Monte Carlo calculation of the primary radical and molecular yields of liquid water radiolysis in the linear energy transfer range 0.3-6.5 keV/micrometer: application to 137Cs gamma rays

Monte Carlo simulations of the radiolysis of neutral liquid water and 0.4 M H(2)SO(4) aqueous solutions at ambient temperature are used to calculate the variations of the primary radical and molecular yields (at 10(-6)s) as a function of linear energy transfer (LET) in the range approximately 0.3 to 6.5 keV/micrometer. The early energy deposition is approximated by considering short (approximately 20-100 micrometer) high-energy (approximately 300-6.6 MeV) proton track segments, over which the LET remains essentially constant. The subsequent nonhomogeneous chemical evolution of the reactive species formed in these tracks is simulated by using the independent reaction times approximation, which has previously been used successfully to model the radiolysis of water under various conditions. The results obtained are in good general agreement with available experimental data over the whole LET range studied. After normalization of our computed yields relative to the standard radical and molecular yields for (60)Co gamma radiation (average LET approximately 0.3 keV/micrometer), we obtain empirical relationships of the primary radiolytic yields as a function of LET over the LET range studied. Such relationships are of practical interest since they allow us to predict a priori values of the radical and molecular yields for any radiation from the knowledge of the average LET of this radiation only. As an application, we determine the corresponding yields for the case of (137)Cs gamma radiation. For this purpose, we use the value of approximately 0.91 keV/micrometer for the average LET of (137)Cs gamma rays, chosen so that our calculated yield G(Fe(3+)) for ferrous-ion oxidation in air-saturated 0.4 M sulfuric acid reproduces the value of 15.3 molecules/100 eV for this radiation recommended by the International Commission on Radiation Units and Measurements. The uncertainty range on those primary radical and molecular yields are also determined knowing the experimental error (approximately 2%) for the measured G(Fe(3+)) value. The following values (expressed in molecules/100 eV) are obtained: (1) for neutral water: G(e(-)(aq)) = 2.50 +/- 0.16, G(H(.)) = 0.621 +/- 0.019, G(H(2)) = 0.474 +/- 0.025, G((.)OH) = 2.67 +/- 0.14, G(H(2)O(2)) = 0.713 +/- 0.031, and G(-H(2)O) = 4.08 +/- 0.22; and (2) for 0.4 M H(2)SO(4) aqueous solutions: G(H(.)) = 3.61 +/- 0.09, G(H(2)) = 0.420 +/- 0.019, G((.)OH) = 2.78 +/- 0.12, G(H(2)O(2)) = 0.839 +/- 0.037, and G(-H(2)O) = 4.46 +/- 0.16. These computed values are found to differ from the standard yields for (60)Co gamma rays by up to approximately 6%.     

High-LET ion radiolysis of water: visualization of the formation and evolution of ion tracks and relevance to the radiation-induced bystander effect

Ionizing radiation-induced bystander effects, commonly observed in cell populations exposed to high-linear energy transfer (LET) radiations, are initiated by damage to a cellular molecule which then gives rise to a toxic signal exported to neighboring cells not directly hit by radiation. A major goal in studies of this phenomenon is the identification of this initial radiation-induced lesion. Liquid water being the main constituent of biological matter, reactive species produced by water radiolysis in the cellular environment are likely to be major contributors to the induction of this lesion. In this context, the radiation track structure is of crucial importance in specifying the precise location and identity of all the radiolytic species and their subsequent signaling or damaging effects. We report here Monte Carlo track structure simulations of the radiolysis of liquid water by four different impacting ions 1H+, 4He2+, 12C6+ and 20Ne10+, with the same LET ( approximately 70 keV/ microm). The initial radial distribution profiles of the various water decomposition products (eaq(-), *OH, H*, H2 and H2O2) for the different ions considered are presented and discussed briefly in the context of track structure theory. As an example, the formation and temporal evolution of simulated 24 MeV 4He2+ ion tracks (LET approximately 26 keV/microm) are reported for each radiolytic species from 1 ps to 10 micros. The calculations reveal that the ion track structure is completely lost by approximately 1 micros.     

