Effects of mycotoxins on cytokine production and proliferation in EL-4 thymoma cells

The thymoma cell line EL4.IL-2 (EL-4) was used as a T-cell model to assess the immunotoxic effects of several mycotoxins produced by the Aspergillus-Penicillium and the Fusarium groups. EL-4 cells were stimulated with phorbol 12-myristate 13-acetate (PMA) in the presence of mycotoxins at various concentrations for 5 d and culture supernatants were analyzed for interleukins (IL) IL-2 and IL-5 by enzyme-linked immunosorbent assay (ELISA). The cytokine effects were further related to proliferation and cell viability using the MTT [3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyltetrazolium bromide] assay with absorbance at 570 nm (A(570)) as the endpoint indicator. IL-2 and IL-5 levels were dramatically increased by cyclopiazonic acid at 50-1000 ng/ml, whereas IL-2 was significantly decreased at 10 mu g/ml. Proliferation was slightly increased at 100-1000 ng/ml cyclopiazonic acid but markedly depressed at 5 and 10 mu g/ml. When EL-4 cells were exposed to 5 and 10 mu g/ml of ochratoxin A, IL-2 production was markedly increased while IL-5 production was significantly decreased. The A(570) was significantly decreased by ochratoxin A at 10 mu g/ml. IL-2 and IL-5 production was almost totally suppressed by patulin at concentrations greater than or equal to 500 ng/ml and by T-2 toxin at greater than or equal to 5 ng/ml. These effects occurred concurrently with marked depression of A(570) in the MTT assay. Although A(570) was unaffected by either zearalenone or alpha-zearalenol exposure, both IL-2 and IL-5 levels were significantly elevated by these toxins at 5 or 10 mu g/ml. IL-2 and IL-5 production were not affected in EL-4 cells cultured with either the Aspergillus-Penicillium toxins aflatoxin B-1 and secalonic acid or the Fusarium toxins wortmannin, fumonisin B-1, or fusaric acid at concentrations up to 70 mu g/ml. In total, the EL-4 culture studies indicated that cyclopiazonic acid, ochratoxin A, zearalenone, and alpha-zearalenol could stimulate cytokine production whereas patulin and T-2 toxin were inhibitory. Cytokine dysregulation was not always related directly to perturbations in proliferation. The results suggest that the EL-4 thymoma cell line could be a simple and effective in vitro model for evaluating immunotoxicity of various classes of environmental chemicals.