Cytogenetic effect of the pulse reactor BARS-6 continuous or one-time irradiation with ultrahigh dose rate on human lymphocytes

The results of the comparative study of radiation effects of the pulse reactor BARS-6 either in single pulse or continuos irradiation mode on human G0 lymphocytes are presented. Under identical doses the cytogenetic efficiency was observed to be higher for continuous irradiation (1 hour) than for single pulse irradiation with ultrahigh dose rate (0.5-3) x 106 Gy/minutes (pulse duration 65 x 10(-6) s). The difference averaged about 37% on total aberration frequency and 27% on the sum of dicentrics and centric rings. The influence of the dose rate and of the mixed gamma-neutron irradiation on the obtained results is discussed.     

Effect of temperature on the development of chromosome aberrations in human blood lymphocytes irradiated in pulse and continuous modes at BARS-6 reactor

Summarized results of 5 repeated experiments of comparative study of radiation effects of the pulse reactor BARS-6 either in a single pulse or a continuos irradiation mode on human lymphocytes are presented. Higher efficiency (30-40%) of continuous irradiation (exposure duration 1 h) rather than pulse irradiation with ultra-high dose rates (1-2.5) x 10(6) Gy/min (pulse duration 65 micros) was confirmed. The efficiency ratio did not depend on the temperature, 20 degrees C or 0 degrees C, during the exposure. Cell repair system and chromatin conformation influence on the results obtained is discussed.     

Liposomal Delivery Systems for Encapsulation of Ferrous Sulfate: Preparation and Characterization

Liposomal delivery systems for water-soluble bioactives were prepared using the pro-liposome and the microfluidization technologies. Iron, an essential micronutrient as ferrous sulfate and ascorbic acid, as an antioxidant for iron were encapsulated in the liposomes. Liposomes prepared by the microfluidization technology using 6% (w/w) concentration of the lipid encapsulated with ferrous sulfate and ascorbic acid had particle size distributions around 150 to 200 nm, whereas liposomes from the pro-liposome technology resulted in particle sizes of about 5 μm. The encapsulation efficiency of ferrous sulfate was 58% for the liposomes prepared by the microfluidization using 6% (w/w) lipid and 7.5% of ferrous sulfate concentrations, and it was 11% for the liposomes from pro-liposome technology using 1.5% (w/v) lipid and 15% of ferrous-sulfate concentration. Both the liposomes exhibited similar levels of oxidative stability, demonstrating the feasibility of microfluidization-based liposomal delivery systems for large-scale food/nutraceutical applications.     

Pulse-periodic generation of supershort avalanche electron beams and X-ray emission

Pulse-periodic generation of supershort avalanche electron beams (SAEBs) and X-ray emission in nitrogen, as well as the transition from a single-pulse mode to a pulse-periodic mode with a high repetition frequency, was studied experimentally. It is shown that, in the pulse-periodic mode, the full width at halfmaximum of the SAEB is larger and the decrease rate of the gap voltage is lower than those in the single-pulse mode. It is found that, when the front duration of the voltage pulse at a nitrogen pressure of 90 Torr decreases from 2.5 to 0.3 ns, the X-ray exposure dose in the pulse-periodic mode increases by more than one order of magnitude and the number of SAEB electrons also increases. It is shown that, in the pulse-periodic mode of a diffuse discharge, gas heating in the discharge gap results in a severalfold increase in the SAEB amplitude (the number of electrons in the beam). At a generator voltage of 25 kV, nitrogen pressure of 90 Torr, and pulse repetition frequency of 3.5 kHz, a runaway electron beam was detected behind the anode foil.     

