micronucleus assay, comprising 11 laboratories using human lymphocytes, was coordinated by an organizing committee supported by the SFTG (the French branch of the European Environmental Mutagen Society). Nine coded substances were assessed for their ability to induce micronuclei in human lymphocytes in vitro, mitomycin C being used as a positive control. Cultures were exposed to the test substances for a short (early or late) time or for a long time, followed by a short or long recovery period, in the presence of cytochalasin B. Each chemical was evaluated, generally in two laboratories, using three treatment schedules at least twice. The data were assessed for acceptability, and then classified as negative, positive or equivocal. Two of seven genotoxic compounds, namely colchicine and bleomycin, clearly induced micronuclei. Reproducible results were difficult to obtain for some substances, which tended to be those acting at specific stages of the cell cycle. Cytosine arabinoside, diethylstilboestrol and 5-fluorouracil were classified as equivocal. Urethane and thiabendazole were classified as negative. The two presumed non-genotoxic compounds, mannitol and clofibrate, did not induce micronuclei. Repeat testing, exposing cells at both an early and late time after mitogenic stimulation, was needed to detect substances classified as equivocal. These results show the importance of achieving sufficient inhibition of nuclear division to avoid the possibility of missing an effect. The evaluation of micronuclei in mononucleated as well as binucleated cells was particularly useful to detect aneugens. There were no false positive results using lymphocytes, indicating a high specificity. It is concluded that the clastogenic or aneugenic potential in vitro of the substances tested was correctly identified in this study, but that refining the protocol to take into account factors such as the stages of the cell cycle exposed to the compound, or the duration of recovery would be likely to improve the sensitivity of detection using lymphocytes.
这项关于体外微核试验的研究由欧洲环境诱变剂协会法国分会（SFTG）支持的一个组委会协调，有11个实验室使用人淋巴细胞参与。对9种编码物质进行了评估，以确定它们在体外诱导人淋巴细胞微核的能力，丝裂霉素C用作阳性对照。在细胞松弛素B存在的情况下，将培养物短时间（早期或晚期）或长时间暴露于受试物质，随后进行短时间或长时间的恢复期。每种化学物质通常在两个实验室进行评估，使用三种处理方案，至少重复两次。对数据进行可接受性评估，然后分为阴性、阳性或可疑。七种遗传毒性化合物中的两种，即秋水仙碱和博来霉素，明显诱导了微核。对于一些物质，尤其是那些作用于细胞周期特定阶段的物质，难以获得可重复的结果。阿糖胞苷、己烯雌酚和5-氟尿嘧啶被归类为可疑。氨基甲酸乙酯和噻苯达唑被归类为阴性。两种假定的非遗传毒性化合物，甘露醇和氯贝丁酯，未诱导微核。对于被归类为可疑的物质，需要在有丝分裂刺激后的早期和晚期都对细胞进行重复检测。这些结果表明，实现对核分裂的充分抑制以避免漏检效应的可能性非常重要。对单核细胞和双核细胞中的微核进行评估对于检测非整倍体特别有用。使用淋巴细胞没有假阳性结果，表明特异性很高。结论是，本研究正确识别了受试物质的体外致断裂或致非整倍体潜力，但完善方案以考虑诸如暴露于化合物时的细胞周期阶段或恢复期持续时间等因素，可能会提高使用淋巴细胞检测的灵敏度。
