Peculiarities of the effect of low-dose-rate radiation simulating high-altitude flight conditions on mice in vivo

In the present work, the effect of a low-dose rate of high-LET radiation in polychromatic erythrocytes of mice bone marrow was investigated in vivo. The spectral and component composition of the radiation field used was similar to that present in the atmosphere at an altitude of about 10 km. The dose dependence, adaptive response, and genetic instability in the F₁ generation born from males irradiated under these conditions were examined using the micronucleus test. Irradiation of the mice was performed for 24 h per day in the radiation field behind the concrete shield of the Serpukhov accelerator. Protons of 70 GeV were used over a period of 15–31 days, to accumulate doses of 11.5–31.5 cGy. The experiment demonstrated that irradiation of mice in vivo in this dose range leads to an increase in cytogenetic damage to bone marrow cells, but does not induce any adaptive response. In mice pre-irradiated with a dose of 11.5 cGy, an increase in sensitivity was observed after an additional irradiation with a dose of 1.5 Gy. The absence of an adaptive response suggests existence of genetic instability.     

Depression of p53-independent Akt survival signals in human oral cancer cells bearing mutated p53 gene after exposure to high-LET radiation

Although mutations and deletions in the p53 tumor suppressor gene lead to resistance to low linear energy transfer (LET) radiation, high-LET radiation efficiently induces cell lethality and apoptosis regardless of the p53 gene status in cancer cells. Recently, it has been suggested that the induction of p53-independent apoptosis takes place through the activation of Caspase-9 which results in the cleavage of Caspase-3 and poly (ADP-ribose) polymerase (PARP). This study was designed to examine if high-LET radiation depresses serine/threonine protein kinase B (PKB, also known as Akt) and Akt-related proteins. Human gingival cancer cells (Ca9-22 cells) harboring a mutated p53 (mp53) gene were irradiated with 2 Gy of X-rays or Fe-ion beams. The cellular contents of Akt-related proteins participating in cell survival signaling were analyzed with Western Blotting 1, 2, 3 and 6h after irradiation. Cell cycle distributions after irradiation were assayed with flow cytometric analysis. Akt-related protein levels decreased when cells were irradiated with high-LET radiation. High-LET radiation increased G(2)/M phase arrests and suppressed the progression of the cell cycle much more efficiently when compared to low-LET radiation. These results suggest that high-LET radiation enhances apoptosis through the activation of Caspase-3 and Caspase-9, and suppresses cell growth by suppressing Akt-related signaling, even in mp53 bearing cancer cells.     

Activation of antioxidative enzymes induced by low-dose-rate whole-body gamma irradiation: adaptive response in terms of initial DNA damage

An adaptive response induced by long-term low-dose-rate irradiation in mice was evaluated in terms of the amount of DNA damage in the spleen analyzed by a comet assay. C57BL/ 6N female mice were irradiated with 0.5 Gy of (137)Cs gamma rays at 1.2 mGy/h; thereafter, a challenge dose (0.4, 0.8 or 1.6 Gy) at a high dose rate was given. Less DNA damage was observed in the spleen cells of preirradiated mice than in those of mice that received the challenge dose only; an adaptive response in terms of DNA damage was induced by long-term low-dose-rate irradiation in mice. The gene expression of catalase and Mn-SOD was significantly increased in the spleen after 23 days of the low-dose-rate radiation (0.5 Gy). In addition, the enzymatic activity of catalase corresponded to the gene expression level; the increase in the activity was observed at day 23 (0.5 Gy). These results suggested that an enhancement of the antioxidative capacities played an important role in the reduction of initial DNA damage by low-dose-rate radiation.     

Transgenerational genomic instability in the first generation offspring of mice exposed to low-intensity red and near-infrared irradiation in vivo

Transgenerational genomic instability in the first generation offspring of mice exposed to lowintensity infrared laser (632.8 nm) and light-emitting-diode infrared irradiation (850 nm) was investigated in vivo. It was found that the level of spontaneous damage in bone marrow according to the micronucleus test, the level of reactive oxygen species in whole blood, and the mass index of lymphoid organs in all of the descendants of irradiated mice did not increase. After additional X-ray exposure of the progeny at a dose rate of 1.5 Gy, a decrease in the level of damage and the absence of an adaptive response were revealed upon testing according to “radiosensitivity” and the radiation-induced adaptive-response scheme (0.1 + 1.5 Gy), respectively, compared to the descendants of nonirradiated mice. The rate of tumor growth in the offspring of irradiated mice did not differ from that in the descendants of nonirradiated mice, although inhibition of the tumor growth rate was observed in their irradiated parents. The survival rate after irradiation at a dose rate of 6.5 Gy did not differ from both the parents and the control.

