Behavioral performance and microglial status in mice after moderate dose of proton irradiation.

Cognitive impairment is a remote effect of gamma radiation treatment of malignancies. The major part of the studies on the effect of proton irradiation (a promising alternative in the treatment of radio-resistant tumors and tumors located close to critical organs) on the cognitive abilities of laboratory animals and their relation to morphological changes in the brain is rather contradictory. The aim of this study was to investigate cognitive functions and the dynamics of changes in morphological parameters of hippocampal microglial cells after 7.5 Gy of proton irradiation. Two months after the cranial irradiation, 8- to 9-week-old male SHK mice were tested for total activity, spatial learning, as well as long- and short-term hippocampus-dependent memory. To estimate the morphological parameters of microglia, brain slices of control and irradiated animals each with different time after proton irradiation (24 h, 7 days, 1 month) were stained for microglial marker Iba-1. No changes in behavior or deficits in short-term and long-term hippocampus-dependent memory were found, but an impairment of episodic memory was observed. A change in the morphology of hippocampal microglial cells, which is characteristic of the transition of cells to an activated state, was detected. One day after proton exposure in the brain tissue, a slight decrease in cell density was observed, which was restored to the control level by the 30th day after treatment. The results obtained may be promising with regard to the future use of using high doses of protons per fraction in the irradiation of tumors.

Growth Induction of Solid Ehrlich Ascitic Carcinoma in Mice after Proton Irradiation of Tumor Cells Ex Vivo.

This study presents data on the growth rate and frequency of induction of the solid form of Ehrlich's ascites carcinoma (EAC) in mice in the short and long term after inoculation of ascitic cells irradiated ex vivo with a proton beam in the dose range of 30-150 Gy. It was shown that the growth rate of solid tumors after inoculation of irradiated cells ex vivo coincided with the growth of tumors in the control group. The frequency of tumor induction in mice after inoculation of EAC cells irradiated at a dose of 30 Gy was 80%, 60 Gy-60%, 90 Gy-25%, and 120 Gy-10%; at irradiation at a dose of 150 Gy, no tumors appeared during the entire observation period. Thus, we determined the dose of proton radiation required to eliminate tumor cells and/or signaling factors that can lead to the induction of tumor growth of EAC in mice.

Anti-tumor Effect of High Doses of Carbon Ions and X-Rays during Irradiation of Ehrlich Ascites Carcinoma Cells Ex Vivo.

The effect of carbon ions (12C) with the energy of 400 MeV/nucleon on the dynamics of induction and growth rate of solid tumors in mice under irradiation of Ehrlich ascites carcinoma cells (EAC) ex vivo at doses of 5-30 Gy relative to the action of equally effective doses of X-ray radiation was studied. The dynamics of tumor induction under the action of 12C and X-rays had a similar character and depended on the dose during 3 months of observation. The value of the latent period, both when irradiating cells with 12C and X-ray, increased with increasing dose, and the interval for tumor induction decreased. The rate of tumor growth after ex vivo irradiation of EAC cells was independent of either dose or type of radiation. The dose at which EAC tumors are not induced within 90 days was 30 Gy for carbon ions and 60 Gy for X-rays. The value of the relative biological effectiveness of carbon ions, calculated from an equally effective dose of 50% probability of tumors, was 2.59.

The Effect of Xylazine/Zoletil Anesthesia on the Radiosensitivity of Mice under Total Irradiation with X-Rays, Protons, and Carbon Ions.

The effect of xylazine-zoletyl anesthesia on the radiosensitivity of mice irradiated with protons, carbon ions in two regions of the Bragg curve, and X-ray radiation was studied according to the criteria of 30-day survival, dynamics of death, and the average life span of dead mice. The maximum effect of anesthesia by 3.3 times was observed by a decrease in the death of animals during irradiation with carbon ions at the Bragg peak; in the case of irradiation before the peak, the effect was 1.2 times. In the case of proton irradiation at the Bragg peak, the protective effect of anesthesia by a factor of 1.7 was observed only at a dose of 8.5 Gy. When mice were irradiated with X-rays in the dose range of 6.0-8.5 Gy, the anesthesia effect coefficient was 1.7-2. According to the 30-day survival method, it was shown that the use of a xylazine-zoletil mixture significantly changes the radiosensitivity of mice depending on the radiation dose and the radiation source quality.

Radioprotective Effect of Nanocerium by Irradiation of Mice with Carbon Ions in Medium and Lethal Doses.

