56Fe Particles Research Articles

Galactic Cosmic Ray Particle Exposure Does Not Increase Protein Levels of Inflammation or Oxidative Stress Markers in Rat Microglial Cells In Vitro.

Astronauts on exploratory missions will be exposed to galactic cosmic rays (GCR), which can induce neuroinflammation and oxidative stress (OS) and may increase the risk of neurodegenerative disease. As key regulators of inflammation and OS in the CNS, microglial cells may be involved in GCR-induced deficits, and therefore could be a target for neuroprotection. This study assessed the effects of exposure to helium (4He) and iron (56Fe) particles on inflammation and OS in microglia in vitro, to establish a model for testing countermeasure efficacy. Rat microglia were exposed to a single dose of 20 cGy (300 MeV/n) 4He or 2 Gy 56Fe (600 MeV/n), while the control cells were not exposed (0 cGy). Immediately following irradiation, fresh media was applied to the cells, and biomarkers of inflammation (cyclooxygenase-2 [COX-2], nitric oxide synthase [iNOS], phosphorylated IκB-α [pIκB-α], tumor necrosis factor-α [TNFα], and nitrite [NO2-]) and OS (NADPH oxidase [NOX2]) were assessed 24 h later using standard immunochemical techniques. Results showed that radiation did not increase levels of NO2- or protein levels of COX-2, iNOS, pIκB-α, TNFα, or NOX2 compared to non-irradiated control conditions in microglial cells (p > 0.05). Therefore, microglia in isolation may not be the primary cause of neuroinflammation and OS following exposures to helium or iron GCR particles.

Effects of HZE-Particle Exposure Location and Energy on Brain Inflammation and Oxidative Stress in Rats.

Astronauts on exploratory missions will be exposed to particle radiation of high energy and charge (HZE particles), which have been shown to produce neurochemical and performance deficits in animal models. Exposure to HZE particles can produce both targeted effects, resulting from direct ionization of atoms along the particle track, and non-targeted effects (NTEs) in cells that are distant from the track, extending the range of potential damage beyond the site of irradiation. While recent work suggests that NTEs are primarily responsible for changes in cognitive function after HZE exposures, the relative contributions of targeted and non-targeted effects to neurochemical changes after HZE exposures are unclear. The present experiment was designed to further explore the role of targeted and non-targeted effects on HZE-induced neurochemical changes (inflammation and oxidative stress) by evaluating the effects of exposure location and particle energy/linear energy transfer (LET). Forty-six male Sprague-Dawley rats received head-only or body-only exposures to 56Fe particles [600 MeV/n (75 cGy) or 1,000 MeV/n (100 cGy)] or 48Ti particles [500 MeV/n (50 cGy) or 1,100 MeV/n (75 cGy)] or no irradiation (0 cGy). Twenty-four h after irradiation, rats were euthanized, and the brain was dissected for analysis of HZE-particle-induced neurochemical changes in the hippocampus and frontal cortex. Results showed that exposure to 56Fe and 48Ti ions produced changes in measurements of brain inflammation [glial fibrillary astrocyte protein (GFAP)], oxidative stress [NADPH-oxidoreductase-2 (NOX2)] and antioxidant enzymes [superoxide dismutase (SOD), glutathione S-transferase (GST), nuclear factor erythroid 2-related factor 2 (Nrf2)]. However, radiation effects varied depending upon the specific measurement, brain region, and exposure location. Although overall exposures of the head produced more detrimental changes in neuroinflammation and oxidative stress than exposures of the body, body-only exposures also produced changes relative to no irradiation, and the effect of particle energy/LET on neurochemical changes was minimal. Results indicate that both targeted and non-targeted effects are important contributors to neurochemical changes after head-only exposure. However, because there were no consistent neurochemical changes as a function of changes in track structure after head-only exposures, the role of direct effects on neuronal function is uncertain. Therefore, these findings, although in an animal model, suggest that NTEs should be considered in the estimation of risk to the central nervous system (CNS) and development of countermeasures.

Astrocytes regulate vascular endothelial responses to simulated deep space radiation in a human organ-on-a-chip model.