     

Radiation adaptive response of glial cells.

There are various types of radiation in space including high energy particles. It is, therefore, becoming to be important to study the low dose and low dose-rate effects in space radiation biology. Radiation adaptive response (RAR) for cell growth and its mechanism were examined using cultured glial cells. The cells from hippocampus of Wistar rats were irradiated with a low dose (0.1 Gy) of X-rays and 3 h after with a high dose (2 Gy). Decrease in the rate of cell growth with 2 Gy was suppressed by the 0.1 Gy preirradiation, when cells were counted 2 days after irradiation. The inhibitors of protein kinase C (PKC) and DNA-dependent protein kinase (DNAPK) or phosphatidylinositol 3-kinase (PI3K) suppressed RAR. The treatment with the activators of PKC instead of 0.1 Gy-preirradiation also caused adaptive response to 2 Gy-irradiation. Moreover, glial cells cultured from severe combined immunodeficiency (scid) mice, which have lost DNAPK activity, and AT-2KY cells, fibroblasts of an ataxia-telangiectasia (AT) patient, showed no RAR. These results indicated that PKC, ATtM, DNAPK and/or PI3K were involved in RAR for growth of cultured glial cells. Proteomics [correction of preteomics] analysis of these cells exposed to low dose irradiation in now underway.     

Indicators of hippocampal neurogenesis are altered by 56Fe-particle irradiation in a dose-dependent manner

The health risks to astronauts exposed to high-LET radiation include possible cognitive deficits. The pathogenesis of radiation-induced cognitive injury is unknown but may involve loss of neural precursor cells from the subgranular zone (SGZ) of the hippocampal dentate gyrus. To address this hypothesis, adult female C57BL/6 mice received whole-body irradiation with a 1 GeV/nucleon iron-particle beam in a single fraction of 0, 1, 2 and 3 Gy. Two months later mice were given BrdU injections to label proliferating cells. Subsequently, hippocampal tissue was assessed using immunohistochemistry for detection of proliferating cells and immature neurons. Routine histopathological methods were used to qualitatively assess tissue/cell morphology in the hippocampal formation and adjacent areas. When compared to controls, irradiated mice showed progressively fewer BrdU-positive cells as a function of dose. This observation was confirmed by Ki-67 immunostaining in the SGZ showing reductions in a dose-dependent fashion. The progeny of the proliferating SGZ cells, i.e. immature neurons, were visualized by doublecortin staining and were significantly reduced by irradiation, with the decreases ranging from 34% after 1 Gy to 71% after 3 Gy. Histopathology showed that in addition to cell changes in the SGZ, (56)Fe particles induced a chronic and diffuse astrocytosis and changes in pyramidal neurons in and around the hippocampal formation. The present data provide the first evidence that high-LET radiation has deleterious effects on cells associated with hippocampal neurogenesis.     

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.     

The remote effects of radiation after hypofractionated irradiation with protons of Ehrlich ascites carcinoma in mice

This work focuses on the study of remote effects (duration of remission, recurrence rate, and average lifespan) in mice with Ehrlich ascites carcinoma exposed to oligofractionated irradiation with a pencil scanning beam of protons depending on the volume of the tissue being irradiated and the interval between dose fractions. The results show higher antitumor efficacy and a considerable increase in the average life span of mice after hypofractionated irradiation with a pencil scanning beam of protons at a total dose of 60 Gy of the gross tumor volume compared with the planning target volume.     