The data of the study of the radioprotective properties of nanocerium (nCeO2) after total irradiation of mice with carbon ions in medium and lethal doses according to the micronucleus test and the criterion of 30-day survival are presented. A significant protective effect of nCeO2 upon irradiation at medium doses was observed at per os administration for 5 days before irradiation (that is, at long-term prophylactic use). Mouse survival data showed no protective effect of per os administration of nCeO2 in contrast to the micronucleus test results. After injections of both nCeO2 and saline solution 24 h before or immediately after irradiation, the radioprotective effect was detected using both methods. The data obtained revealed the dependence of the observed effects on the mode and time of nCeO2 administration, the influence of the solvent, the level of doses and the quality of radiation, as well as demonstrated the possibility of using nanocerium preparations to protect organisms from radiation with high LET values and the importance of further studies of the radioprotective properties of new nanomaterials.

Low-Intensity Femtosecond Radiation Activates the Natural Defenses of Mice in vivo.

The possibility of induction of cytogenetic damage in the bone marrow, changes in the cellularity of lymphoid organs and blood composition in mice irradiated with low-intensity femtosecond laser radiation at a power flux density of 5.1, 10.4, and 52 mJ/cm2 (0.5 mW for 5, 10, and 50 s) in vivo was shown. Using the radiation adaptive response test (0.1 Gy + 1.5 Gy), it was found that, when mice were exposed to femtosecond laser radiation in high doses, the body's natural defenses were activated in the same narrow range of energy flux density (2-16 mJ/cm2) as in the case of X-ray irradiation in a dose of 0.1 Gy (4 mJ/cm2). The data obtained suggest a similar mechanism of activation of the body's natural defense upon exposure to low doses of both ionizing and non-ionizing radiation.

Assessment of the Relative Biological Efficiency of Pencil Beam Scanning of Protons in Mice in Vivo.

The effect of proton pencil beam scanning in the dose range of 4.5-15 Gy on the radiosensitivity of mice under irradiation in two regions of the Bragg curve was studied according to the criteria of 30-day survival, dynamics of death, and average lifespan of mice. The relative biological effectiveness (RBE) value of protons relative to X-ray radiation before and at the Bragg peak determined by the LD50/30 index was 0.86 and 0.94, respectively, and by the criterion of 30-day survival at a dose of 6.5 Gy it was 0.83 and 0.84, respectively. With similar RBE values for protons in different regions of the Bragg curve, significant differences in the dynamics of the course of radiation sickness were revealed, which indicates different damage to critical systems and organs of animals and the induction of compensatory mechanisms involved in the formation of stress responses at the organismal level.

Combined Effect of Neutron and Proton Radiations on the Growth of Solid Ehrlich Ascites Carcinoma and Remote Effects in Mice.

The combined effect of the irradiation with a proton pencil scanning beam (PBS) at a total dose of 80 Gy and neutron radiation at a dose of 5 Gy on the growth of solid Ehrlich ascites carcinoma (EAC) and the remote effects in tumor-bearing mice was studied. Combined irradiation of mice with neutrons before and after irradiation with PBS, as well as irradiation only with PBS, effectively suppressed the growth of solid EAC within 1 month. In terms of the frequency and severity of radiation-induced skin reactions of mice observed 15-40 days after therapy, neutron irradiation after the irradiation with PBS showed better values of these parameters as compared to only PBS; however, exposure to neutrons before PBS was more damaging as compared to the other two options. It was also shown that the tumor relapse rate in the groups of animals with combined irradiation was higher, and the total lifespan was lower than the group of mice irradiated with PBS alone.

The Effect of Low and Medium Doses of Proton Pencil Scanning Beam on the Blood-Forming Organs during Total Irradiation of Mice.

The aim of this work was to study the effect of proton pencil beam scanning in the Bragg peak in the dose range of 0.1-1.5 Gy on the induction of cytogenetic damage in the bone marrow, reactive oxygen species (ROS) production in whole blood, and the state of lymphoid organs after total body irradiation of mice. Irradiation was carried out in the Prometeus proton synchrotron (Protvino) in the Bragg peak with proton energy at the output of 90-116 MeV. It was found that, under irradiation of mice in the range of low and medium doses of proton pencil beam scanning in the Bragg peak, the relative biological effectiveness (RBE) according to the criterion of cytogenetic changes was 1.15. In addition, it was found that the pathophysiological effect on the lymphoid organs and the production of ROS by blood cells were different as compared with the effect of X-rays.

Combined Effect of Low-Intensity Helium-Neon Laser and X-Ray Radiation on in Vivo Cellular Response of the Whole Blood and Lymphoid Organs in Mice.

We studied the effect of exposure to helium-neon laser (dose range 0.16-50 mJ/cm2) on activation of natural protection reserve in mice using the adaptive response test. DNA comets method revealed a protective response manifested in DNA damage level in whole blood leukocytes of mice and in lymphoid organs by the thymus and spleen weight index; preexposure to laser did not induce the adaptive response. ROS level in the whole blood was assessed by the level of zymosan-induced luminol chemiluminescence. In mice subjected to adaptive laser irradiation in doses of 0.16-5 mJ/cm2 followed by X-ray irradiation in a dose of 1.5 Gy, the activation index calculated as the ratio of induced to spontaneous area of luminescence was by 1.4 times lower than that in non-irradiated animals, which attested to reduced ROSgeneration reserve capacity of neutrophils.

