Differential impacts of individual and combined exposures of deoxynivalenol and zearalenone on the HepaRG human hepatic cell proteome.

Numerous surveys have highlighted the natural co-occurrence of deoxynivalenol (DON) and zearalenone (ZEA) mycotoxins in food and feed. Nevertheless, data regarding cellular mechanisms involved in response to their individual and simultaneous exposures are lacking. In this study, in order to analyze how low mycotoxin doses could impact cellular physiology and homeostasis, proteomic profiles of proliferating human hepatic cells (HepaRG) exposed for 1h and 24h to low DON and ZEA cytotoxicity levels (0.2 and 20μM respectively), alone or in combination, were analyzed by LC-MS/MS. Proteome analyses of mycotoxin-treated cells identified 4000 proteins with about 1.4% and 3.7% of these proteins exhibiting a significantly modified abundance compared to controls after 1h or 24h, respectively. Analysis of the Gene Ontology biological process annotations showed that cell cycle, proliferation and/or development as well as on DNA metabolic processes were affected for most treatments. Overall, different proteins, and thus biological processes, were impacted depending on the considered mycotoxin and exposure duration. Finally, despite the important proteome changes observed following 24h exposure to both mycotoxins, only the uptake of ZEA by the cells was suggested by the mycotoxin quantification in cell supernatants.

BIOLOGICAL SIGNIFICANCE: This study investigated the proteomic changes that occurred after DON and ZEA (individually and in combination) short exposures at low cytotoxicity levels in proliferating HepaRG cells using LC-MS/MS. The obtained results showed that the cellular response is time- and mycotoxin or mixture-dependent. In particular, after 1h exposure, the DON+ZEA combination led to more proteomic changes than DON or ZEA alone, whereas the opposite was observed after 24h. In addition, the significant cellular response to stress induced by ZEA after 24h exposure seemed to be reduced when combined with DON. Thus, these results supported a possible mitigation by the hepatocytes when exposed to the mycotoxin mixture for a long duration. These findings represent an essential step to further explore adaptive cell response to mycotoxin exposure using with more complex incubation kinetics and combining different "omics" tools. Moreover, as mycotoxin quantification in cell supernatants showed different behaviors for DON and ZEA, this also raises the question about how mycotoxins actually trigger the cell response.