Purpose Pediatric patients undergoing cranial irradiation are at risk of developing neurocognitive impairments, with radiation-induced damage to the hippocampi playing a central role in this process. This damage is primarily driven by loss of neural stem/progenitor cells (NSCs/NPCs) and neuroinflammation. Noradrenergic agonists can potentially protect against radiation-induced hippocampal damage by reducing cell death and neuroinflammation. This study aims to assess whether pharmacologic boosting of noradrenaline (NA) can reduce radiation-induced hippocampal injury and prevent memory decline. Materials and Methods Mice were exposed to cranial irradiation of 10 Gy at postnatal day 22, and treated with noradrenergic agonists, reboxetine (REB), or atipamezole (ATI). At 6 hours, 1 week, and 1 month postirradiation, hippocampal cell death, phosphorylated cAMP responsive element-binding protein (pCREB), and brain-derived neurotrophic factor (BDNF) expression, loss of hippocampal NSCs/NPCs, and microglial inflammation were histologically investigated. At 1 month postirradiation, memory deficits and hippocampal atrophy were investigated by a behavioral test battery and ex vivo magnetic resonance imaging, respectively. Results We report that REB and ATI significantly mitigate radiation-induced hippocampal toxicity. Treatment with noradrenergic agonists reduces radiation-induced apoptosis, microglial inflammation, and preserves pCREB and BDNF expression in the hippocampus early after irradiation. It also attenuates the loss of hippocampal NSCs/NPCs and immature neurons. After radiation therapy, considerable hippocampal volume loss is observed, which is prevented by REB treatment. Finally, behavioral testing revealed that REB treatment after radiation therapy successfully mitigates radiation-induced memory dysfunction. Conclusions The present findings uncover the marked benefits of NA treatment against radiation-induced hippocampal injury. We demonstrate that REB is able to mitigate radiation-induced memory dysfunction, which highlights the potential of NA treatment for addressing long-standing complications faced by pediatric brain tumor survivors.

The skeletal muscle and the immune system are heavily affected by the space environment. The crosstalk between these organs, although established, is not fully understood. This study determined the nature of immune cell changes in the murine skeletal muscle following (hindlimb) unloading combined with an acute session of irradiation (HLUR). Our findings show that 14 days of HLUR induces a significant increase of myeloid immune cell infiltration in skeletal muscle.

Human spaceflight is associated with several health-related issues as a result of long-term exposure to microgravity, ionizing radiation, and higher levels of psychological stress. Frequent reported skin problems in space include rashes, itches, and a delayed wound healing. Access to space is restricted by financial and logistical issues; as a consequence, experimental sample sizes are often small, which limits the generalization of the results. Earth-based simulation models can be used to investigate cellular responses as a result of exposure to certain spaceflight stressors. Here, we describe the development of an in vitro model of the simulated spaceflight environment, which we used to investigate the combined effect of simulated microgravity using the random positioning machine (RPM), ionizing radiation, and stress hormones on the wound-healing capacity of human dermal fibroblasts. Fibroblasts were exposed to cortisol, after which they were irradiated with different radiation qualities (including X-rays, protons, carbon ions, and iron ions) followed by exposure to simulated microgravity using a random positioning machine (RPM). Data related to the inflammatory, proliferation, and remodeling phase of wound healing has been collected. Results show that spaceflight stressors can interfere with the wound healing process at any phase. Moreover, several interactions between the different spaceflight stressors were found. This highlights the complexity that needs to be taken into account when studying the effect of spaceflight stressors on certain biological processes and for the aim of countermeasures development.