低水平辐射诱导的细胞遗传学适应性反应

先用0.01GY x-射线(剂量率:0.01GY/分)体外照射人、兔外周血,经不同时间后再用1.5GY X-射线(0.44GY/分)照射,发现在G_0、G_1、S和G_2期受0.01GY X-射线照射后再给大剂量照射者,其染色体畸变率明显低于单纯受1.5GY X-射线照射组(P<0.01)。这一适应性反应能持续3个细胞周期,在接受小剂量照射后超过3个细胞周期再受大剂量照射者,染色体畸变率未见减少。若在第三细胞周期以后再次给予小剂量照射,可再次诱导适应性反应。用小鼠整体小剂量照射后骨髓细胞和生殖细胞亦出现这种适应性反应。另外也探讨了不同剂量和不同剂量率的预先照射对适应性反应的影响。     

Effects of gamma-irradiation on the infectivity and chromosome aberration of Clonorchis sinensis

Effects of gamma irradiation on the worm survival and chromosomal aberration of Clonorchis sinensis were studied. The metacercariae irradiated with various amounts of gamma radiation (ranging from 5 Gy to 50 Gy) were fed to rats, and the effects were compared with those of non-irradiated controls. Recovery rates of adult worms in irradiated groups were reduced gradually as increasing of the irradiation doses. No worm was recovered from rats which were fed with 50 Gy irradiated metacercariae. The chromosome number was 2n = 56 in all worms from all experimental groups. However, the groups irradiated with 20 Gy, 25 Gy or 30 Gy showed variations in the chromosome number, depending on different cells in the same individual. Radiation doses used in this study did not appear to induce chromosome aberrations, however, irradiation with 30 Gy showed slightly reduced chromosome size.     

Relative cataractogenic effects of X rays, fission-spectrum neutrons, and 56Fe particles: a comparison with mitotic effects

The eyes of Sprague-Dawley rats were irradiated with doses of 2.5-10 Gy 250-kVp X rays, 1.25-2.25 Gy fission-spectrum neutrons (approximately 0.85 MeV), or 0.1-2.0 Gy 600-MeV/A 56Fe particles. Lens opacifications were evaluated for 51-61 weeks following X and neutron irradiations and for 87 weeks following X and 56Fe-particle irradiations. Average stage of opacification was determined relative to time after irradiation, and the time required for 50% of the irradiated lenses to achieve various stages (T50) was determined as a function of radiation dose. Data from two experiments were combined in dose-effect curves as T50 experimental values taken as percentages of the respective T50 control values (T50-% control). Simple exponential curves best describe dose responsiveness for both high-LET radiations. For X rays, a shallow dose-effect relationship (shoulder) up to 4.5 Gy was followed at higher doses by a steeper exponential dose-effect relationship. As a consequence, RBE values for the high-LET radiations are dose dependent. Dose-effect curves for cataracts were compared to those for mitotic abnormalities observed when quiescent lens epithelial cells were stimulated mechanically to proliferate at various intervals after irradiation. Neutrons were about 1.6-1.8 times more effective than 56Fe particles for inducing both cataracts and mitotic abnormalities. For stage 1 and 2 cataracts, the X-ray Dq was 10-fold greater and the D0 was similar to those for mitotic abnormalities initially expressed after irradiation.     

Suppression of thymic lymphoma induction by life-long low-dose-rate irradiation accompanied by immune activation in C57BL/6 mice

The induction of thymic lymphomas by whole-body X irradiation with four doses of 1.8 Gy (total dose: 7.2 Gy) in C57BL/6 mice was suppressed from a high frequency (90%) to 63% by preirradiation with 0.075 Gy X rays given 6 h before each 1.8-Gy irradiation. This level was further suppressed to 43% by continuous whole-body irradiation with 137Cs gamma rays at a low dose rate of 1.2 mGy/h for 450 days, starting 35 days before the challenging irradiation. Continuous irradiation at 1.2 mGy/h resulting in a total dose of 7.2 Gy over 258 days yielded no thymic lymphomas, indicating that this low-dose-rate radiation does not induce these tumors. Further continuous irradiation up to 450 days (total dose: 12.6 Gy) produced no tumors. Continuously irradiated mice showed no loss of hair and a greater body weight than unirradiated controls. Immune activities of the mice, as measured by the numbers of CD4+ T cells, CD40+ B cells, and antibody-producing cells in the spleen after immunization with sheep red blood cells, were significantly increased by continuous 1.2-mGy/h irradiation alone. These results indicate the presence of an adaptive response in tumor induction, the involvement of radiation-induced immune activation in tumor suppression, and a large dose and dose-rate effectiveness factor (DDREF) for tumor induction with extremely low-dose-rate radiation.     

