Insights into cellular metabolic pathways of the combined toxicity responses of Caco-2 cells exposed to deoxynivalenol, zearalenone and Aflatoxin B.

Metabolic profiling in Caco-2 cells was studied for the combined toxic effects of deoxynivalenol (DON), zearalenone (ZEN), and Aflatoxin B (AFB) through untargeted GC-MS. The GC-MS spectra of Caco-2 cells treated with individual 6.7 μM DON, 20 μM ZEN, 20 μM AFB and the combined DON + AFB (6.7 + 20 μM) and DON + ZEN + AFB (6.7 + 20 + 20 μM) for 24 h were deconvoluted, aligned and identified with MS DIAL. The metabolic pathway analysis was analyzed with MetaMapp and visualized with CytoScape. Results show that the combined DON + AFB and DON + ZEN + AFB treatment has an obvious "synergistic effect". The apoptosis-related gene mRNA test result indicates that the combined mycotoxins downregulate Bcl-2 gene and upregulate Bax, p53, caspase-3, caspase-8 and caspase-utilized 9 genes, more significantly than any individual mycotoxins group. The cellular metabolism illustrated that the combined mycotoxin groups, DON + ZEN + AFB seriously effect glycine, serine and threonine metabolism, pyruvate metabolism, etc. while no metabolic disorders were presented in individual mycotoxin group. Our hypothesis was validated that the combined mycotoxins with low concentrations can have a synergistic effect in the metabolism, which may lead to cellular apoptosis or necrosis.