Previous studies suggested a causal link between pre-natal exposure to ionizing radiation and birth defects such as microphthalmos and exencephaly. In mice, these defects arise primarily after high-dose X-irradiation during early neurulation. However, the impact of sublethal (low) X-ray doses during this early developmental time window on adult behavior and morphology of central nervous system structures is not known. In addition, the efficacy of folic acid (FA) in preventing radiation-induced birth defects and persistent radiation-induced anomalies has remained unexplored. To assess the efficacy of FA in preventing radiation-induced defects, pregnant C57BL6/J mice were X-irradiated at embryonic day (E)7.5 and were fed FA-fortified food. FA partially prevented radiation-induced (1.0 Gy) anophthalmos, exencephaly and gastroschisis at E18, and reduced the number of pre-natal deaths, fetal weight loss and defects in the cervical vertebrae resulting from irradiation. Furthermore, FA food fortification counteracted radiation-induced impairments in vision and olfaction, which were evidenced after exposure to doses ≥0.1 Gy. These findings coincided with the observation of a reduction in thickness of the retinal ganglion cell and nerve fiber layer, and a decreased axial length of the eye following exposure to 0.5 Gy. Finally, MRI studies revealed a volumetric decrease of the hippocampus, striatum, thalamus, midbrain and pons following 0.5 Gy irradiation, which could be partially ameliorated after FA food fortification. Altogether, our study is the first to offer detailed insights into the long-term consequences of X-ray exposure during neurulation, and supports the use of FA as a radioprotectant and antiteratogen to counter the detrimental effects of X-ray exposure during this crucial period of gestation.

In preparation for upcoming space missions to the Moon and Mars, there is a need to understand how space stressors (e.g. microgravity, radiation) affect different physiological systems. As skeletal muscle is a critical organ, not only for locomotion but also for overall body homeostasis, defining the molecular impact of microgravity and radiation on this tissue will help developing new, or fine-tuning current countermeasures to maintain health and physiological function of space travellers. PURPOSE: To investigate the effects of combined radiation and unloading on anabolic/catabolic and immune/inflammatory processes on skeletal muscle in mice. METHODS: Ten C57/BL6J mice were subjected to 14-d hind-limb unloading by tail suspension with an acute radiation session (dose=25 mGy, X-ray) on day 7 of unloading (HLUR). Ten mice were used as control (CTRL; similar cages, sham radiation). Mice were sacrified and soleus muscle was immediately dissected, weighed and frozen. Then, RNA was extracted and converted to cDNA. Gene expression of anabolic/catabolic (i.e. myostatin, MuRF-1, Atrogin-1, PGC-1α) and immune/inflammatory markers (i.e. CD4, CD8, IFNγ, CD11b, MHCII, TNF, IL-6) was assessed by RT-PCR. Independent t-tests were use to compare HLUR vs. CTRL. RESULTS: Soleus muscle weight was ~30% lower in HLUR vs. CTRL (P<0.001). Myostatin expression was greater in HLUR vs. CTRL (1.8-fold, P=0.014). MHCII expression was higher in HLUR vs. CTRL (2.4-fold, P<0.001). There was a trend for group differences (P<0.08) in CD11b and TNF mRNA content with HLUR showing greater values than CTRL. Gene expression of CD4, CD8 and IFNγ was barely detected in either group. CONCLUSIONS: The combination of unloading and radiation has a major impact on skeletal muscle. Apart from inducing muscle atrophy, as indicated by the decreased muscle weight and increased myostatin levels, these two space stressors altered the immune profile within the muscle. The increased gene expression of MHCII and CD11b indicates that the myeloid component of the immune system is activated upon unloading and radiation in skeletal muscle. In contrast, the almost undetected mRNA levels of CD4, CD8 and IFNγ may imply that unloading and acute radiation have little impact on the lymphoid component. These findings should be followed up with immunohistochemical analysis.

Spaceflight-induced bone losses have been reliably reproduced in Hind-Limb-Unloading (HLU) rodent models. However, a considerable knowledge gap exists regarding the effects of low-dose radiation and microgravity together. Ten-week-old male C57BL/6J mice, randomly allocated to Control (CONT), Hind-Limb Unloading (HLU), and Hind-Limb Unloading plus Irradiation (HLUIR), were acclimatized at 28 °C, close to thermoneutral temperature, for 28 days prior to the 14-day HLU protocol. HLUIR mice received a 25 mGy dose of X-ray irradiation, simulating 14 days of exposure to the deep space radiation environment, on day 7 of the HLU protocol. Trabecular bone mass was similarly reduced in HLU and HLUIR mice when compared to CONT, with losses driven by osteoclastic bone resorption rather than changes to osteoblastic bone formation. Femoral cortical thickness was reduced only in the HLUIR mice (102 μm, 97.5–107) as compared to CONT (108.5 μm, 102.5–120.5). Bone surface area was also reduced only in the HLUIR group, with no difference between HLU and CONT. Cortical losses were driven by osteoclastic resorption on the posterior endosteal surface of the distal femoral diaphysis, with no increase in the numbers of dead osteocytes. In conclusion, we show that low-dose radiation exposure negatively influences bone physiology beyond that induced by microgravity alone.