Production of superoxide radicals with pulse radiolysis of water with high linear energy transfer

The radiolysis of water with heavy ions of high linear energy transfer (LET) (-dE/dx) is characterized, in deaerated medium, by the production of superoxide anions, the radiolytic yields of which increase with the LET. Radiobiological interest in such radical species comes from the oxidative stress which may be generated by their dismutation in O2 and H2O2 in anoxic medium (radiotherapy with heavy ions). A brief review of the measurements of superoxide free radicals in aqueous solution by indirect or direct methods is presented. Moreover, some experimental results obtained by pulse radiolysis with Ar18+ ions (TEL = 290 keV x microm(-1)), are described. The interpretation of the kinetics takes into account the superoxide absorbance and that of hydrogen peroxide, which is present at the millisecond time scale.     

High LET heavy ion radiation induces lower numbers of initial chromosome breaks with minimal repair than low LET radiation in normal human cells

We investigated the earliest possible chromosome break and repair process in normal human fibroblasts irradiated with low and high LET (linear energy transfer) heavy ion radiation using the modified premature chromosome condensation (PCC) technique utilizing wortmannin (WM) during the fusion incubation period [M. Okada, S. Saito, R. Okayasu, Facilitated detection of chromosome break and repair at low levels of ionizing radiation by addition of wortmannin to G1-type PCC fusion incubation, Mutat. Res., 562 (2004) 11-17]. The initial numbers of breaks were approximately 10/cell/Gy in X-irradiated samples, followed by carbon (LET: 70 keV/microm), neon, and the number was around 5/cell/Gy in silicon (LET: 70 and 200 keV/microm) and iron (LET: 200 keV/microm) samples. If WM was not used, the initial numbers of breaks with silicon and iron were higher than those of X-rays. To quantify these data, we used initial repair ratio (IRR) defined as the number of G1 PCC breaks with WM divided by the number of breaks without WM. X-irradiation gave the maximum IRR ( approximately 2.0), while iron as well as silicon irradiation showed the minimum IRR ( approximately 1.0), suggesting almost no rejoining at the initial stage. Although there is a comparatively good correlation between the IRR value and the cell survival, the survival fraction with the repair data at 2 or 6h correlates better statistically. Our data indicate that high LET heavy ion irradiation induces a lower number of initial chromosome breaks with minimal repair when compared with low LET irradiation. These results at the chromosome level substantiate and extend the notion that high LET radiation produces complex-type DNA double strand breaks (DSBs).     

Track structure in DNA irradiated with heavy ions

The spatial properties of trapped radicals produced in heavy-ion-irradiated solid DNA at 77 K have been probed using pulsed electron paramagnetic double resonance (PELDOR or DEER) techniques. Salmon testes DNA hydrated to 12 water molecules per nucleotide was irradiated with 40Ar ions of energy 100 MeV/nucleon and LET ranging from 300 to 400 keV/microm. Irradiated samples were maintained at cryogenic temperature at all times. PELDOR measurements were made using a refocused echo detection sequence that allows dipolar interaction between trapped radicals to be observed. The EPR spectrum is attributed to electron loss/gain DNA base radicals and neutral carbon-centered radicals that likely arise from sugar damage. We find a radical concentration of 13.5 x 10(18) cm(-3) in the tracks and a track radius of 6.79 nm. The cross section of these tracks is 144 nm2, yielding a lineal radical density of 2.6 radicals/nm. Based on the yields determined previously for particles having calculated LET values of 300-400 keV/microm and our measured lineal density, we obtain an LET of 270 keV/microm, which is in good agreement with the calculated range of values. These measurements of radical density and spatial extent provide the first direct experimental determination of track characteristics in irradiated DNA.     

On (3)H beta-particle and (60)Co gamma irradiation of aqueous systems

The chemistry of water and aqueous solutions is very different after irradiation with (3)H beta particles and high-energy electrons or (60)Co gamma rays. The greater the linear energy transfer (LET) of the medium for (3)H beta particles compared to high-energy electrons or (60)Co gamma rays leads to an increased local concentration of reactants. There is an increased amount of intratrack chemistry, which reduces the escape yield of and OH by about 50%, but increases the yield of H(2) by about 50% and of H(2)O(2) by about 35%. Analysis of stochastic-diffusion kinetic calculations employing simulated track structures reveals that the yield of H(2) produced by diffusion-kinetic processes increases significantly for (3)H beta particles compared to (60)Co gamma radiation, while production of H(2) by sub-picosecond processes is essentially the same. In both (3)H beta-particle and (60)Co gamma radiolysis, the reactions + and are equally important in the production of H(2). In the former case, each reaction has a yield of approximately 0.18, and in the latter a yield of approximately 0.08. In neutral water, the reaction (H + H) is negligible. The yield of Fe(III) in (3)H beta-particle radiolysis of the Fricke dosimeter is much smaller than in radiolysis with more energetic electrons. Simulations show that this change is primarily due to the reduced escape yield of H, formed from the scavenging of by the bulk H(3)O(+) of the acid. The chemical differences observed in experiments, and in calculations, reflect the underlying structure of the electron tracks: Examination of the track structure simulations demonstrates that primary events are considerably more well-separated in high-energy electron tracks compared to (3)H beta-particle tracks.     