Liposomal delivery systems for encapsulation of ferrous sulfate: Preparation and characterization

Liposomal delivery systems for water-soluble bioactives were prepared using the pro-liposome and the microfluidization technologies. Iron, an essential micronutrient as ferrous sulfate and ascorbic acid, as an antioxidant for iron were encapsulated in the liposomes. Liposomes prepared by the microfluidization technology using 6% (w/w) concentration of the lipid encapsulated with ferrous sulfate and ascorbic acid had particle size distributions around 150 to 200 nm, whereas liposomes from the pro-liposome technology resulted in particle sizes of about 5 microm. The encapsulation efficiency of ferrous sulfate was 58% for the liposomes prepared by the microfluidization using 6% (w/w) lipid and 7.5% of ferrous sulfate concentrations, and it was 11% for the liposomes from pro-liposome technology using 1.5% (w/v) lipid and 15% of ferrous-sulfate concentration. Both the liposomes exhibited similar levels of oxidative stability, demonstrating the feasibility of microfluidization-based liposomal delivery systems for large-scale food/nutraceutical applications.     

Relation of lipid peroxidation to loss of cations trapped in liposomes

Lipid peroxidation and alterations in cation loss have been induced in liposomes by ferrous ion, ascorbic acid, reduced and oxidized glutathione, and gamma radiation. Modifications of these effects by tocopherol and 2,6-di-tert-butyl-4-methylphenol (BHT) were studied when these antioxidants were either incorporated in the membrane or were added to already formed liposomes prior to the addition of the chemical agent or to irradiation. Lipid peroxidation, as indicated by the thiobarbituric acid test for malonic dialdehyde, did not correlate with alterations in cation loss. The largest amounts of lipid peroxidation induced by ascorbic acid and glutathione were associated with decreased cation loss. Inhibition of Fe(2+)- and radiation-induced lipid peroxidation by antioxidants did not inhibit the associated increase in cation loss. Tocopherol was a more effective antioxidant than BHT when it was incorporated in the membrane, whereas BHT was more effective when it was added to the liposomes after formation.     

Action of mixed gamma-neutron radiation with various dose rates on the number of cells in thymus and chromosomes of mice bone marrow and human lymphocytes

The irradiation with mixed gamma-neutron radiation was carried out at the pulse nuclear reactor on fast neutrons BARS-6 in a regimen of one pulse (100 micros) and in a regimen of continuous irradiation during 60 minutes. Was shown, that the irradiation of mice with pulse radiation was 1.3-1.8 times more effective in the induction of the chromosome aberrations in bone marrow cells in comparison with the continuous regimen of irradiation. At the same time, other biological tests (yield of chromosome aberrations in human lymphocytes, decreasing the number of cells in thymus) demonstrated that pulsed and continuous regimens have almost equal biological effectiveness.     

Study of dose-rate effects of neutron radiation in a wild-type and a repair-deficient yeasts saccharomyces

We report here a comparative analysis of RBE for lethality of a single pulse (duration 65 micros) of fast neutron with ultra high dose rates (up to 6 x 10(6) Gy/s) and continuous neutron radiation (3.6 x 10(3) s) of the pulse reactor BARS-6. Three diploid strains, one haploid strain and three diploid repair-deficient strains (rad52-1/rad52-1; rad54/rad54; rad2/rad2) were used. The RBE values (D(0gamma)/1D(0n)) of a single pulse and continuous neutron irradiation were equal (1.7-1.8) with maximum RBE (4.1-3.1) in region of low doses (shoulder region). Haploid cells were found to be more (3 times) sensitive to both gamma-rays and neutrons than the wild type. There was no obvious decrease in the RBE of 1.9 in highly sensitive haploid cells as compared with highly resistant diploid cells. The repair-deficient strains (rad52-1/rad52-1; rad54/rad54) were more (up to 10 fold) sensitive to both neutrons and gamma-rays as compared with their parent line. The RBE values of 1.5-1.7 of neutrons for these mutants (independent by of the mode of irradiation) were found. The repair-deficient mutant rad2/rad2 had similar sensitivity as a wild type and a RBE value was 2.0. We have concluded that biological effectiveness of the neutrons of pulse reactor BARS-6 was independent of the dose-rate, differing up to 10(8) fold. The RBE didn't vary significantly with the capacity of cells to repair DNA damages.     