Prenatal irradiation is known to perturb brain development. Epidemiological studies revealed that radiation exposure during weeks 8-15 of pregnancy was associated with an increased occurrence of mental disability and microcephaly. Such neurological deficits were reproduced in animal models, in which rodent behavioral testing is an often used tool to evaluate radiation-induced defective brain functionality. However, up to now, animal studies suggested a threshold dose of around 0.30 Gray (Gy) below which no behavioral alterations can be observed, while human studies hinted at late defects after exposure to doses as low as 0.10 Gy. Here, we acutely irradiated pregnant mice at embryonic day 11 with doses ranging from 0.10 to 1.00 Gy. A thorough investigation of the dose-response relationship of altered brain function and architecture following in utero irradiation was achieved using a behavioral test battery and volumetric 3D T2-weighted magnetic resonance imaging (MRI). We found dose-dependent changes in cage activity, social behavior, anxiety-related exploration, and spatio-cognitive performance. Although behavioral alterations in low-dose exposed animals were mild, we did unveil that both emotionality and higher cognitive abilities were affected in mice exposed to ≥0.10 Gy. Microcephaly was apparent from 0.33 Gy onwards and accompanied by deviations in regional brain volumes as compared to controls. Of note, total brain volume and the relative volume of the ventricles, frontal and posterior cerebral cortex, cerebellum, and striatum were most strongly correlated to altered behavioral parameters. Taken together, we present conclusive evidence for persistent low-dose effects after prenatal irradiation in mice and provide a better understanding of the correlation between their brain size and performance in behavioral tests.

Background: In humans, in utero exposure to ionising radiation results in an increased prevalence of neurological aberrations, such as small head size, mental retardation and decreased IQ levels. Yet, the association between early damaging events and long-term neuronal anomalies remains largely elusive.Methods: Mice were exposed to different X-ray doses, ranging between 0.0 and 1.0 Gy, at embryonic days (E) 10, 11 or 12 and subjected to behavioural tests at 12 weeks of age. Underlying mechanisms of irradiation at E11 were further unravelled using magnetic resonance imaging (MRI) and spectroscopy, diffusion tensor imaging, gene expression profiling, histology and immunohistochemistry.Results: Irradiation at the onset of neurogenesis elicited behavioural changes in young adult mice, dependent on the timing of exposure. As locomotor behaviour and hippocampal-dependent spatial learning and memory were most particularly affected after irradiation at E11 with 1.0 Gy, this condition was used for further mechanistic analyses, focusing on the cerebral cortex and hippocampus. A classical p53-mediated apoptotic response was found shortly after exposure. Strikingly, in the neocortex, the majority of apoptotic and microglial cells were residing in the outer layer at 24 h after irradiation, suggesting cell death occurrence in differentiating neurons rather than proliferating cells. Furthermore, total brain volume, cortical thickness and ventricle size were decreased in the irradiated embryos. At 40 weeks of age, MRI showed that the ventricles were enlarged whereas N-acetyl aspartate concentrations and functional anisotropy were reduced in the cortex of the irradiated animals, indicating a decrease in neuronal cell number and persistent neuroinflammation. Finally, in the hippocampus, we revealed a reduction in general neurogenic proliferation and in the amount of Sox2-positive precursors after radiation exposure, although only at a juvenile age.Conclusions: Our findings provide evidence for a radiation-induced disruption of mouse brain development, resulting in behavioural differences. We propose that alterations in cortical morphology and juvenile hippocampal neurogenesis might both contribute to the observed aberrant behaviour. Furthermore, our results challenge the generally assumed view of a higher radiosensitivity in dividing cells. Overall, this study offers new insights into irradiation-dependent effects in the embryonic brain, of relevance for the neurodevelopmental and radiobiological field.

