Background: The nitroxide Tempol (4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl) is a stable, free radical that exhibits protection from ionizing radiation damage and from oxidative stress mediated through exposure of cells to superoxide or hydrogen peroxide. Radiation protection has been observed in both in vivo and in vitro models. To understand the mechanism of Tempol-mediated radioprotection better, the production of radiation-induced chromosome aberrations was evaluated. This study analyzed Tempol-mediated radioprotection of human peripheral blood lymphocytes (PBLs).
Methods: Peripheral blood lymphocytes were exposed to control (0mM), 10 mM (Tp10), and 50 mM (Tp50) concentrations of Tempol for 20 minutes before irradiation with 0, 150, 300, and 450 cGy. One quarter ml whole blood was cultured in F12 medium and phytohemagglutinin at 37 degrees C for 49, 54, 59, and 64 hours. Colcemide was added to each sample for the last 5 hours before harvest. Cells were harvested, treated with hypotonic solution, and fixed before dropping on cold clean slides. Mitotic indices and frequency of dicentric, ring, and triradial chromosomal aberrations were determined at 1000x magnification for each treatment group at each collection point.
Results: Treatment of cells with Tempol alone did not induce the chromosomal aberration frequency above that for unirradiated controls. Radiation dose response curves for total chromosome aberration production revealed radioprotection for Tempol treatment for both 10 and 50 mM exposures. Tempol protection factors (assessed at 0.2 aberrations/cell level) for Tp 10 and Tp 50 were 2.2 and 2.8, respectively.
Conclusions: Tempol protects against radiation-induced chromosome aberrations in human PBLs. This finding is consistent with and lends support to previous studies in which Tempol was reported to enhance cell survival and reduce radiation-induced DNA double strand breaks.