Influence of radiation quality on the yield of DNA strand breaks in SV40 DNA irradiated in solution.

Using highly energetic particles to irradiate plasmid DNA in aerobic aqueous solution, we have compiled an extensive database on how yields of DNA single- and double-strand breaks (SSBs and DSBs) vary with radiation quality. This study was performed in a low-scavenging buffer system and covers a wide range of ion species (helium to uranium) and LETs (5 to 16,000 keV/microm). For LETs up to around 40 keV/microm for SSBs and 400 keV/microm for DSBs, the total energy deposition determines cross section. At higher LET, cross sections level off and individual plateaus for particles of different atomic numbers are observed. For each ion species this is more pronounced and occurs at lower LET for SSBs than for DSBs, leading to an increase in the DSB:SSB ratio from 1:70 for X rays to 1:6 at 500 keV/microm. At this LET, the influence of track structure becomes evident, with high local concentrations of ionization events favoring the formation of DSBs and also intratrack recombination reactions. For lower-energy ions, a saturation in production of measurable DSBs is apparent, due to correlated lesion induction within densely ionizing particle tracks. For very heavy low-energy ions, both SSB and DSB cross sections decrease with particle velocity at nearly constant LET, forming individual hooked curves when plotted as a function of LET.     

Simulation of DNA damage after proton irradiation

The biophysical radiation track simulation model PARTRAC was improved by implementing new interaction cross sections for protons in water. Computer-simulated tracks of energy deposition events from protons and their secondary electrons were superimposed on a higher-order DNA target model describing the spatial coordinates of the whole genome inside a human cell. Induction of DNA double-strand breaks was simulated for proton irradiation with LET values between 1.6 and 70 keV/microm and various reference radiation qualities. The yield of DSBs after proton irradiation was found to rise continuously with increasing LET up to about 20 DSBs per Gbp and Gy, corresponding to an RBE up to 2.2. About half of this increase resulted from a higher yield of DSB clusters associated with small fragments below 10 kbp. Exclusion of experimentally unresolved multiple DSBs reduced the maximum DSB yield by 30% and shifted it to an LET of about 40 keV/microm. Simulated fragment size distributions deviated significantly from random breakage distributions over the whole size range after irradiation with protons with an LET above 10 keV/microm. Determination of DSB yields using equations derived for random breakage resulted in an underestimation by up to 20%. The inclusion of background fragments had only a minor influence on the distribution of the DNA fragments induced by radiation. Despite limited numerical agreement, the simulations reproduced the trends in proton-induced DNA DSBs and fragment induction found in recent experiments.     

Biological effectiveness of accelerated particles for the induction of chromosome damage measured in metaphase and interphase human lymphocytes

Chromosome aberrations were investigated in human lymphocytes after in vitro exposure to 1H-, 3He-, 12C-, 40Ar-, 28Si-, 56Fe-, or 197Au-ion beams, with LET ranging from approximately 0.4-1393 keV/microm in the dose range of 0.075-3 Gy. Dose-response curves for chromosome exchanges, measured at the first mitosis postirradiation using fluorescence in situ hybridization (FISH) with whole-chromosome probes, were fitted with linear or linear-quadratic functions. The relative biological effectiveness (RBE) was estimated from the initial slope of the dose-response curve for chromosomal damage with respect to low- or high-dose-rate gamma rays. Estimates of RBEmax values for mitotic spreads, which ranged from near 0.7 to 11.1 for total exchanges, increased with LET, reaching a maximum at about 150 keV/microm, and decreased with further increase in LET. RBEs for complex aberrations are undefined due to the lack of an initial slope for gamma rays. Additionally, the effect of mitotic delay on RBE values was investigated by measuring chromosome aberrations in interphase after chemically induced premature chromosome condensation (PCC), and values were up to threefold higher than for metaphase analysis.     

