The cytotoxic effects of T-2 toxin on normal human lymphocytes.
T -2 toxin is an immunosuppressive mycotoxin that has been conjoined with several symptoms and diseases as early as the turn of the century, but whose mechanisms of action are still being investigated. Accordingly, this study was an attempt to determine the cytotoxic effects of T -2 toxin on normal human lymphocytes in vitro, with particular emphasis on mitochondrial viability, cellular and nuclear morphology as well as the localisation of the subcellular sites of toxin interaction. The cytotoxicity of T -2 toxin was assessed with the use of a methylthiazol tetrazolium (MTT) assay. This assay targeted the succinate dehydrogenase activity of the lymphocytic mitochondria, over a range of concentrations of T-2 toxin at various incubation times. The morphology of treated lymphocytes was analysed with the use of transmission electron microscopy and the localisation of the toxin was accomplished via immunocytochemistry. DNA fragmentation studies formed an integral part of the analyses. The cytotoxicity assay indicated that not only was cell viability inversely proportional to both the dose and exposure time, but that the eftects of the different doses were only evident at prolonged incubation times (12-24 hours). The electron microscopy studies showed that T-2 toxin (1,56 ug/ml) induced apoptosis (cell suicide) in normal human lymphocytes. This was determined by the observation of chromatin condensation and nuclear disintegration within the toxin treated lymphocytes. Apoptosis seemed to occur independently of mitochondrial damage at 6 hours of exposure to T-2 toxin. The presence of polyribosomes within the treated lymphocytes indicated that protein synthesis was not inhibited. Anti-T-2 toxin conjugated gold label was present in all areas of damage, particularly within the nuclei of the T-2 toxin treated lymphocytes. The DNA fragmentation results showed that T-2 toxin induced fragmentation in lymphocytes, the extent of which was directly proportional to the exposure time. It appears that the early signs of T-2 toxin induced apoptosis in normal human lymphocytes can be determined by damage to the nucleus.