Central nervous system (CNS) damage by galactic cosmic ray radiation is a major health risk for human deep space exploration. Simulated galactic cosmic rays or their components, especially high Z-high energy particles such as 56Fe ions, cause neurodegeneration and neuroinflammation in rodent models. CNS damage can be partially mediated by the blood-brain barrier, which regulates systemic interactions between CNS and the rest of the body. Astrocytes are major cellular regulators of blood-brain barrier permeability that also modulate neuroinflammation and neuronal health. However, astrocyte roles in regulating CNS and blood-brain barrier responses to space radiation remain little understood, especially in human tissue analogs. In this work, we used a novel high-throughput human organ-on-a-chip system to evaluate blood-brain barrier impairments and astrocyte functions 1-7 days after exposure to 600 MeV/n 56Fe particles and simplified simulated galactic cosmic rays. We show that simulated deep space radiation causes vascular permeability, oxidative stress, inflammation and delayed astrocyte activation in a pattern resembling CNS responses to brain injury. Furthermore, our results indicate that astrocytes have a dual role in regulating radiation responses: they exacerbate blood-brain barrier permeability acutely after irradiation, followed by switching to a more protective phenotype by reducing oxidative stress and pro-inflammatory cytokine and chemokine secretion during the subacute stage.

Beamline and Flight Comparisons of the ARMAS Flight Module With the Tissue Equivalent Proportional Counter for Improving Atmospheric Radiation Monitoring Accuracy

AbstractIonizing radiation at aircraft and commercial suborbital spaceflight altitudes is driven by space weather and is a health concern for crew and passengers. We compare the response functions of two radiation detectors that were exposed to four different ground‐based laboratory radiation fields as well as flown alongside each other on aircraft. The detectors were a tissue equivalent proportional counter (TEPC) and a Teledyne silicon micro dosimeter chip that was integrated into an Automated Radiation Measurements for Aerospace Safety Flight Module (ARMAS FM). Both detectors were flown onboard commercial and research aircraft. In addition, both detectors were exposed neutrons at the Los Alamos Neutron Science Center, protons at Loma Linda University Medical Center, 56Fe particles at the NASA Space Radiation Laboratory, and also a gamma radiation source at Lawrence Livermore National Laboratory. The response of each of these instruments as well as derived dosimetric quantities are compared for each radiation exposure and the ratio for converting ARMAS absorbed dose in silicon to an estimated absorbed dose in tissue is obtained. This process resulted in the first definitive calibration of the silicon‐based detector like ARMAS to TEPC. In particular, with seven flights of both instruments together, the ARMAS‐derived dose in tissue was then validated with the TEPC‐measured dose in tissue and these results are reported. This work provides a method for significantly improving the accuracy of radiation measurements relevant to human tissue safety using a silicon detector that is easy to deploy and can report data in real time.

Effects of partial- or whole-body exposures to 56Fe particles on brain function and cognitive performance in rats

On exploratory class missions, such as a mission to Mars, astronauts will be exposed to particles of high energy and charge (HZE particles). Exposure to HZE particles produces changes in neuronal function and can disrupt cognitive performance. Cells throughout the entire body, not just the brain, will be impacted by these particles. To determine the possible effects that irradiation of the body might have on neuronal function and cognitive performance, rats were given head-only, body-only or whole-body exposures to 56Fe particles. Cognitive performance (novel object recognition, operant responding) was tested in one set of animals; changes in brain function (oxidative stress, neuroinflammation) was tested in a second set of rats. The results indicated that there were no consistent differences in either behavioral or neurochemical endpoints as a function of the location of the irradiation. These results suggest that radiation to the body can impact the brain, therefore it may be necessary to re-evaluate the estimates of the risk of HZE particle-induced changes in neuronal function and cognitive performance.

Exposure to ≤15 cGy of 600 MeV/n 56Fe Particles Impairs Rule Acquisition but not Long-Term Memory in the Attentional Set-Shifting Assay.

On future missions into deep space, astronauts will be required to work more autonomously than on previous missions, and thus their ability to perform executive functions could be critical to mission success. In this study, we determined the effect that ≤15 cGy of 600 MeV/n 56Fe particles has on attentional set-shifting (ATSET) performance of ∼10 month-old (at the time of irradiation) male Wistar rats that had been prescreened for their ability to perform the task. Exposure to 1-15 cGy of 56Fe particles leads to a significant impairment in compound discrimination (CD) performance. Should similar effects occur in astronauts, an impaired ability to execute CD would result in a decreased ability to identify and maintain focus on relevant aspects of the task being performed. The use of rats that had been prescreened for ATSET performance helped to establish that working memory of the rules for the food reward remained intact (for at least 100 days) even after 15 cGy irradiation with 600 MeV/n 56Fe particles, but that 56Fe radiation exposure affected associative cue learning/acquisition rather than an intrinsic inability to perform the CD tasks. Our data suggest that declarative memory, and the ability to transitively infer established rules, also remained intact in the irradiated rats. Thus, should similar effects occur in astronauts, 56Fe-induced CD performance deficits may only be manifested in scenarios where astronauts are required to transitively apply their knowledge to solve problems that they have not previously encountered; nevertheless, potentially one-third of astronauts may not be able to perform event-critical tasks correctly. The implication of this data, from a probabilistic risk assessment perspective, is that cognitive performance studies that use naïve rodents, thus requiring task rule acquisition as well as task performance, are likely to overestimate the risk of 56Fe-induced cognitive deficits.

