Radiation-induced DNA Damage and Repair in Lens Epithelial Cells of both Ptch1(+/-) and Ercc2(+/-) Mutated Mice.

Epidemiological studies suggest an increased incidence and risk of cataract after low-dose (<2 Gy) ionizing radiation exposures. However, the biological mechanism(s) of this process are not fully understood. DNA damage and repair are thought to have a contributing role in radiation-induced cataractogenesis. Recently we have reported an inverse dose-rate effect, as well as the low-dose response, of DNA damage and repair in lens epithelial cells (LECs). Here, we present further initial findings from two mutated strains (Ercc2+/- and Ptch1+/-) of mice, both reportedly susceptible to radiation-induced cataract, and their DNA damage and repair response to low-dose and low-dose-rate gamma rays. Our results support the hypothesis that the lens epithelium responds differently to radiation than other tissues, with reported radiation susceptibility to DNA damage not necessarily translating to the LECs. Genetic predisposition and strain(s) of mice have a significant role in radiation-induced cataract susceptibility.

Radiation-Induced Lens Opacity and Cataractogenesis: A Lifetime Study Using Mice of Varying Genetic Backgrounds.

Recent epidemiological findings and reanalysis of historical data suggest lens opacities resulting from ionizing radiation exposures are likely induced at lower doses than previously thought. These observations have led to ICRP recommendations for a reduction in the occupational dose limits for the eye lens, as well as subsequent implementation in EU member states. The EU CONCERT LDLensRad project was initiated to further understand the effects of ionizing radiation on the lens and identify the mechanism(s) involved in radiation-induced cataract, as well as the impact of dose and dose-rate. Here, we present the results of a long-term study of changes to lens opacity in male and female adult mice from a variety of different genetic (radiosensitive or radioresistant) backgrounds, including mutant strains Ercc2 and Ptch1, which were assumed to be susceptible to radiation-induced lens opacities. Mice received 0.5, 1 and 2 Gy 60Co gamma-ray irradiation at dose rates of 0.063 and 0.3 Gy min-1. Scheimpflug imaging was used to quantify lens opacification as an early indicator of cataract, with monthly observations taken postirradiation for an 18-month period in all strains apart from 129S2, which were observed for 12 months. Opacification of the lens was found to increase with time postirradiation (with age) for most mouse models, with ionizing radiation exposure increasing opacities further. Sex, dose, dose rate and genetic background were all found to be significant contributors to opacification; however, significant interactions were identified, which meant that the impact of these factors was strain dependent. Mean lens density increased with higher dose and dose rate in the presence of Ercc2 and Ptch1 mutations. This project was the first to focus on low (<1 Gy) dose, multiple dose rate, sex and strain effects in lens opacification, and clearly demonstrates the importance of these experimental factors in radiobiological investigations on the lens. The results provide insight into the effects of ionizing radiation on the lens as well as the need for further work in this area to underpin appropriate radiation protection legislation and guidance.

Influence of Select Antibiotics on Vibrio fischeri and Desmodesmus subspicatus at µg L-1 Concentrations.

The presence of pharmaceuticals in the aquatic environment is a contemporary reality and it is necessary to understand more about the effects of this presence on organisms. The purpose of this work was to assess the ecotoxicity of antibiotics metronidazole, nitrofurantoin, trimethoprim, and sulphamethoxazole (single and mixture) in Vibrio fischeri and Desmodesmus subspicatus at µg L-1 concentrations. The evaluation of the toxic effect of the antibiotics on V. fischeri and D. subspicatus was based on fluorescence and bioluminescence tests, respectively, using nominal concentrations. When tested individually, the four antibiotics gave rise to a toxic effect on the evaluated organisms. Sulphamethoxazole caused a higher toxic effect on V. fischeri and D. subspicatus from 7.81 to 500 µg L-1. Trimethoprim and sulphamethoxazole showed hormesis for the concentrations, which ranged from 7.81 to 62.5 µg L-1. The mixture of antibiotics induced a toxic effect on the V. fischeri and D. subspicatus organisms (from 0.03 to 1 µg L-1 concentrations) than when the antibiotics were evaluated individually. These results were significant since water quality problems are widespread all over the word, and emerging pollutants such as antibiotics have been detected in the aquatic environment in very low concentrations.

The German Mouse Clinic: a platform for systemic phenotype analysis of mouse models.

The German Mouse Clinic (GMC) is a large scale phenotyping center where mouse mutant lines are analyzed in a standardized and comprehensive way. The result is an almost complete picture of the phenotype of a mouse mutant line--a systemic view. At the GMC, expert scientists from various fields of mouse research work in close cooperation with clinicians side by side at one location. The phenotype screens comprise the following areas: allergy, behavior, clinical chemistry, cardiovascular analyses, dysmorphology, bone and cartilage, energy metabolism, eye and vision, host-pathogen interactions, immunology, lung function, molecular phenotyping, neurology, nociception, steroid metabolism, and pathology. The German Mouse Clinic is an open access platform that offers a collaboration-based phenotyping to the scientific community (www.mouseclinic.de). More than 80 mutant lines have been analyzed in a primary screen for 320 parameters, and for 95% of the mutant lines we have found new or additional phenotypes that were not associated with the mouse line before. Our data contributed to the association of mutant mouse lines to the corresponding human disease. In addition, the systemic phenotype analysis accounts for pleiotropic gene functions and refines previous phenotypic characterizations. This is an important basis for the analysis of underlying disease mechanisms. We are currently setting up a platform that will include environmental challenge tests to decipher genome-environmental interactions in the areas nutrition, exercise, air, stress and infection with different standardized experiments. This will help us to identify genetic predispositions as susceptibility factors for environmental influences.

Radical intermediates during the oxidation of nitropropanes. The formation of NO2 from 2-nitropropane, its reactivity with nucleosides, and implications for the genotoxicity of 2-nitropropane.

The chemistry of the nonenzymatic oxidation of the rat liver carcinogen, 2-nitropropane, and its anionic form, propane-2-nitronate, was investigated using pulse radiolysis and EPR/spin trapping with 3,5-dibromo-4-nitrosobenzenesulfonic acid as the trapping agent. The results suggest that, following initial oxidation to a secondary alkyl radical, propane-2-nitronate is effectively degraded in a peroxidative chain reaction with the intermediary formation of peroxyl and NO2.radicals. The latter radical was shown to react appreciably fast with ribonucleosides, deoxyribonucleosides, and guanosine nucleotides. It is proposed that nonenzymatic formation of NO2.radicals after enzymatic oxidation of propane-2-nitronate to the corresponding secondary alkyl radical accounts for the induction of DNA damage observed after exposure of rats to 2-nitropropane.

Propane 2-nitronate is more rapidly denitrified and is more genotoxic than 2-nitropropane in cultured rat hepatoma cells.

We have investigated the importance of nitronate formation from 2-nitropropane (2-NP) for the oxidative metabolism and the genotoxicity of 2-NP in 2sFou rat hepatoma cells. Treatment of the cells with 2-NP for up to 3 h resulted in the time-dependent appearance of nitrite in the culture medium and in a weak induction of DNA repair synthesis. Both nitrite formation and repair induction were markedly enhanced in cells exposed to the anionic form of 2-NP, propane 2-nitronate. These observations suggest that propane 2-nitronate, rather than 2-NP itself, is oxidized by the liver cells to yield a DNA-damaging product. The results also indicate that the nitro/nitronate equilibration in intact liver cells is slow, suggesting that nitronate formation represents the rate-limiting step in the metabolic activation of 2-NP.