Abstract
The radioprotector 2-[(aminopropyl)amino] ethanethiol (WR1065) was investigated with respect to its ability to affect radiation-induced DNA damage and repair in V79 cells. Studies were performed to evaluate the protector under conditions in which it is known to be effective in reducing the cytotoxic and mutagenic effects of gamma-irradiation. At a concentration of 4 mM, WR1065 protected against the formation of single strand breaks (SSB), as determined by the method of alkaline elution, when it was present during irradiation. The protector appeared, however, to inhibit the subsequent postirradiation repair or rejoining of SSB. While repair was complete within 24 h, the protector reduced the rate of repair by a factor of 3. This inhibitory effect on the rate of repair did not correlate with either measured differences in cell survival or mutagenesis. The radioprotector was also investigated with respect to its ability to affect cell cycle progression. WR1065 present in the growth medium inhibited the progression of cells through S-phase, and cell-doubling time following a 3 h exposure to the protector was increased from 11 to 18 h. These data are consistent with the well characterized property of thiols to inhibit DNA polymerase activity. It was concluded that, while the presence of WR1065 during irradiation reduced SSB-DNA damage, its effect on the subsequent rejoining of these breaks could not be correlated with its observed effect on protecting against radiation-induced mutagenesis. It may be that the inhibition of cell-cycle progression by the protector allowed more time to enhance the fidelity of repair as measured by the protector's ability to protect against radiation-induced mutagenesis.
Highlights
Since it has been reported that thiol-containing compounds such as cysteamine can induce single-strand-break (SSB) damage in DNA as well as inhibit its repair (Sawada & Okada, 1970), we have focused this investigation towards characterizing the role of the radioprotector WR1065 with respect to (a) the induction of single strand breaks (SSB) in DNA of unirradiated and irradiated cells, (b) the rejoining or repair of that damage, and (c) the modulation of cell-cycle kinetics
To determine whether the radioprotector WR1065 can induce SSB in DNA of exposed V79 cells, cells were treated with 4 mM for times ranging from 30 min to 180 min
Cells irradiated in the presence of WR1065 appeared to have fewer SSB
Summary
V79-B310H Chinese hamster cells were maintained as stock cultures in a minimal essential medium (Gibco) with 10% foetal calf serum (Biologos) in a humidified atmosphere containing 5% CO2 and 95% air at 37°C. Growing cells were harvested and irradiated at ice bath temperatures using 60Co yrays from a Gamma beam 650 irradiator (Atomic Energy of Canada) at a dose rate of 0.5Gymin-m (Han et al, 1980). With respect to dose-response studies, irradiated cells were immediately diluted into ice-cold Solution A (8.0 g NaCl, 0.4 g KCl, 1.0 g glucose, 0.35gNaHCO3, per litre) containing 5mM EDTA to insure an inhibition of DNA repair (Meyn & Jenkins, 1983). DNA repair studies were performed using cells irradiated with 10 Gy. Following irradiation, cells were added to prewarmed culture medium and were incubated at 37°C for the desired time
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