The effects of ionizing radiation on the pulmonary vasculature of intact rats and isolated pulmonary endothelium

We studied the effects of ionizing radiation on the morphology of the pulmonary circulation using an in vivo rat model and an in vitro pulmonary artery endothelial cell model. Gamma radiation was given as either an acute (30 Gy) or fractionated (5 X 6 Gy) dose to one hemithorax of rats. An acute 30-Gy dose delivered resulted in a 70% decrease in pulmonary arterial perfusion, using technetium-99m microaggregated albumin (99mTc-MAA), in the irradiated lung by 2-3 weeks after irradiation. Pulmonary microradiographs, using a barium sulfate perfusion method, obtained 2-3 weeks after irradiation demonstrated widespread loss of capillary filling and segmentation of the vessels. Histologic examination demonstrated intact capillaries, suggesting that the alterations in pulmonary perfusion were at the precapillary level. Similar abnormalities in lung perfusion and morphology were found after delivery of fractionated doses of radiation, but the onset of the changes was delayed, occurring 4-6 weeks postirradiation. Using cultured pulmonary endothelial cell monolayers, cell sloughing and retraction from the surface substrate were observed within 24 h after in vitro delivery of 30 Gy. Similar findings occurred in monolayers given fractionated doses (5 X 6 Gy) of radiation 2-3 days after the final dose. The in vivo animal and in vitro endothelial cell models offer a useful means of examining the morphologic alterations involved in radiation lung vascular damage.     

Laser impact on bacterial ATP: insights into the mechanism of laser-bacteria interactions

The mechanisms of laser action on bacteria are not adequately understood. Here, an attempt has been made to study the fluctuation in ATP (adenosine triphosphate) concentration following laser irradiation from a pulsed Nd:YAG laser on a marine biofilm-forming bacterium Pseudoalteromonas carrageenovora. A stationary phase bacterial suspension (density 10(7-8) ml-1) was exposed to pulsed laser irradiations at a fluence of 0.1 J cm-2 (pulse width 5 ns, repetition rate 10 Hz) for different durations, ranging from 2 s to 15 min. The total viable count (TVC) and ATP concentration of the irradiated samples were determined immediately after the laser irradiation. While the maximum reduction in the TVC observed with respect to the control was 59% immediately after 15 min irradiation, the ATP concentration showed a reduction of about 86% for the same duration. The ATP concentration showed an abrupt reduction from 3 min of laser irradiation and continued to reduce significantly with increasing duration of irradiation. Thus, 3 min irradiation at a fluence of 0.1 J cm-2 is considered as an approximate threshold for ATP production in this bacterium. As the decreased level of ATP production continued, bacterial mortality resulted. The reduction in ATP production could be due to damage caused by the laser irradiations on bacterial metabolic processes such as cellular respiration.     

Localization of lipid peroxidation products in liposomes after γ-irradiation

The distribution of lipid peroxidation products in liposomes after γ-irradiation at various doses was studied. Increases in thiobarbituric-acid-reactive substances, in the absorbance at 232 nm and in hydroperoxides were observed mainly in liposomal membranes after relatively low doses of irradiation, while carbonyl compounds were distributed both inside and outside the membranes. After higher doses of irradiation, however, the absorbance at 232 nm and the amount of hydroperoxides reached a maximal level in the membrane portion and then decreased when the decomposition products were released from the membranes. Under this condition, malondialdehyde and other carbonyl compounds were increased mainly in the medium of liposomal suspension. These results are discussed with reference to the lipid peroxidation process which is induced quantitatively by ionizing radiation.     

