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Application of Cytogenetic Endpoints and Comet Assay on Human Lymphocites Treated with Vincristine in Vitro

Vincristine belongs to vinca alkaloids, antimitotic drugs, and has been used in various chemotherapeutic regimens. The genotoxic potential of vincristine is assessed on human peripheral blood lymphocytes following administration of the drug at doses within the therapeutic range (0.0875 g/ml) by use of analysis of structural chromosome aberrations, micronucleus assay, sister chromatid exchange (SCE) analysis and Comet assay. In vitro treatment of human lymphocytes with vincristine was performed on cells in G0 phase as well on lymphocyte cultures 24 hours after stimulation with mitogen phytohaemagglutinine. For the Comet assay 24h, 48h and 72h after treatment treated cells were embedded in agarose on slides, lysed with alkaline lysis solution and exposed to an electric field. DNA migrated within the agarose and formed comets whose length depend on the amount of DNA damage. For the analysis of structural chromosome aberrations cells were grown on F-10 medium for 48 hours, and for micronucleus assay and SCE analysis for 72 hours. The results on Comet assay showed an increase in tail length compared to control both in cells treated in G0 and in cells treated 24 h after mitogen stimulation. The amount of DNA damage was higher in cells treated with vincristine 24 h after mitogen stimulation. Administered concentration of drug caused total inhibition of lymphocyte growth in 72 h cultures for micronucleus assay and SCE analysis indicating strong microtubule distruptive effects of vincristine. Analysis of structural chromosome aberrations reveal chromatid breaks and acentric fragments as the main aberration types both in cells treated in G0 and in cells treated 24 h after mitogen. Number of these aberrations was higher in cells treated in G0 phase. Results obtained in this study by use of different cytogenetic endpoints confirmed that vincristine exhibit both aneugenic and clastogenic effects on human lymphocytes.

vincristine; cytogenetic endpoints; human lymphocytes

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