Synergistic Action of the Nephrotoxic Mycotoxins Ochratoxin A and Citrinin at Nanomolar Concentrations in Human Proximal Tubule-derived Cells

Increased ochratoxin A (OTA) or citrinin (CIT) concentrations in food correlate with increased prevalence of tubule-interstitial nephropathy. We tested the hypothesis that co-exposure of human proximal tubule-derived epithelial cells (HK-2) to OTA and CIT promotes synergistic events indicative for inflammation, epithelial-to-mesenchymal-transition (EMT) or fibrosis. We measured markers of inflammation, EMT and fibrosis and investigated the role of MAP-kinases. Only concurrent but not individual exposure to OTA and CIT at nanomolar concentrations led to (i) an increase of TNF protein and mRNA, (ii) a decrease of COX-2 protein and mRNA, (iii) a decrease of E-cadherin protein and (iv) an increase of vimentin and α-SMA protein. Cell shape shifted from a cobblestone- to a spindle-like phenotype indicating EMT. Extra- and intracellular collagen III protein content was increased. Concomitant mRNA expression changes were observed for TNF, COX-2, E-cadherin and α-SMA indicating transcriptional regulation. This was not the case for vimentin and collagen III mRNA indicating posttranscriptional regulation. Inhibition of ERK 1/2 and JNK 1/2 reduced the effect on TNF but not on α-SMA mRNA indicating an involvement of these kinases. Phosphorylation of ERK1/2 was increased by CIT, OTA alone and the mycotoxin combination. In contrast, the phosphorylation of JNK1/2 was unchanged. In conclusion, nanomolar OTA and CIT act synergistically favouring nephropathic processes.