Influence of calcium on lipid mixing mediated by influenza hemagglutinin

We studied the influence of calcium on lipid mixing mediated by influenza hemagglutinin (HA). Lipid mixing between HA-expressing cells and liposomes containing disialoganglioside, influenza virus receptor, was studied at 37 degrees C and neutral pH after a low-pH pulse at 4 degrees C. With DSPC/cholesterol liposomes, calcium present after raising the temperature significantly promoted lipid mixing only when it was triggered by a short low-pH application. In case of DOPC/cholesterol liposomes, calcium promotion was observed regardless of the duration of the low-pH pulse. Calcium present during a short, but not long, low-pH application to HA-expressing cells with bound DSPC/cholesterol liposomes at 4 degrees C inhibited subsequent lipid mixing. We hypothesize that calcium influences lipid mixing because it binds to a vestigial esterase domain of hemagglutinin or causes expulsion of the fusion peptide from an electronegative cavity. We suggest that calcium promotes the transition from early and reversible conformation(s) of low pH-activated HA towards an irreversible conformation that underlies both HA-mediated lipid mixing and HA inactivation.     

Induction and repair of DNA double-strand breaks assessed by gamma-H2AX foci after irradiation with pulsed or continuous proton beams

In particle tumor therapy including beam scanning at accelerators, the dose per voxel is delivered within about 100 ms. In contrast, the new technology of laser plasma acceleration will produce ultimately shorter particle packages that deliver the dose within a nanosecond. Here, possible differences for relative biological effectiveness in creating DNA double-strand breaks in pulsed or continuous irradiation mode are studied. HeLa cells were irradiated with 1 or 5 Gy of 20-MeV protons at the Munich tandem accelerator, either at continuous mode (100 ms), or applying a single pulse of 1-ns duration. Cells were fixed 1 h after 1-Gy irradiation and 24 h after 5-Gy irradiation, respectively. A dose–effect curve based on five doses of X-rays was taken as reference. The total number of phosphorylated histone H2AX (gamma-H2AX) foci per cell was determined using a custom-made software macro for gamma-H2AX foci counting. For 1 h after 1-Gy 20-MeV proton exposures, values for the relative biological effectiveness (RBE) of 0.97 ± 0.19 for pulsed and 1.13 ± 0.21 for continuous irradiations were obtained in the first experiment 1.13 ± 0.09 and 1.16 ± 0.09 in the second experiment. After 5 Gy and 24 h, RBE values of 0.99 ± 0.29 and 0.91 ± 0.23 were calculated, respectively. Based on the gamma-H2AX foci numbers obtained, no significant differences in RBE between pulsed and continuous proton irradiation in HeLa cells were detected. These results are well in line with our data on micronucleus induction in HeLa cells.     

A Study of the Effect of Terahertz Electromagnetic Radiation on Microbial Cell Viability

Biophysics - Abstract—The effect of pulsed terahertz radiation at a wavelength of 66 μm, a pulse duration of 100 ns, and a pulse energy of 200 mJ on a suspension of microbial cells was...     

The effectiveness of 20 mev protons at nanosecond pulse lengths in producing chromosome aberrations in human-hamster hybrid cells

Laser accelerated radiotherapy is a potential cancer treatment with proton and carbon-ion beams that is currently under development. Ultra-fast high-energy laser pulses will accelerate ion beams that deliver their dose to a patient in a "pulsed mode" that is expected to differ from conventional irradiation by increasing the dose delivery rate to a tissue voxel by approximately 8 orders of magnitude. In two independently performed experiments at the ion microprobe SNAKE of the 14 MV Munich tandem accelerator, A(L) cells were exposed either to protons with 1-ns pulse durations or to protons applied over 150 ms in continuous irradiation mode. A slightly but consistently lower aberration yield was observed for the pulsed compared to the continuous mode of proton irradiation. This difference was not statistically significant when each aberration type was analyzed separately (P values between 0.61 and 0.85 in experiment I and P values between 0.32 and 0.64 in experiment II). However, excluding the total aberrations, which were not analyzed as independent radiation-induced effects, the mean ratio of the yields of dicentrics, centric rings and excess acentrics scored together showed (with 95% CI) a significant difference of 0.90 (0.81; 0.98) between the pulsed and the continuous irradiation modes. A similar tendency was also determined for the corresponding RBE values relative to 70 kV X rays. Since the different findings for the comparisons of individual chromosome aberration types and combined comparisons could be explained by different sample sizes with the consequence that the individual comparisons had less statistical power to identify a difference, it can be concluded that 20 MeV protons may be slightly less effective in the pulsed mode.     

