Effect of Oxygen on Radiation-Induced DNA Damage in Isolated Nuclei

Horan, Annamarie D.; Giandomenico, Albert R.; Koch, Cameron J.

doi:10.2307/3580087

Cited by 18 publications

(20 citation statements)

References 44 publications

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“…When the chambers were opened, 100 M deferoximine was added to the ice-cold medium and included in all subsequent steps. The cells were rinsed and scraped into versene, centrifuged, resuspended in 20 l of phosphate-buffered saline, and lysed in hypotonic buffer with CHAPS, phenylmethylsulfonyl fluoride, and Sigma protease inhibitor (27). For whole cell extracts, urea was added followed by a high speed spin, and the supernatant was frozen.…”

Section: Methodsmentioning

confidence: 99%

Detection of Reactive Oxygen Species via Endogenous Oxidative Pentose Phosphate Cycle Activity in Response to Oxygen Concentration

Tuttle

Maity

Oprysko

et al. 2007

Journal of Biological Chemistry

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The oxidative pentose phosphate cycle (OPPC) is necessary to maintain cellular reducing capacity during periods of increased oxidative stress. Metabolic flux through the OPPC increases stoichiometrically in response to a broad range of chemical oxidants, including those that generate reactive oxygen species (ROS). Here we show that OPPC sensitivity is sufficient to detect low levels of ROS produced metabolically as a function of the percentage of O 2 . We observe a significant decrease in OPPC activity in cells incubated under severe and moderate hypoxia (ranging from <0.01 to 4% O 2 ), whereas hyperoxia (95% O 2 ) results in a significant increase in OPPC activity. These data indicate that metabolic ROS production is directly dependent on oxygen concentration. Moreover, we have found no evidence to suggest that ROS, produced by mitochondria, are needed to stabilize hypoxia-inducible factor 1␣ (HIF-1␣) under moderate hypoxia. Myxothiazol, an inhibitor of mitochondrial electron transfer, did not prevent HIF-1␣ stabilization under moderate hypoxia. Moreover, the levels of HIF-1␣ that we observed after exposure to moderate hypoxia were comparable between 0 cells, which lack functional mitochondria, and the wild-type cells. Finally, we find no evidence for stabilization of HIF-1␣ in response to the non-toxic levels of H 2 O 2 generated by the enzyme glucose oxidase. Therefore, we conclude that the oxygen dependence of the prolyl hydroxylase reaction is sufficient to mediate HIF-1␣ stability under moderate as well as severe hypoxia.The primary role of the oxidative pentose phosphate cycle (OPPC) 2 in mammalian cells is to maintain the [NADPH/ NADPϩ] ratio, thereby helping to regulate the cellular redox equilibrium (1-3). Glucose-6-phosphate dehydrogenase (G6PD), the initial and rate-limiting enzyme of the OPPC, exists in a dimer-tetramer equilibrium with the tetramer being the catalytically active conformation. Each of four identical G6PD monomers contains a structural NADPϩ binding site (4, 5). When NADPϩ is bound at this site, formation of the active tetramer is favored. In non-stressed cells, the [NADPH]/ [NADPϩ] ratio is very high (approaching 1000) and flux through the OPPC is minimal. However, even a slight increase in [NADPϩ] can increase the number of active G6PD tetramers. Therefore, G6PD activity, by regulating flux through the OPPC, is uniquely sensitive to reactive oxygen species (ROS) as well as other chemical oxidants.The importance of the OPPC for the cellular response to ROS is evident from the elevated incidence of apoptosis that we observed in G6PD Ϫ Chinese hamster ovary cells following exposure to ionizing radiation (2). Likewise, Efferth et al. (6) observed an elevated incidence of oxidant-induced apoptosis in macrophages isolated from patients suffering from G6PD deficiency syndrome, while Fico et al. (42) observed H 2 O 2 -induced apoptosis in G6PD Ϫ mouse embryo fibroblasts. Notably, a 10-fold reduction in cloning efficiency was seen in G6PD Ϫ mouse embryo fibroblasts (MEFs) incubated in a...

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Section: Methodsmentioning

confidence: 99%

Detection of Reactive Oxygen Species via Endogenous Oxidative Pentose Phosphate Cycle Activity in Response to Oxygen Concentration

Tuttle

Maity

Oprysko

et al. 2007

Journal of Biological Chemistry

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“…[1][2][3] For this reason, radiation physics and chemistry studies have attempted to link biological damage to fundamental processes including the production of electrons, ions, electronic excited states [4][5][6] and subsequent reactions. 1,[6][7][8][9][10][11] Many studies involved the reactions of radiation products in aqueous solutions with the DNA molecules.…”

Section: Introductionmentioning

confidence: 99%

Measurements ofGValues for DNA Damage Induced by Low-Energy Electrons

Alizadeh

Sanche

2011

J. Phys. Chem. B

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We address the problem of measurement of G-values (damage per deposited energy) for low energy electrons (LEEs) below 30 eV. Such values (G LEE ) usually have to be derived from damage yields in nanometer (~10nm) thick films, which are too thin to allow complete absorption of the energy of LEEs. In this work, we determine optimum corrections to obtain reliable G LEE in 2-80 nm thick films of plasmid DNA which are not uniform. G LEE is found to increase with film average thickness and reaches a plateau at 260±50 nmol/J around 20 nm, which corresponds to the most reliable value. The previously G LEE measured for films thinner than 20nm that were underestimated can be corrected from a factor derived from the present results. This method could be used to obtain reliable G LEE for other biomolecules so as to be able to compare LEE-induced damage to that produced by other types of radiation under various experimental conditions.

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“…These toxic effects of O 2 exposure [14][15][16] were interpreted as due to the reaction of O 2 with damage sites within DNA, a phenomenon called the "Oxygen-Effect". 13,17 More specifically, when plasmid DNA is surrounded by oxygen at atmospheric pressure, the damage to DNA induced by LEEs is found to double. 18 Thus, O 2 considerably sensitises DNA to LEE derived damage.…”

Section: Introductionmentioning

confidence: 99%

Low energy electron stimulated desorption from DNA films dosed with oxygen

Mirsaleh-Kohan

Bass

Cloutier

et al. 2012

The Journal of Chemical Physics

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Desorption of anions stimulated by 1-18 eV electron impact on self-assembled monolayer (SAM) films of single DNA strands is measured as a function of film temperature (50-250 K). The SAMs, composed of 10 nucleotides, are dosed with O 2 . The OH − desorption yields increase markedly with exposure to O 2 at 50 K and are further enhanced upon heating. In contrast, the desorption yields of O − , attributable to dissociative electron attachment to trapped O 2 molecules decrease with heating. Irradiation of the DNA films prior to the deposition of O 2 shows that this surprising increase in OH − desorption, at elevated temperatures, arises from the reaction of O 2 with damaged DNA sites. These results thus appear to be a manifestation of the so-called "oxygen fixation" effect, well known in radiobiology.

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Effect of Oxygen on Radiation-Induced DNA Damage in Isolated Nuclei

Cited by 18 publications

References 44 publications

Detection of Reactive Oxygen Species via Endogenous Oxidative Pentose Phosphate Cycle Activity in Response to Oxygen Concentration

Detection of Reactive Oxygen Species via Endogenous Oxidative Pentose Phosphate Cycle Activity in Response to Oxygen Concentration

Measurements ofGValues for DNA Damage Induced by Low-Energy Electrons

Low energy electron stimulated desorption from DNA films dosed with oxygen

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