Combined use of Monte Carlo DNA damage simulations and deterministic repair models to examine putative mechanisms of cell killing

A kinetic repair-misrepair-fixation (RMF) model is developed to better link double-strand break (DSB) induction to reproductive cell death. Formulas linking linear-quadratic (LQ) model radiosensitivity parameters to DSB induction and repair explicitly account for the contribution to cell killing of unrejoinable DSBs, misrepaired and fixed DSBs, and exchanges formed through intra- and intertrack DSB interactions. Information from Monte Carlo simulations is used to determine the initial yields and complexity of DSBs formed by low- and high-LET radiations. Our analysis of published survival data for human kidney cells suggests that intratrack DSB interactions are negligible for low-LET radiations but increase rapidly with increasing LET. The analysis suggests that no class of DSB is intrinsically unrejoinable or that DSB reparability is not strictly determined by the number of lesions forming the DSB. For radiations with LET >110 keV/mum, the model predicts that the relative cell killing efficiency, per unit absorbed dose, should continue to increase, whereas data from published experiments indicate a reduced cell killing efficiency. This observation suggests that the Monte Carlo simulation overestimates the DSB yield beyond 110 keV/microm or that other biological phenomena not included in the model, such as proximity effects, are important. For 200-250 kVp X rays ( approximately 1.9 keV/microm), only about 1% of the one-track killing is attributed to intratrack binary misrepair interactions. The analysis indicates that the remaining 99% of the lethal damage is due to other types of one-track damage, including possible unrepairable, misrepaired and fixed damage. Compared to the analysis of the X-ray results, 48% of the one-track lethal damage caused by 5.1 MeV alpha particles (approximately 88 keV/microm) is due to intratrack DSB interactions while the remainder is due to other forms of one-track damage.     

Quantifying rates of protein synthesis in humans by use of 2H2O: application to patients with end-stage renal disease

A method is introduced for quantitating protein synthetic rates in humans by use of (2)H(2)O. Its validity was tested in subjects with end-stage renal disease. Six clinically stable subjects, hemodialyzed three times weekly, ingested (2)H(2)O to a body water (2)H enrichment of approximately 0.4%. On dialysis, body water enrichment declined to approximately 0.1%. Enrichment of the alpha-hydrogen of plasma free alanine was also approximately 0.4% before and approximately 0.1% after dialysis. Beta-hydrogen enrichment was approximately 80-100% of alpha-hydrogen enrichment. (2)H(2)O was ingested to replace (2)H(2)O removed after each dialysis for 15-51 days, returning enrichment to approximately 0.4%. Enrichment of alanine from plasma albumin gradually increased, with again approximately 80-100% as much (2)H in beta- as in alpha-hydrogens. With continued dialyses, without (2)H(2)O replacement, alanine from albumin enrichment gradually declined, whereas free alanine and water enrichments were negligible. The fractional albumin synthesis rate, calculated from the increase in enrichment in alanine from albumin, was 4.0 +/- 0.5%/day, and from the decrease, 4.6 +/- 0.2%/day. Thus body water enrichment in a subject given (2)H(2)O can be maintained constant long term. A rapid exchange, essentially complete, occurs between the hydrogens of alanine and body water. An integrated measure over a long period of albumin's synthetic rate can be estimated from both the rise in enrichment of alanine from the protein during (2)H(2)O ingestion and fall on (2)H(2)O withdrawal, while the subject's living routine is uninterrupted. Estimates are in subjects with renal disease, but the method should be applicable to estimates of protein synthetic rates in normal subjects and in other pathological states.     

Respiration-dependent removal of exogenous H2O2 in brain mitochondria: inhibition by Ca2+

