Abstract
The release of tumor necrosis factor α (TNF-α) by ochratoxin A (OTA) was studied in various macrophage and non-macrophage cell lines and compared with E. coli lipopolysaccharide (LPS) as a standard TNF-α release agent. Cells were exposed either to 0, 2.5 or 12.5 µmol/L OTA, or to 0.1 µg/mL LPS, for up to 24 h. OTA at 2.5 µmol/L and LPS at 0.1 µg/mL were not toxic to the tested cells as indicated by viability markers. TNF-α was detected in the incubated cell medium of rat Kupffer cells, peritoneal rat macrophages, and the mouse monocyte macrophage cell line J774A.1: TNF-α concentrations were 1,000 pg/mL, 1,560 pg/mL, and 650 pg/mL, respectively, for 2.5 µmol/L OTA exposure and 3,000 pg/mL, 2,600 pg/mL, and 2,115 pg/mL, respectively, for LPS exposure. Rat liver sinusoidal endothelial cells, rat hepatocytes, human HepG2 cells, and mouse L929 cells lacked any cytokine response to OTA, but showed a significant release of TNF-α after LPS exposure, with the exception of HepG2 cells. In non-responsive cell lines, OTA lacked both any activation of NF-κB or the translocation of activated NF-κB to the cell nucleus, i.e., in mouse L929 cells. In J774A.1 cells, OTA mediated TNF-α release via the pRaf/MEK 1/2-NF-κB and p38-NF-κB pathways, whereas LPS used pRaf/MEK 1/2–NF-κB, but not p38-NF-κB pathways. In contrast, in L929 cells, LPS used other pathways to activate NF-κB. Our data indicate that only macrophages and macrophage derived cells respond to OTA and are considered as sources for TNF-α release upon OTA exposure.
Highlights
Ochratoxin A (OTA) is a natural mycotoxin produced from Aspergillus and Penicillium species.OTA causes diverse toxicological responses: it is genotoxic, carcinogenic, nephrotoxic, hepatotoxic, embryotoxic, teratogenic, and immunotoxic
Our previous studies have shown that OTA releases tumor necrosis factor α (TNF-α) from blood-free perfused rat livers in a dose- and time-dependent fashion without effects on liver vitality [7,9,13]
LPS did not alter the survival of HepG2 cells, L929 cells, and J774A.1 cells in comparison with untreated cells
Summary
Ochratoxin A (OTA) is a natural mycotoxin produced from Aspergillus and Penicillium species. Previous risk assessments have studied its genotoxicity and OTA-mediated nephrotoxicity [1,2,3,4]. A marked release of TNF-α and IL-6 has been observed during perfusion of the isolated rat liver with OTA via the portal vein [8]. In blood-free perfused rat livers, a synergistic effect on OTA-mediated TNF-α release by co-addition of low doses of E. coli lipopolysaccharides (LPS) has been observed [10] the question arises whether in general OTA mimics LPS-induced TNF-α release in all LPS-sensitive macrophage and non-macrophage cells, indicating a release pathway similar to the LPS-triggered cascade [11,12] in this study we compared the ability of OTA to release TNF-α from macrophage and non-macrophage cells with LPS, and investigated the possible mediator mechanism of this release
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