The induction by 60Co gamma-rays of DNA breaks, revealed by relaxation (single-strand breaks, SSBs) and linearization (double-strand breaks, DSBs) of supercoiled plasmid DNA, was measured under three irradiation conditions, the DNA being in a dry, humid, or aqueous state in the absence of oxygen, at 25 or -196 degrees C (77 K). Yields of strand breaks (3.0 x 10(-10) SSB/Gy.Da and 2.6 x 10(-11) DSB/Gy.Da) in DNA exposed to a stream of humidified nitrogen were higher than those in the dry condition (5.7 x 10(-11) SSB/Gy.Da and 3.2 x 10(-12) DSB/Gy.Da), but both these yields were markedly lower than those measured for DNA in aqueous solution at a concentration of 73 micrograms/cm3 (1.14 x 10(-7) SSB/Gy.Da and 5.4 x 10(-9) DSB/Gy.Da). Over 100-fold fewer SSBs were observed in the frozen aqueous system compared with the non-frozen liquid state, whereas in the dry and humid states, freezing did not affect the yield as much. The same trend was observed for DSBs. However, the induction of SSBs was more affected than that of DSBs by freezing in the aqueous systems. An interesting reverse relationship was observed in humid systems. The observed linearity of DSB induction with radiation dose supported a single-event mechanism. A comparison of G values for humid systems revealed that the role of bound water in radiation damage becomes significant in the nonfrozen state. Based on these and other measurements of strand breaks under different conditions, the significance of bound and free water on the yields of DNA strand breaks by gamma-rays is discussed, and the relevance of these results to the in vivo situation outlined.
Action spectra were determined for cell killing and mutation by monochromatic ultraviolet and visible radiations (254-434 nm) in cultured human epithelial P3 cells. Cell killing was more efficient following radiation at the shorter wavelengths (254-434 nm) than at longer wavelengths (365-434 nm). At 254 nm, for example, a fluence of 11 Jm-2 gave 37% cell survival, while at 365 nm, 17 X 10(5) Jm-2 gave equivalent survival. At 434 nm little killing was observed with fluences up to 3 X 10(6) Jm-2. Mutant induction, determined at the hypoxanthine-guanine phosphoribosyltransferase locus, was caused by radiation at 254, 313, and 365 nm. There was no mutant induction at 334 nm although this wavelength was highly cytotoxic. Mutagenesis was not induced by 434 nm radiation, either. There was a weak response at 405 nm; the mutant frequencies were only slightly increased above background levels. For the mutagenic wavelengths, log-log plots of the mutation frequency against fluence showed linear regressions with positive slopes of 2.5, consistent with data from a previous study using Escherichia coli. The data points of the action spectra for lethality and mutagenesis were similar to the spectrum for DNA damage at wavelengths shorter than 313 nm, whereas at longer wavelengths the lethality spectrum had a shoulder, and the mutagenesis spectrum had a secondary peak at 365 nm. No correlation was observed for the P3 cells between the spectra for cell killing and mutagenesis caused by wavelengths longer than 313 nm and the induction of DNA breakage or the formation of DNA-to-protein covalent bonds in these cells.
Abstract— The reduced pyridine coenzymes NADPH and NADH produced superoxide anion(“CK”) from ground state molecular oxygen when irradiated by ultraviolet (UV) radiation extending from 290 to 405 nm as detected by cytochrome c reduction. Superoxide dismutase (SOD), but not catalase or heat‐inactivated SOD, decreased the amount of cytochrome c reduced, indicating that O2− was responsible for the reduction of cytochrome c. Decreased oxygen tension during irradiation also inhibited production of O2−. Quantum yields for the production of the anion were in the region of 10−7 to 10−9 mol per photon. These data indicate that NADH and NADPH can act as type II photosensitizers of both far‐and near‐UV radiation, and that the deleterious biological effects of exposure to these radiations such as erythema and dermal carcinogenesis may be mediated at least in part through the generation of O2−.
Abstract— An action spectrum for the immediate induction in DNA of single‐strand breaks (SSBs, frank breaks plus alkali‐labile sites) in human P3 teratoma cells in culture by monochromatic 254‐, 270‐, 290‐, 313‐, 334‐, 365‐, and 405‐nm radiation is described. The cells were held at +0.5d̀C during irradiation and were Iysed immediately for alkaline sedimentation analysis following the irradiation treatments. Linear fluence responses were observed over the fluence ranges studied for all energies. Irradiation of the cells in a D2O environment (compared with the normal H2O environment) did not alter the rate of induction of SSBs by 290‐nm radiation, whereas the D2O environment enhanced the induction of SSBs by 365‐ and 405‐nm irradiation. Analysis of the relative efficiencies for the induction of SSBs, corrected for quantum efficiency and cellular shielding, revealed a spectrum that coincided closely with nucleic acid absorption below 313 nm. At longer wavelengths, the plot of relative efficiency vs. wavelength contained a minor shoulder in the same wavelength region as that observed in a previously obtained action spectrum for stationary phase Bacillus subtilis cells. Far‐UV radiation induced few breaks relative to pyrimidine dimers, whereas in the near‐UV region of radiation, SSBs account for a significant proportion of the lesions relative to dimers, with a maximum number of SSBs per lethal event occurring at 365‐nm radiation.
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