Ionizing radiation is a potent activator of the tumor suppressor gene p53, which itself regulates the transcription of genes involved in canonical pathways such as the cell cycle, DNA repair and apoptosis as well as other biological processes like metabolism, autophagy, differentiation and development. In this study, we performed a meta-analysis on gene expression data from different in vivo and in vitro experiments to identify a signature of early radiation-responsive genes which were predicted to be predominantly regulated by p53. Moreover, we found that several genes expressed different transcript isoforms after irradiation in a p53-dependent manner. Among this gene signature, we identified novel p53 targets, some of which have not yet been functionally characterized. Surprisingly, in contrast to genes from the canonical p53-regulated pathways, our gene signature was found to be highly enriched during embryonic and post-natal brain development and during in vitro neuronal differentiation. Furthermore, we could show that for a number of genes, radiation-responsive transcript variants were upregulated during development and differentiation, while radiation non-responsive variants were not. This suggests that radiation exposure of the developing brain and immature cortical neurons results in the p53-mediated activation of a neuronal differentiation program. Overall, our results further increase the knowledge of the radiation-induced p53 network of the embryonic brain and provide more evidence concerning the importance of p53 and its transcriptional targets during mouse brain development.

At the gastrula phase of development, just after the onset of implantation, the embryo proper is characterized by extremely rapid cell proliferation. The importance of DNA repair is illustrated by embryonic lethality at this stage after ablation of the genes involved. Insight into mutation induction is called for by the fact that women often do not realize they are pregnant, shortly after implantation, a circumstance which may have important consequences when women are subjected to medical imaging using ionizing radiation. We screened gastrula embryos for DNA synthesis, nuclear morphology, growth, and chromosome aberrations (CA) shortly after irradiation with doses up to 2.5 Gy. In order to obtain an insight into the importance of DNA repair for CA induction, we included mutants for the non-homologous end joining (NHEJ) and homologous recombination repair (HRR) pathways, as well as Parp1-/- and p53+/- embryos. With the pUR288 shuttle vector assay, we determined the radiation sensitivity for point mutations and small deletions detected in young adults. We found increased numbers of abnormal nuclei 5 h after irradiation; an indication of disturbed development was also observed around this time. Chromosome aberrations 7 h after irradiation arose in all genotypes and were mainly of the chromatid type, in agreement with a cell cycle dominated by S-phase. Increased frequencies of CA were found for NHEJ and HR mutants. Gastrula embryos are unusual in that they are low in exchange induction, even after compromised HR. Gastrula embryos were radiation sensitive in the pUR288 shuttle vector assay, giving the highest mutation induction ever reported for this genetic toxicology model. On theoretical grounds, a delayed radiation response must be involved. The compromised developmental profile after doses up to 2.5 Gy likely is caused by both apoptosis and later cell death due to large deletions. Our data indicate a distinct radiation-sensitive profile of gastrula embryos, including some stage-specific aspects that are not as yet understood. (C) 2015 Elsevier B.V. All rights reserved.

Microgravity and cosmic rays as found in space are difficult to recreate on earth. However, ground-based models exist to simulate space flight experiments. In the present study, an experimental model was utilized to monitor gene expression changes in fetal skin fibroblasts of murine origin. Cells were continuously subjected for 65 h to a low dose (55 mSv) of ionizing radiation (IR), comprising a mixture of high‑linear energy transfer (LET) neutrons and low-LET gamma-rays, and/or simulated microgravity using the random positioning machine (RPM), after which microarrays were performed. The data were analyzed both by gene set enrichment analysis (GSEA) and single gene analysis (SGA). Simulated microgravity affected fetal murine fibroblasts by inducing oxidative stress responsive genes. Three of these genes are targets of the nuclear factor‑erythroid 2 p45-related factor 2 (Nrf2), which may play a role in the cell response to simulated microgravity. In addition, simulated gravity decreased the expression of genes involved in cytoskeleton remodeling, which may have been caused by the downregulation of the serum response factor (SRF), possibly through the Rho signaling pathway. Similarly, chronic exposure to low-dose IR caused the downregulation of genes involved in cytoskeleton remodeling, as well as in cell cycle regulation and DNA damage response pathways. Many of the genes or gene sets that were altered in the individual treatments (RPM or IR) were not altered in the combined treatment (RPM and IR), indicating a complex interaction between RPM and IR.