In brain mitochondria, state 4 respiration supported by the NAD-linked substrates glutamate/malate in the presence of EGTA promotes a high rate of exogenous H2O2 removal. Omitting EGTA decreases the H2O2 removal rate by almost 80%. The decrease depends on the influx of contaminating Ca2+, being prevented by the Ca2+ uniporter inhibitor ruthenium red. Arsenite is also an inhibitor (maximal effect approximately 40%, IC50, 12 microm). The H2O2 removal rate (EGTA present) is decreased by 20% during state 3 respiration and by 60-70% in fully uncoupled conditions. H2O2 removal in mitochondria is largely dependent on glutathione peroxidase and glutathione reductase. Both enzyme activities, as studied in disrupted mitochondria, are inhibited by Ca2+. Glutathione reductase is decreased by 70% with an IC50 of about 0.9 microm, and glutathione peroxidase is decreased by 38% with a similar IC50. The highest Ca2+ effect with glutathione reductase is observed in the presence of low concentrations of H2O2. With succinate as substrate, the removal is 50% less than with glutamate/malate. This appears to depend on succinate-supported production of H2O2 by reverse electron flow at NADH dehydrogenase competing with exogenous H2O2 for removal. Succinate-dependent H2O2 is inhibited by rotenone, decreased DeltaPsi, as described previously, and by ruthenium red and glutamate/malate. These agents also increase the measured rate of exogenous H2O2 removal with succinate. Succinate-dependent H2O2 generation is also inhibited by contaminating Ca2+. Therefore, Ca2+ acts as an inhibitor of both H2O2 removal and the succinate-supported H2O2 production. It is concluded that mitochondria function as intracellular Ca2+-modulated peroxide sinks.     

Effect of linear energy transfer (LET) on the complexity of alpha-particle-induced chromosome aberrations in human CD34+ cells

The aim of this study was to assess the relative influence of the linear energy transfer (LET) of alpha particles on the complexity of chromosome aberrations in the absence of significant other differences in track structure. To do this, we irradiated human hemopoietic stem cells (CD34+) with alpha particles of various incident LETs (110-152 keV/microm, with mean LETs through the cell of 119-182 keV/microm) at an equi-fluence of approximately one particle/cell and assayed for chromosome aberrations by mFISH. Based on a single harvest time to collect early-division mitotic cells, complex aberrations were observed at comparable frequencies irrespective of incident LET; however, when expressed as a proportion of the total exchanges detected, their occurrence was seen to increase with increasing LET. Cycle analysis to predict theoretical DNA double-strand break rejoining cycles was also carried out on all complex chromosome aberrations detected. By doing this we found that the majority of complex aberrations are formed in single non-reducible cycles that involve just two or three different chromosomes and three or four different breaks. Each non-reducible cycle is suggested to represent "an area" of finite size within the nucleus where double-strand break repair occurs. We suggest that the local density of damage induced and the proximity of independent repair areas within the interphase nucleus determine the complexity of aberrations resolved in metaphase. Overall, the most likely outcome of a single nuclear traversal of a single alpha particle in CD34+ cells is a single chromosome aberration per damaged cell. As the incident LET of the alpha particle increases, the likelihood of this aberration being classed as complex is greater.     

Does the limiting F value at very low doses depend systematically on linear energy transfer?

The debate on the validity of the ratios of radiation-induced yields of chromosome aberrations, in particular the F value (dicentrics/ring chromosomes), as a chromosomal fingerprint of radiation quality is still in progress. From a recent analysis of their experimental data, Sasaki et al. (Radiat. Res. 150, 253-258, 1998) noted that despite a considerable variability in the data, the limiting F value at the lowest doses, or the F(0) value, obviously decreased with increasing LET, indicating that the LET could be a factor that determines the F value. We have reassessed here our own 13 cytogenetic data sets that cover a range of dose-averaged LET of 0.5 to 150 keV/microm in terms of this F(0)-value approach, but we could not confirm such a dependence on LET at very low doses. The validity of the F value as a biomarker therefore remains questionable. For a final evaluation, scoring of a far greater number of cells at low doses would be necessary to reduce the large error ranges of F values.     

Radioprotection by DMSO of mammalian cells exposed to X-rays and to heavy charged-particle beams

Populations of G1-phase Chinese hamster cells in stirred suspensions containing various concentrations of DMSO were irradiated with 250 kV X-rays or various heavy charged-particle beams. Chemical radioprotection of cell inactivation was observed for all LET values studied. When cell survival data were resolved into linear and quadratic components, the extent and concentration dependence of DMSO protection were found to be different for the two mechanisms. The chemical kinetics of radioprotection for single-events were similar for LET values up to those which gave the maximum RBE. DMSO protected to a lesser extent against energetic argon ions at an median LET of approximately 220 keV/micron. These data could indicate the contribution of indirect action by hydroxyl radicals and hydrogen atoms to cell inactivation by single-hit and double-hit mechanisms for various radiation qualities. The decrease in RBE observed at very high LET may result, in part, from reduced yields of water radicals at 10(-9)-10(-8) s resulting from radical recombination mechanisms within the charged particle tracks.     

