Cytotoxic and inflammatory effects of individual and combined exposure of HepG2 cells to zearalenone and its metabolites

Zearalenone (ZEA), a mycotoxin produced by several Fusarium spp., is most commonly found as a contaminant in stored grain. ZEA derivatives (alpha-zearalenol (alpha-ZOL), beta-zearalenol (beta-ZOL)) can also be produced by Fusarium spp. in corn stems infected by fungi in the field. Also, following oral exposure, zearalenone is metabolized in various tissues, particularly in the liver, the major metabolites being alpha-ZOL and beta-ZOL. The co-exposure of cells to mixture of a combination of mycotoxins may cause an increase of toxicity produced by these mycotoxins. In this in vitro study, we investigated the combined effects of ZEA, alpha-ZOL, beta-ZOL in binary mixtures on the viability and inflammatory response of human liver cancer cell line (HepG2). Cell viability was assessed after 72 h using a neutral red assay. Effect of the toxins and their binary combinations on the expression of genes involved in inflammation (IL-1 beta, TNF-alpha, and IL-8) were assessed through qPCR. Our viability data showed that irrespective of the toxin combinations, the toxins have synergistic effect. ZEA + alpha-ZOL and ZEA + beta-ZOL mixtures have induced a slight to high antagonistic response on inflammatory cytokines at low concentrations that have turned into strong synergism for high concentrations. alpha-ZOL + beta-ZOL showed antagonistic effects on inflammation for IL-1 beta and TNF-alpha, but act synergic for IL-8 at high toxin concentrations. This study clearly shows that co-contamination of food and feed with ZEA metabolites should be taken into consideration, as the co-exposure to mycotoxins might result in stronger adverse effect than resulted from the exposure to individual toxin.