Induction of the Adaptive Response in Mice Exposed to He-Ne Laser and X-Ray Radiation.

We studied the dose-dependent induction of in vivo adaptive response in the bone marrow and blood of mice exposed to low-intensity radiation of He-Ne laser (633 nm) and X-ray radiation by the severity of cytogenetic injury and intensity of ROS production, respectively. Induction of the adaptive response in mice preexposed to He-Ne laser and X-ray radiation depended on the adaptive dose and the interval between the adaptive and main doses and correlated with changes in ROS generation. The adaptive response after exposure to low-intensity ionizing and non-ionizing radiation was observed in the same dose range, which attests to similar mechanisms of its induction.

[Induction of DNA damage in blood leucocytes and of cytogenetic injuries in bone marrow polychromatic erythrocytes in mice exposed to low-LET and high-LET radiation and in their progeny].

The present work was aimed at studying the molecular and cellular levels of the response of the hematopoietic system in mice and their progeny to the action of low-LET and high-LET radiation at different times after exposure. The damage to the genome at the molecular level was assessed by the comet assay in peripheral blood leucocytes, whereas at the cellular level it was estimated by means of the micronuclear test in the marrow cells, after exposure of mice to X-radiation of 1, 3 and 5 Gy and to a high-LET low-intensity radiation at thedoses of 0.14 and 0.35 Gy, as well as to a combined effect of these types of radiation. When accessing the level of the DNA damage to individual cells by the comet assay, we also used, apart from a commonly accepted parameter %TDNA, additional characteristics: the proportions of leucocytes with an intact and highly fragmented DNA. Using these parameters, we detected the changes characterizing the dynamics of the leukocyte population in mouse blood at different times after the action of X-ray and high-LET radiation. It was found that: (1) the DNA damage increases with the dose of high-LET radiation; (2) the level of damage in the progeny of the animals exposed to high-LET radiation does not differ from that in unirradiated animals both at the molecular and cytogenetic levels; and (3) a decrease in the radiosensitivity of the progeny of the mice exposed to high-LET radiation at a dose of 0.35 Gy makes itself evident only at the molecular level, which may point to the possible transgeneration transmission of genomic lesions.

[Influence of low-dose-rate red and near-infrared radiations on the level of reactive oxygen species, the genetic apparatus and the tumor growth in mice in vivo].

The effect of low-dose-rate red and near-infrared radiations from the matrix of light emitted diode (650 nm and 850 nm) and a He-Ne laser (633 nm) on activation of the reserve of a natural defense system in the mice exposed to radiation in vivo was studied by the level of reactive oxygen species (ROS) production in blood cells, the induction of cytogenetic adaptive response in bone marrow cells, thymus and spleen, and the rate of Ehrlich ascites carcinoma growth in a solid form. As a positive control animals were irradiated with X-rays by the scheme of the radiation-induced adaptive response (0.1 Gy + 1.5 Gy). The levels of ROS production was assessed in whole blood by luminol-dependent chemiluminescence, of cytogenetic damage--by the "micronucleus test" in the bone marrow, the weight of the thymus and spleen--by index of organ, and the rate of tumor growth--according to its size for 30 days after inoculation. Adaptogenic and anticarcinogenic effects of studied radiations were revealed. The values of these effects were not different from those in animals pre-irradiated with the X-rays. The relationship between the level of ROS production and adaptive response induction in the mice under the influence of non-ionizing radiation was first ascertained. The experimental data obtained may indicate a similar mechanism of induction of protective responses to ionizing and non-ionizing radiations in mice in vivo.

Effect of infrared and X-ray radiation on thymus cells and the rate of growth of Ehrlich carcinoma.

We studied the effect of infrared light with a wavelength of 850 nm and modulated frequency of 101 Hz and X-ray radiation on the induction of cross-adaptive and radiation responses in the thymus and on the rate of tumor growth in mice in vivo. Preliminary exposure to infrared and X-ray radiation was shown to result in recovery in thymus weight after irradiation in a dose of 1.5 Gy and also inhibited the growth rate of Ehrlich carcinoma. These data attest to common mechanisms of the adaptive response induced by infrared and X-ray radiation in mice. Infrared light can be used as an adaptogen to adapt the animals to adverse factors.

Delayed effects of chronic low-dose high linear energy transfer (LET) radiation on mice in vivo.