Lipid interaction of alpha-synuclein during the metal-catalyzed oxidation in the presence of Cu2+ and H2O2

Alpha-synuclein co-exists with lipids in the Lewy bodies, a pathological hallmark of Parkinson's disease. Molecular interaction between alpha-synuclein and lipids has been examined by observing lipid-induced protein self-oligomerization in the presence of a chemical coupling reagent of N-(ethoxycarbonyl)-2-ethoxy-1,2-dihydroquinoline. Lipids such as phosphatidic acid, phosphatidylinositol, phosphatidylserine, phosphatidylethanolamine, and even arachidonic acid induced the self-oligomerization whereas phosphatidylcholine did not affect the protein. Because the oligomerizations occurred from critical micelle concentrations of the lipids, the self interaction of alpha-synuclein was shown to be a lipid-surface dependent phenomenon with head group specificity. By employing beta-synuclein and a C-terminally truncated alpha-synuclein (alpha-syn97), the head-group dependent self-oligomerization was demonstrated to occur preferentially at the N-terminal region while the fatty acid interaction leading to the protein self-association required the presence of the acidic C-terminus of alpha-synuclein. In the presence of Cu2+ and H2O2, phosphatidylinositol (PI), along with other acidic lipids, actually enhanced the metal-catalyzed oxidative self-oligomerization of alpha-synuclein. The dityrosine crosslink formation responsible for the PI-enhanced covalent self-oligomerization was more sensitive to variation of copper concentrations than that of H2O2 during the metal-catalyzed oxidation. The enhancement by PI was shown to be due to facilitation of copper localization to the protein because actual binding affinity between copper and alpha-synuclein increased from Kd of 44.7 microm to 5.9 microm in the presence of the lipid. Taken together, PI not only affects alpha-synuclein to be more self-interactive by providing the lipid surface, but also enhances the metal-catalyzed oxidative protein self-oligomerization by facilitating copper localization to the protein when the metal and H2O2 are provided. This observation therefore could be implicated in the formation of Lewy bodies as lipids and metal-catalyzed oxidative stress have been considered to be a part of pathological causes leading to the neurodegeneration.     

Heavy ion-induced DNA double-strand breaks in yeast

Induction of DSBs in the diploid yeast, Saccharomyces cerevisiae, was measured by pulsed-field gel electrophoresis (PFGE) after the cells had been exposed on membrane filters to a variety of energetic heavy ions with values of linear energy transfer (LET) ranging from about 2 to 11,500 keV/microm, (241)Am alpha particles, and 80 keV X rays. After irradiation, the cells were lysed, and the chromosomes were separated by PFGE. The gels were stained with ethidium bromide, placed on a UV transilluminator, and analyzed using a computer-coupled camera. The fluorescence intensities of the larger bands were found to decrease exponentially with dose or particle fluence. The slope of this line corresponds to the cross section for at least one double-strand break (DSB), but closely spaced multiple breaks cannot be discriminated. Based on the known size of the native DNA molecules, breakage cross sections per base pair were calculated. They increased with LET until they reached a transient plateau value of about 6 x 10(-7) microm(2) at about 300-2000 keV/microm; they then rose for the higher LETs, probably reflecting the influence of delta electrons. The relative biological effectiveness for DNA breakage displays a maximum of about 2.5 around 100-200 keV/microm and falls below unity for LET values above 10(3) keV/microm. For these yeast cells, comparison of the derived breakage cross sections with the corresponding cross section for inactivation derived from the terminal slope of the survival curves shows a strong linear relationship between these cross sections, extending over several orders of magnitude.     

Induction of DNA double-strand breaks by 1H and 4He lons in primary human skin fibroblasts in the LET range of 8 to 124 keV/microm.

Yields of DNA double-strand breaks were determined in primary human skin fibroblasts exposed to 1H and 4He ions at various linear energy transfers (LETs) and to 15 MeV electrons as the reference radiation. The values obtained for the relative biological effectiveness (RBE) were 2.03, 1.45 and 1.36 for 1H ions at LETs of 35, 23 and 7.9 keV/microm, respectively, and 1.2, 1.18, 1.38 and 1.31 for 4He ions at LETs of 124, 76, 35 and 27 keV/microm, respectively. The data were obtained using pulsed-field gel electrophoresis of DNA released from cells using the chromosomes of the yeast Saccharomyces cerevisiae as length markers and fitting the experimental mass distributions of fragmented DNA to those obtained by computer simulation of the random breakage of human chromosomes. The RBE values for induction of DSBs in mammalian cells cannot be fitted to a common RBE-LET relationship for electrons and 1H, 4He and light ions. Comparison of the RBEs for mammalian cells with the corresponding RBEs obtained for yeast cells shows similar RBEs of electrons for yeast and mammalian cells; however, for 4He and light ions in the LET range of 100 to 1000 keV/microm, the RBEs for yeast are significantly higher compared with mammalian cells. These characteristics of the RBE-LET relationships for yeast and mammalian cells are attributed to the fraction of small DNA fragments induced by particles when traversing the higher-order chromatin structures which are different to some extent in these two cell types.     