Ionizing radiation can elicit harmful effects on the cardiovascular system at high doses. Endothelial cells are critical targets in radiation-induced cardiovascular damage. Astronauts performing a long-term deep space mission are exposed to consistently higher fluences of ionizing radiation that may accumulate to reach high effective doses. In addition, cosmic radiation contains high linear energy transfer (LET) radiation that is known to produce high values of relative biological effectiveness (RBE). The aim of this study was to broaden the understanding of the molecular response to high LET radiation by investigating the changes in gene expression in endothelial cells. For this purpose, a human endothelial cell line (EA.hy926) was irradiated with accelerated nickel ions (Ni) (LET, 183 keV/µm) at doses of 0.5, 2 and 5 Gy. DNA damage was measured 2 and 24 h following irradiation by γ-H2AX foci detection by fluorescence microscopy and gene expression changes were measured by microarrays at 8 and 24 h following irradiation. We found that exposure to accelerated nickel particles induced a persistent DNA damage response up to 24 h after treatment. This was accompanied by a downregulation in the expression of a multitude of genes involved in the regulation of the cell cycle and an upregulation in the expression of genes involved in cell cycle checkpoints. In addition, genes involved in DNA damage response, oxidative stress, apoptosis and cell-cell signaling (cytokines) were found to be upregulated. An in silico analysis of the involved genes suggested that the transcription factors, E2F and nuclear factor (NF)-κB, may be involved in these cellular responses.

BACKGROUND:Radioadaptation is a phenomenon whereby cells exposed to a low dose of ionizing radiation are more resistant to a much higher dose delivered some time thereafter. This phenomenon could result from the activation of damage repair and/or antioxidant defense systems by the low dose.MATERIALS AND METHODS:The existence of a cytogenetic adaptive response in female germ cells was investigated using a recently developed in vitro system. Mouse ovarian follicles were cultured from an early preantral stage up to ovulation. The follicles were X-irradiated with either 2 or 4 Gy ("challenge dose") preceded or not by 50 mGy ("conditioning dose", 5 h earlier), on days 0 or 12 of the culture. Ovulated oocytes were collected on day 13, fixed and analyzed for the presence of chromosome aberrations.RESULTS:Irradiation with 2 or 4 Gy on days 0 or 12 did not influence ovulation but had dose-dependent effects on the germinal vesicle breakdown of the oocytes. It also caused dose-dependent chromosome damage, with a greater sensitivity of oocytes to this effect when irradiation occurred on day 12 than on day 0. Prior irradiation of oocytes with the dose of 50 mGy led to a reduction in the yield of chromosome aberrations when irradiation occurred on day 12 but not on day 0.CONCLUSION:These results suggest that pre-irradiation of mouse pre-ovulatory oocytes with a low conditioning dose could confer on them some protection against radiation-induced chromosomal damage by a subsequent challenge dose of a few Gy.

Many space missions have shown that prolonged space flights may increase the risk of cardiovascular problems. Using a three‐dimensional clinostat, we investigated human endothelial EA.hy926 cells up to 10 days under conditions of simulated microgravity (µg) to distinguish transient from long‐term effects of µg and 1g. Maximum expression of all selected genes occurred after 10 min of clinorotation. Gene expression (osteopontin, Fas, TGF‐β1) declined to slightly upregulated levels or rose again (caspase‐3) after the fourth day of clinorotation. Caspase‐3, Bax, and Bcl‐2 protein content was enhanced for 10 days of microgravity. In addition, long‐term accumulation of collagen type I and III and alterations of the cytoskeletal alpha‐ and beta‐tubulins and F‐actin were detectable. A significantly reduced release of soluble factors in simulated microgravity was measured for brain‐derived neurotrophic factor, tissue factor, vascular endothelial growth factor (VEGF), and interestingly for endothelin‐1, which is important in keeping cardiovascular balances. The gene expression of endothelin‐1 was suppressed under µg conditions at days 7 and 10. Alterations of the vascular endothelium together with a decreased release of endothelin‐1 may entail post‐flight health hazards for astronauts. J. Cell. Biochem. J. Cell. Biochem. 101: 1439–1455, 2007. © 2007 Wiley‐Liss, Inc.