Efficiency of radiation-induced base lesion excision and the order of enzymatic treatment.

PURPOSE: To clarify whether initial base excision repair processes at clustered DNA damage sites comprising multiple base lesions affect subsequent excision processes via the formation of additional strand breaks by glycosylase and apurinic/apyrimidinic (AP) endonuclease base excision enzymes. MATERIALS AND METHODS: Plasmid DNA (pUC18) as a model DNA molecule was exposed to high-linear-energy-transfer (LET) ionizing radiation (He2+ or C6+ ions) or low-LET ionizing radiation (X-rays) under various conditions to produce varied radical-scavenging effects. pUC18 was then treated sequentially or simultaneously with two bacterial base excision enzymes (glycosylases), namely, endonuclease III and formamidopyrimidine-DNA glycosylase, which convert pyrimidine (or abasic [AP] site) and purine (or AP site) lesions to single-strand breaks (SSB), respectively. Yields of additional SSB or double-strand breaks (DSB) as digestion products were examined after changing the order of enzymatic treatment. RESULTS: There were few differences among the enzymatic treatments, indicating that treatment order did not affect the final yields of additional SSB or DSB formed by glycosylase activity. This suggests that of the total damage, the fraction of clustered damage sites with a persistent base lesion dependent on the order of glycosylase treatment was insignificant if present. CONCLUSION: Base lesion clusters induced by high- or low-LET radiation appear three or more base pairs apart, and are promptly converted to a DSB by glycosylase, regardless of the order of enzymatic treatment.

Gamma radiation at a human relevant low dose rate is genotoxic in mice.

Even today, 70 years after Hiroshima and accidents like in Chernobyl and Fukushima, we still have limited knowledge about the health effects of low dose rate (LDR) radiation. Despite their human relevance after occupational and accidental exposure, only few animal studies on the genotoxic effects of chronic LDR radiation have been performed. Selenium (Se) is involved in oxidative stress defence, protecting DNA and other biomolecules from reactive oxygen species (ROS). It is hypothesised that Se deficiency, as it occurs in several parts of the world, may aggravate harmful effects of ROS-inducing stressors such as ionising radiation. We performed a study in the newly established LDR-facility Figaro on the combined effects of Se deprivation and LDR γ exposure in DNA repair knockout mice (Ogg1(-/-)) and control animals (Ogg1(+/-)). Genotoxic effects were seen after continuous radiation (1.4 mGy/h) for 45 days. Chromosomal damage (micronucleus), phenotypic mutations (Pig-a gene mutation of RBC(CD24-)) and DNA lesions (single strand breaks/alkali labile sites) were significantly increased in blood cells of irradiated animals, covering three types of genotoxic activity. This study demonstrates that chronic LDR γ radiation is genotoxic in an exposure scenario realistic for humans, supporting the hypothesis that even LDR γ radiation may induce cancer.

Site-directed photoproteolysis of 8-oxoguanine DNA glycosylase 1 (OGG1) by specific porphyrin-protein probe conjugates: a strategy to improve the effectiveness of photodynamic therapy for cancer.

The specific light-induced, non-enzymatic photolysis of mOGG1 by porphyrin-conjugated or rose bengal-conjugated streptavidin and porphyrin-conjugated or rose bengal-conjugated first specific or secondary anti-IgG antibodies is reported. The porphyrin chlorin e6 and rose bengal were conjugated to either streptavidin, rabbit anti-mOGG1 primary specific antibody fractions or goat anti-rabbit IgG secondary antibody fractions. Under our experimental conditions, visible light of wavelengths greater than 600 nm induced the non-enzymatic degradation of mOGG1 when this DNA repair enzyme either directly formed a complex with chlorin e6-conjugated anti-mOGG1 primary specific antibodies or indirectly formed complexes with either streptavidin-chlorin e6 conjugates and biotinylated first specific anti-mOGG1 antibodies or first specific anti-mOGG1 antibodies and chlorin e6-conjugated anti-rabbit IgG secondary antibodies. Similar results were obtained when rose bengal was used as photosensitizer instead of chlorin e6. The rate of the photochemical reaction of mOGG1 site-directed by all three chlorin e6 antibody complexes was not affected by the presence of the singlet oxygen scavenger sodium azide. Site-directed photoactivatable probes having the capacity to generate reactive oxygen species (ROS) while destroying the DNA repair system in malignant cells and tumors may represent a powerful strategy to boost selectivity, penetration and efficacy of current photodynamic (PDT) therapy methodologies.

Effects of low-dose gamma radiation on DNA damage, chromosomal aberration and expression of repair genes in human blood cells.

Exposure to low-dose gamma radiation is common in certain occupations but the biological and health effects from such exposure remain to be determined. The aim of this study was to investigate the effects of low-dose gamma radiation on DNA damage, chromosomal aberration and DNA repair gene expressions in whole blood and peripheral lymphocytes. The study revealed a dose-dependent effect of gamma radiation on DNA damage. Significant increases in DNA strand breaks and oxidative base damage, determined as formamidopyrimidine-DNA-glycosylase (FPG)-sensitive sites, were observed at absorbed doses of 5 and 10cGy, respectively. However, gamma radiation at doses up to 500cGy did not significantly increase the level of 8-oxo-7, 8-dihydro-2'-deoxyguanosine (8-oxodG) determined by HPLC with electrochemical detection (HPLC-ECD). Gamma radiation as low as 5cGy caused chromosomal aberrations determined as dicentric and deletion frequencies. This finding is significant since the genotoxic effects of gamma radiation can be observed even at a low dose of 5cGy. Furthermore, gamma radiation decreased the mRNA expression of both hOGG1 and XRCC1 repair genes determined by reverse transcriptase-polymerase chain reaction (RT-PCR), with a significant decrease of expression being observed at 20cGy. The expression levels of hOGG1 and XRCC1 mRNA were inversely correlated with the levels of FPG-sensitive sites and DNA strand breaks. The finding of decreased expression levels for hOGG1 and XRCC1 in gamma-irradiated lymphocytes has not been reported elsewhere. Our observations suggest that the genotoxic effects of gamma radiation may be due to a combination of DNA-damaging effects and reduced DNA repair capacity, and may explain the significant increase in health risk from high doses of ionizing radiation.