Contribution of indirect action to radiation-induced mammalian cell inactivation: dependence on photon energy and heavy-ion LET

The contribution of indirect action mediated by OH radicals to cell inactivation by ionizing radiations was evaluated for photons over the energy range from 12.4 keV to 1.25 MeV and for heavy ions over the linear energy transfer (LET) range from 20 keV/microm to 440 keV/microm by applying competition kinetics analysis using the OH radical scavenger DMSO. The maximum level of protection provided by DMSO (the protectable fraction) decreased with decreasing photon energy down to 63% at 12.4 keV. For heavy ions, a protectable fraction of 65% was found for an LET of around 200 keV/microm; above that LET, the value stayed the same. The reaction rate of OH radicals with intracellular molecules responsible for cell inactivation was nearly constant for photon inactivation, while for the heavy ions, the rate increased with increasing LET, suggesting a reaction with the densely produced OH radicals by high-LET ions. Using the protectable fraction, the cell killing was separated into two components, one due to indirect action and the other due to direct action. The inactivation efficiency for indirect action was greater than that for direct action over the photon energy range and the ion LET range tested. A significant contribution of direct action was also found for the increased RBE in the low photon energy region.     

Increased H2O2, vascular endothelial growth factor and receptors in the retina of the BBZ/Wor diabetic rat

Hyperglycemia in diabetes induces increased levels of hydrogen peroxide (H2O2), a reactive oxygen species generated by reduced nicotinamide adenine dinucleotide (NADH) oxidase. Nontoxic levels of H2O2 increase endothelial cell permeability. Using a model of non-insulin-dependent diabetes, the BBZ/Wor rat, we investigated retinal levels of H2O2, vascular endothelial growth factor (VEGF) and its receptors, VEGF-R1 and VEGF-R2 by transmission electron microscopy at sites of the blood-retinal barrier (BRB). H2O2 localization was done by the cerium NADH oxidase method, and extravasation of endogenous serum albumin was used to document disruption of the BRB. Higher levels of H2O2 were detected in blood vessels of diabetic (78.7 +/- 4.84%) as compared with vessels from nondiabetic rats (39.0 +/- 4.47%). VEGF immunoreactivity was statistically higher in the inner BRB (24.67 +/- 0.33 colloidal gold particles/63 microm2 vs. 21.52 +/- 0.43 colloidal gold particles/63 microm2, p = .0001) and outer BRB (42.56 +/- 0.45 colloidal gold particles/63 microm2 vs. 15.51 +/- 0.51 colloidal gold particles/63 microm2, p = .0001) of diabetic rats as compared with age matched nondiabetic control rats. VEGF-R1 immunoreactivity was significantly higher in diabetic retinas in both the inner BRB (21.66 +/- 0.75 colloidal gold particles/63 microm2 vs. 12.69 +/- 0.61 colloidal gold particles/63 microm2, p = .0001) and outer BRB (22.76 +/- 2.36 colloidal gold particles/63 microm2 vs. 8.53 +/- 2.67 colloidal gold particles/63 microm2, p = .0013). VEGF-R2 was statistically higher in the inner BRB (8.97 +/- 0.57 colloidal gold particles/63 microm2 versus 7.03 +/- 0.65 colloidal gold particles/63 microm2, p = .0419) but not in the outer BRB (29.42 +/- 1.25 colloidal gold particles/63 microm2 vs. 28.07 +/- 1.42 colloidal gold particles/63 microm2, p = .4889). H2O2 levels correlated with increased VEGF (correlation coefficient = 0.82, p = .001) in this model of nonproliferative diabetic retinopathy. These results support that hyperglycemia is one factor that induces retinal endothelial cells in vivo to increase H2O2 via NADH oxidase and stimulates increases in VEGF resulting in disruption of the BRB.