The effect of electromagnetic waves of very high frequency of molecular spectra of radiation and absorption of nitric oxide on the functional activity of platelets

A study was made of the effect of electromagnetic EMI MMD-fluctuation on the frequencies of molecular spectra of radiation, and nitric oxide absorption under in vitro conditions on the functional activity of platelets in patients with unstable angina pectoris, with the help of a specially created generator. At amplitude-modulated and continuous modes of EMI MMD-irradiation of platelet-rich plasma for 5, 15 and 30 min the platelet functional activity decreases, which was shown up in reduction of their activation and fall of aggregative ability. The degree, to which platelet functional activity was inhibited, depended on the mode of irradiation and on duration of EMI MMD effect. The most obvious changes in platelet activation and in their readiness to aggregative response were observed at a continuous mode of irradiation within a 15 min interval.     

Effects of long-term storage on phytochrome-mediated germination in lettuce seeds

The effects of long-term seed storage on the physiological properties of phytochrome-mediated germination including water uptake, the temperature and light flunnce dependencies of germination and dark germination were studied. The fluenceresponse relationships of the brief irradiation with monochromatic red (660 nm, 7.5 W m⁻²) and far-red (750 nm, 6.6 W m⁻²) light at various times after sowing were also studied. The samples used consisted of three lots of seeds ofLactuca sativa L. cv. MSU-16, which had been harvested in 1976, 1979 and 1985 and stored dry for 9, 6 and 0 years, respectively, in darkness at 23±2 C until the experiments were carried out in July–August, 1985. Seeds with the longer storage periods showed the higher ability to germinate in both continuous darkness and continuous white fluorescent light at 20–30 C. In the seeds stored for 6 or 9 years, red light irradiation for 20 sec given at 15 min or more after sowing at 25 C induced as high a percent germination (85–95%) as those under continuous white fluorescent light. In the freshly harvested seeds, however, germination under continuous white fluorescent light (46%) was considerably lower than the germination induced by the red pulse (97%). Germination of the seeds decreased when the intervals between sowing and a far-red irradiation for 20 sec increased up to 100 min (or 30 min in the freshly harvested seeds). The far-red pulse given later than 100 min (or 6 hr in the freshly harvested seeds) after sowing resulted in an increased germination up to the dark-germination levels with increasing intervals between sowing and the pulse irradiation. Before or at 3 min after sowing, the seeds stored for 6 or 9 years were responsive to the far-red pulse although they were not or hardly responsive to the red pulse, while the freshly harvested seeds were responsive to both the far-red and the red pulses. These data indicate that normal functions of phytochrome completely survived in the dry seeds during storage at 25 C for as long as 6 or 9 years and that these functions are restored into full operation by means of imbibition. The differences in the dependence of germination on the time and fluence of a single pulse of red or far-red light seems to be related to the smaller water content throughout the imbibition in the seeds with the longer storage periods. The greater ability to germinate in the dark indicates the greater amounts of P_FR or the greater responsivity to P_FR, in the seeds with the longer storage periods.     

Laser Impact on Bacterial ATP: Insights into the Mechanism of Laser-Bacteria Interactions

The mechanisms of laser action on bacteria are not adequately understood. Here, an attempt has been made to study the fluctuation in ATP (adenosine triphosphate) concentration following laser irradiation from a pulsed Nd:YAG laser on a marine biofilm-forming bacterium Pseudoalteromonas carrageenovora. A stationary phase bacterial suspension (density 10^7-8 ml^{m 1}) was exposed to pulsed laser irradiations at a fluence of 0.1 J cm^{m 2} (pulse width 5 ns, repetition rate 10 Hz) for different durations, ranging from 2 s to 15 min. The total viable count (TVC) and ATP concentration of the irradiated samples were determined immediately after the laser irradiation. While the maximum reduction in the TVC observed with respect to the control was 59% immediately after 15 min irradiation, the ATP concentration showed a reduction of about 86% for the same duration. The ATP concentration showed an abrupt reduction from 3 min of laser irradiation and continued to reduce significantly with increasing duration of irradiation. Thus, 3 min irradiation at a fluence of 0.1 J cm^{m 2} is considered as an approximate threshold for ATP production in this bacterium. As the decreased level of ATP production continued, bacterial mortality resulted. The reduction in ATP production could be due to damage caused by the laser irradiations on bacterial metabolic processes such as cellular respiration.     

