MEASUREMENT OF NEUTROPHIL ACTIVATION AND EPIDERMAL-CELL TOXICITY BY PALYTOXIN AND 12-O-TETRADECANOYLPHORBOL-13-ACETATE

Palytoxin is a human and mouse skin irritant and complete mouse skin tumor promoter that differs from the well-studied phorbol ester class of tumor promoters in many respects. In this study, we have found palytoxin to stimulate the production of superoxide by isolated human neutrophils using electron paramagnetic resonance (EPR) spectroscopy with the spin-trap DMPO. This stimulation of oxyradical production by palytoxin is relatively weak, however, when compared to that by 12-O-tetradecanoylphorbol-13-acetate (TPA). The maximal amount of oxyradicals produced by palytoxin-stimulated neutrophils is 10(-4) mumol/10(6) neutrophils, and this stimulation requires nanomolar concentrations of palytoxin, with half maximal stimulation at concentrations of approximately 30 nM. In contrast, the tumor promoter TPA causes human neutrophils to generate in excess of 10(-3) mumol oxyradicals/10(6) neutrophils with concentrations as low as 1 nM. Toxicity to cultured human epidermal cells was observed at very low concentrations of palytoxin, with 50% loss of colony-forming efficiency observed at approximately 3 x 10(-13) M. For TPA, 50% loss of colony-forming efficiency for cultured epidermal cells requires approximately 5 nM. Thus, although palytoxin stimulates superoxide production in isolated neutrophils, epidermal cells are sensitive at much lower concentrations and are likely to be the important target cell in vivo. This is in contrast to TPA, where neutrophils are stimulated at concentrations less than those required to produce pathological effects on epidermal cells, suggesting that neutrophils may be an important target cell for TPA in vivo.