In the present work, the delayed effects of chronic high linear energy transfer (LET) radiation in polychromatic erythrocytes (PCEs) of mice bone marrow were investigated in vivo. Irradiation of the two-month-old SHK white mongrel random-bred male mice was performed in the radiation field behind the concrete shield of the accelerator of 70 GeV protons to accumulate doses of 0.005-0.16 Gy. The dependence of the biological response on dose, adaptive response (AR) and genomic instability (GI) in F(1) and F(2) generations from males irradiated with doses of 0.005 and 0.16 Gy and from males exposed to combined action of immunomodulator-bendazol hydrochloride (BH) and of 0.16 Gy irradiation, were examined using the micronucleus formation test. The data demonstrated that irradiation of mice with these doses lead to an increase in the level of cytogenetic damage and induces no AR. With analysis of the bone marrow radiosensitivity to 1.5 Gy of X rays and the capacity to AR it was found that the chronic high-LET irradiation of parents induced the GI at least two generations. The combined exposure to BH and the dose of 0.16 Gy induces no AR in F(0) generation but induces AR in F(1) and F(2) offspring.

[The effect of infrared and X-ray radiation on the production of reactive oxygen species in blood and induction of cytogenetic damage in the bone marrow of mice in vivo].

The goal of the present work was to study the effect of infrared light (IRL) at a wavelength of 850 nm modulated by a frequency of 101 Hz with a mode of power 22 mW/cm2 and X-rays with a voltage of 200 kV at a dose rate of 1 Gy/min on the production of reactive oxygen species (ROS) in blood cells using luminol-dependent chemiluminescence, as well as on the induction of a cytogenetic damage in bone marrow cells of mice by the in vivo micronucleus test. The experiments performed have shown: 1) the level of the ROS production in blood of the mice exposed to IRL and X-rays at an adapting dose of 0.1 Gy reaches the peak value after 0.5 h and drops to the ROS level in untreated animals 5 h after either exposure; 2) irradiation of mice with IRL and X-rays at a dose of 0.1 Gy induces adaptive responses both in blood cells and bone marrow cells of mice. These adaptive responses were revealed only 5 h after both exposures, when the level of ROS production decreased to the ROS level in untreated animals; they are equal in magnitude and dynamics and persist up to 2 months.

101 Hz frequency-modulated infrared light induces cytogenetic adaptive response in mouse bone marrow in vivo.

The micronucleus test was used tu study the possibility of inducing cross-adaptive response in mouse bone marrow cells in vivo with an 850 nm infrared light modulated by a 101 Hz frequency, emitted by a light therapy device "Kurator". We demonstrated that this exposure led to a substantial reduction of cytogenetic cell damage produced by further exposure of animals to X-radiation in the dose of 1.5 Gy, i.e. it induced an adaptive response which did not differ by the magnitude and time course from the adaptive response to radiation.

[Study of the genetic instability in generations of mice irradiated of a low-dose rate of high-LET radiation].

In present work, we investigated the genetic instability in mice of F1, of F2 and of F3 generations born from males irradiated by a low-dose rate of high-LET radiation that simulates the spectral and component composition of radiation fields formed in the conditions of high-altitude flights in vivo in polychromatic erythrocytes of bone marrow using the micronucleus test. Two-month-old males of SHK white mongrel mice were used. Irradiation was performed for 24 h a day in the radiation field behind the concrete shield of the U-70 accelerator of 70 GeV protons (Serpukhov) to accumulate doses of 11.5, of 21.5 and of 31.5 cGy (1 cGy/day). The experiments demonstrated that in mice of F1 generation born from males irradiated with doses of 11.5, 21.5 and of 31.5 cGy, an increase in sensitivity to additional irradiation with a dose of 1.5 Gy of gamma-radiation and the absence of adaptive response compared with the descendants of unirradiated males occur. In contrast to F1 generation genetic instability in mice of the F2 and F3 generations was revealed only by the absence of adaptive response. These data indicate a genetic instability in F1, F2 and F3 generations born from irradiated males.

[Peculiar effects of low doses of high let radiation on mice].

Experiments with exposure of mice to low doses of chronic high-LET radiation were carried out in the radiation field behind the concrete wall of the Serpukhov accelerators of protons with the energy of 70 GeV. The goal was to study dose dependence, radiation adaptive response (AR), and genetic instability. Mice (SHK strain) were irradiated continuously 15, 24 and 31 days which corresponded to the doses of 11.5, 21.5 and 31.5 Gy. Cytogenetic damages were determined using the micronuclear test in marrow polychromatophil erythrocytes. It was shown that all the experimental doses aggravated the cytogenetic damage; however, no AR induction in marrow cells was observed. Males of the F1 generation born from the males irradiated at 11.5 Gy had same level of spontaneous cytogenetic damage as males born from non-irradiated parents. Yet, they displayed an exaggerated sensitivity to additional exposure to 1.5 Gy and no AR induction by the standard gamma-protocol which is indicative of genetic instability.