The effect of low-dose rate radiation simulating the high-altitude flight conditions on mice in vivo

In present work, we investigated the peculiarities of the effect of a low-dose rate high-LET radiation that simulates the spectral and component composition of the radiation field formed in the atmosphere at a height of 10 km on mice in vivo. The dose dependence and adaptive response were examined. Irradiation of mice was performed for 24 h a day in the radiation field behind the concrete shield of the Serpukhov accelerator of 70 GeV protons for the time (15-31 days) necessary to accumulate the required doses. The experiments demonstrated that irradiation of mice in vivo in the dose range of 11.5-31.5 cGy leads to an increase in cytogenetic damage to bone marrow cells and induces no adaptive response in bone marrow cells.     

Enhanced body radioresistance resulting from the transfusion of autologous blood irradiated with low doses of ionizing radiation]

Preliminary administration of autogenic blood irradiated in vitro with ionizing radiation in small doses of 0.05, 0.3 or 0.5 Gy resulted in a pronounced increase in the radioresistance of mice [correction of rats] subsequently irradiated in a dose of 9 Gy. The optimum was autotransfusion of blood irradiated in a dose of 0.3 Gy a day or 10 days prior to the total irradiation which increased the survival rate of experimental animals to 80% while, in control groups, the survival rate was only 10%.     

Protection against radiation oxidative damage in mice by Triphala

Protection against whole body gamma-irradiation (WBI) of Swiss mice orally fed with Triphala (TPL), an Ayurvedic formulation, in terms of mortality of irradiated animals as well as DNA damage at cellular level has been investigated. It was found that radiation induced mortality was reduced by 60% in mice fed with TPL (1g/kg body weight/day) orally for 7 days prior to WBI at 7.5 Gy followed by post-irradiation feeding for 7 days. An increase in xanthine oxidoreductase activity and decrease in superoxide dismutase activity was observed in the intestine of mice exposed to WBI, which, however, reverted back to those levels of sham-irradiated controls, when animals were fed with TPL for 7 days prior to irradiation. These data have suggested the prevention of oxidative damage caused by whole body radiation exposure after feeding of animals with TPL. To further understand the mechanisms involved, the magnitude of DNA damage was studied by single cell gel electrophoresis (SCGE) in blood leukocytes and splenocytes obtained from either control animals or those fed with TPL for 7 days followed by irradiation. Compared to irradiated animals without administering TPL, the mean tail length was reduced about three-fold in blood leukocytes of animals fed with TPL prior to irradiation. Although, similar protection was observed in splenocytes of TPL fed animals, the magnitude of prevention of DNA damage was significantly higher than that observed in leukocytes. It has been concluded that TPL protected whole body irradiated mice and TPL induced protection was mediated through inhibition of oxidative damage in cells and organs. TPL seems to have potential to develop into a novel herbal radio-protector for practical applications.     

MiR-21 is continually elevated long-term in the brain after exposure to ionizing radiation

Ionizing radiation stimulates miR-21 expression in different types of mammalian cells in culture. However, it remains unclear whether radiation could stimulate miR-21 expression in brain cells and tissue and, if so, how long the upregulation of miR-21 would be maintained after exposure to different types of radiation. To answer these questions, we examined the miR-21 levels in irradiated mouse hippocampal cells and brain tissue from mice at different times up to 1 year after whole-body exposure to 0.5 Gy of X rays [low linear energy transfer (LET)] or (56)Fe ions (high LET). The results showed that radiation stimulated miR-21 expression in mouse hippocampal cells and upregulation of EGFR, which is similar to that in human hepatocytes, as we reported previously. Interestingly, the miR-21 levels gradually increased within 1 year after irradiation, although there was no significant difference in the miR-21 low- and high-LET irradiated mice. The high expression of miR-21 in the brain was also associated with high expression of EGFR in irradiated mice; thus our data strongly support that EGFR and miR-21 are in a positive regulatory loop, because it is known that radiation stimulates miR-21 through the EGFR/Stat3 pathway and miR-21 activates the EGFR pathway. Since the brain is relatively resistant to radiation-induced histomorphological changes, our findings may provide a new way to explore radiation-induced pathological changes in the brain by investigating miR-21 and its multiple targets.     