Utilization of the ferrous sulfate (Fricke) dosimeter for evaluating the radioprotective potential of cystamine: experiment and Monte Carlo simulation

Cystamine, an organic disulfide (RSSR), is among the best of the known radiation-protective compounds and has been used to protect normal tissues in clinical radiation therapy. Recently, it has also proved to be beneficial in the treatment of disorders of the central nervous system in animal models. However, the underlying mechanism of its action at the chemical level is not yet well understood. The present study aims at using the ferrous sulfate (Fricke) dosimeter to quantitatively evaluate, both experimentally and theoretically, the radioprotective potential of this compound. The well-known radiolysis of the Fricke dosimeter by (60)Co γ rays or fast electrons, based on the oxidation of ferrous ions to ferric ions by the oxidizing species (?)OH, HO(2)(?), and H(2)O(2) produced in the radiolytic decomposition of water, forms the basis for our method. The presence of cystamine in Fricke dosimeter solutions during irradiation prevents the radiolytic oxidation of Fe(2+) and leads to decreased ferric yields (or G values). The observed decrease in G(Fe(3+)) increases upon increasing the concentration of the disulfide compound over the range 0-0.1 M under both aerated and deaerated conditions. To help assess the basic radiation-protective mechanism of this compound, a full Monte Carlo computer code is developed to simulate in complete detail the radiation-induced chemistry of the studied Fricke/cystamine solutions. Benefiting from the fact that cystamine is reasonably well characterized in terms of radiation chemistry, this computer model proposes reaction mechanisms and incorporates specific reactions describing the radiolysis of cystamine in aerated and deaerated Fricke solutions that lead to the observable quantitative chemical yields. Results clearly indicate that the protective effect of cystamine originates from its radical-capturing ability, which allows this compound to act by competing with the ferrous ions for the various free radicals--especially (?)OH radicals and H(?) atoms--formed during irradiation of the surrounding water. Most interestingly, our simulation modeling also shows that the predominant pathway in the oxidation of cystamine by (?)OH radicals involves an electron-transfer mechanism, yielding RSSR(?+) and OH(-). A very good agreement is found between calculated G(Fe(3+)) values and experiment. This study concludes that Monte Carlo simulations represent a very efficient method for understanding indirect radiation damage at the molecular level.     

Elementary processes of photoperception by phytochrome A for high-irradiance response of hypocotyl elongation in Arabidopsis

Elementary processes of photoperception by phytochrome A (PhyA) for the high-irradiance response (HIR) of hypocotyl elongation in Arabidopsis were examined using a newly designed irradiator with LED. The effect of continuous irradiation with far-red (FR) light could be replaced by intermittent irradiation with FR light pulses if given at intervals of 3 min or less for 24 h. In this response, the Bunsen-Roscoe law of reciprocity held in each FR light pulse. Therefore, we determined the action spectrum for the response by intermittent irradiation using phyB and phyAphyB double mutants. The resultant action spectrum correlated well with the absorption spectrum of PhyA in far-red-absorbing phytochrome (Pfr). Intermittent irradiation with 550 to 667 nm of light alone had no significant effect on the response. In contrast, intermittent irradiation with red light immediately after each FR light pulse completely reversed the effect of FR light in each cycle. The results indicate that neither red-absorbing phytochrome synthesized in darkness nor photoconverted Pfr are physiologically active, and that a short-lived signal is induced during photoconversion from Pfr to red-absorbing phytochrome. The mode of photoperception by PhyA for HIR is essentially different from that by PhyA for very-low-fluence responses and phytochrome B for low-fluence responses.