INDIVIDUAL AND COMBINED CYTOTOXIC EFFECTS OF THE MAJOR FOUR AFLATOXINS IN DIFFERENT IN VITRO STABILIZED SYSTEMS

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Abstract
The present study aims to investigate the cytotoxic effect of the major aflatoxins (B1, B2, G2 and G2) and also aflatoxin combination, using a simple, rapid and cheap cytotoxicity test like MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay in three in vitro models (human umbilical vein endothelial cells [HUVEC], human lung fibroblasts [HFL] and A2780 cell line) and to extrapolate the data to in vivo situation using a prediction model. A difference in cell sensitivity has been observed for B1 and B1 + B2, in the following order A2789 > HFL > HUVEC, while for B2, G1, G2, Mix (B1 + B2 + G1 + G2) the order was HFL > A2789 > HUVEC when comparing the IC50 (half maximal inhibitory concentration) values. We confirm that in vitro cytotoxicity test MTT assay is able to predict in vivo toxicity, at least for aflatoxins using the prediction model. The values of LD50 (lethal dose 50%) calculated from experiments are different for each cell line. This fact may indicate that some species are more resistant than other and target organs are not necessarily those predicted, because the A2780 ovarian cancer cells seem to be more sensitive to B1 than cells of endothelial or fibroblasts origin. PRACTICAL APPLICATIONS This study is in concordance with the international tendency that refined the current techniques to lessen pain or distress, to reduce the number of animals necessary for a particular test or to replace animals with non-whole-animal models, such as in vitro cell cultures. The practical application of such methodologies may help solve the economic problem related to very expensive in vivo toxicology studies and implement preventive methods based on the calculated data and known mechanism of action of individual or combined toxins easily studied in vitro. The nature of coexistence of many types of mycotoxins in complex environmental samples, such as food and water, has been reported worldwide. How these mycotoxins might affect human health in combination is largely unknown. This study had, as a goal, to test the toxicity of the four aflatoxins and aflatoxin combination on human cells. Due to the lack of aflatoxins mixture data regarding the human cytotoxicity, the aim of this study was to specify, evaluate and predict the combined effects of mycotoxin mixtures.

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