Effects of 1 GeV/nucleon ⁵⁶Fe particles on longevity, carcinogenesis and neuromotor ability in Atm-deficient mice

As therapeutic uses of high-LET radiation become more prevalent and human space exploration continues to be a focus of NASA, it is important to understand the biological effects of high-LET radiation and the role of genetics in sensitivity to high-LET radiation. To study genetic susceptibility to radiation, we used mice deficient in Atm activity (AtmΔSRI). ATM is important in DNA repair, apoptosis and cell cycle regulation. Although homozygous mutations in ATM are rare, the prevalence of ATM heterozygosity is estimated to be 1% and results in an increased cancer risk. We found that the effects of 1 Gy 1 GeV/nucleon ??Fe particles on life span and tumorigenesis are genotype- and sex-specific. Significant effects of 1 Gy 1 GeV/nucleon ??Fe particles on incidence of non-cancer end points were seen; however, 2 Gy 1 GeV/nucleon ??Fe particles significantly affected neuromotor ability. Our results represent an extensive investigation into the late effects of high-LET radiation exposure in a sex- and genotype-dependent manner and provide a baseline for understanding the long-term risks of high-LET radiation.     

The adaptive response modifies latency for radiation-induced myeloid leukemia in CBA/H mice.

We have investigated the effect of the adaptive response on acute myeloid leukemia (AML) induced in CBA/Harwell mice by a chronic radiation exposure. Groups of mice irradiated with a total dose of 1. 0 Gy at two different chronic dose rates (0.5, 0.004 Gy/h) had similar frequencies of AML. Compared to control animals that did not develop AML, irradiation at either of these dose rates did not change the longevity of the mice that did not die of leukemia. The survival rates of irradiated mice that did develop leukemia in the two groups were not different from each other, indicating that the dose rates produced similar responses and therefore were both chronic exposures. We then tested the ability of a chronic 10-cGy (0. 5 Gy/h) exposure to ionizing radiation, mild hyperthermia (40.5 degrees C whole-body, 60 min) or treatment with interleukin-1 (1500 U i.p.) to induce an adaptive response and modify the frequency or latency of AML which resulted from a subsequent (24 h later) 1.0-Gy (0.5 Gy/h) chronic radiation exposure. The frequency of radiation-induced leukemia was not changed in mice given any of the three adapting treatments 24 h prior to the chronic 1.0-Gy dose that induced leukemia. However, the latent period for development of AML was significantly increased by both the prior low radiation dose and mild hyperthermia treatment. Injection of interleukin-1, in contrast, may have reduced the latent period. Similar to the single 1.0-Gy chronic exposure alone, none of the adapting treatments prior to that exposure influenced the survival of animals that did not develop AML. These results indicate that an earlier exposure to a small adapting dose of radiation or to a mild heat stress can influence secondary steps in radiation-induced carcinogenesis.     

不同剂量的X-射线全身照射对小鼠健康状况及涎腺的影响

目的:通过直线加速器全身照射昆明小鼠建立辐射损伤模型,探索不同放射剂量对小鼠健康状况及涎腺功能和结构的影响。方法:选取八种不同剂量对昆明小鼠行体外全身照射,于照射后一个月内观察小鼠生长情况、体重变化;照射后一周、一个月检测各组小鼠血象的变化;测定放射半数致死剂量;照射后两个月,测定各组小鼠的唾液流量及唾液淀粉酶含量,并对下颌下腺组织切片行HE染色。结果:13Gy和15Gy照射组小鼠的体重逐渐下降,一周后死亡,其余组小鼠体重最终呈增加趋势。X-射线全身照射的半数致死量为10Gy。照射后一周,照射组小鼠的白细胞数目明显降低,与对照组比较有明显统计学差异(P0.01);在其他血象方面,除了7Gy组外,其他照射组与对照组比较也均有统计学差异(P0.05)。照射一个月后,各照射组小鼠的血象均恢复正常。照射后两个月,9Gy组和11Gy组小鼠的唾液流量及唾液淀粉酶含量均显著低于0Gy组,且11Gy组较9Gy组亦明显降低,差异均有统计学意义(P0.05)。随照射剂量的增加,小鼠的下颌下腺腺泡细胞数目逐步减少,结构排列紊乱,组织损伤逐渐加重。结论:X-射线全身照射引起小鼠健康状况受损,免疫功能减低,损伤程度与放射线强度呈剂量依赖性,小鼠半数致死量为10Gy,该剂量适合建立全身放射损伤模型。