Impaired Attentional Set-Shifting Performance after Exposure to 5 cGy of 600 MeV/n 28Si Particles.

Astronauts on deep space missions will be required to work more autonomously than on previous missions, and thus their ability to perform executive functions could be critical to mission success. One of the most common measures of executive function in humans is the ability to perform attentional set shifting, which requires contributions from working memory, discrimination, reversal learning, attentional set shifting and attention. Rodent attentional set shifting assays require rats to form an association between the presence of the food reward and an associative cue, which is either the digging media or the scent that is placed in the bowl; by altering the combination of scent and digging media, progressively more complex cognitive processes can be tested. In this study, we have determined the effect that exposure to 5-20 cGy of 600 MeV/n 28Si particles has on the ability of male retired breeder Wistar rats to perform attentional set shifting at three months postirradiation. All doses of Si resulted in a significant impairment in the ability of the rats to perform the first and most simple step of the ATSET assay, the simple discrimination (SD) task. If astronauts were to experience HZE-induced SD impairments, they would be unable to identify key factors to successfully resolve a situation. Performance in at least one other component of the ATSET test was impaired at all doses studied, however, these varied according to the dose. Compared with our previous studies using 1 GeV/n 56Fe and 48Ti particles, 600 MeV/n 28Si ions impaired attentional set-shifting performance at lower doses than the heavier ions. However, when the effect of isofluences of the three HZE ions were compared, there were no significant differences in the severity of the impaired performance; there were, however, ion-specific decrements in the ability of rats to perform within the various stages of the test. This study further supports the notion that "mission-relevant" doses of HZE particles (<20 cGy) can impair certain aspects of attentional set-shifting performance in retired breeder rats, but there may be some ion-specific changes in the specific cognitive domains impaired.

Age as a factor in the responsiveness of the organism to the disruption of cognitive performance by exposure to HZE particles differing in linear energy transfer

Exposure to particles of high energy and charge (HZE particles) can produce decrements in cognitive performance. A series of experiments exposing rats to different HZE particles was run to evaluate whether the performance decrement was dependent on the age of the subject at the time of irradiation. Fischer 344 rats that were 2-, 11- and 15/16-months of age were exposed to 16O, 48Ti, or 4He particles at the NASA Space Radiation Laboratory at Brookhaven National Laboratory. As previously observed following exposure to 56Fe particles, exposure to the higher LET 48Ti particles produced a disruption of cognitive performance at a lower dose in the older subjects compared to the dose needed to disrupt performance in the younger subjects. There were no age related changes in the dose needed to produce a disruption of cognitive performance following exposure to lower LET 16O or 4He particles. The threshold for the rats exposed to either 16O or 4He particles was similar at all ages. Because the 11- and 15-month old rats are more representative of the age of astronauts (45–55 years old) the present results indicate that particle LET may be a critical factor in estimating the risk of developing a cognitive deficit following exposure to space radiation on exploratory class missions.

Neurochemical differences in learning and memory paradigms among rats supplemented with anthocyanin-rich blueberry diets and exposed to acute doses of 56Fe particles

The protective effects of anthocyanin-rich blueberries (BB) on brain health are well documented and are particularly important under conditions of high oxidative stress, which can lead to “accelerated aging.” One such scenario is exposure to space radiation, consisting of high-energy and -charge particles (HZE), which are known to cause cognitive dysfunction and deleterious neurochemical alterations. We recently tested the behavioral and neurochemical effects of acute exposure to HZE particles such as 56Fe, within 24–48h after exposure, and found that radiation primarily affects memory and not learning. Importantly, we observed that specific brain regions failed to upregulate antioxidant and anti-inflammatory mechanisms in response to this insult. To further examine these endogenous response mechanisms, we have supplemented young rats with diets rich in BB, which are known to contain high amounts of antioxidant-phytochemicals, prior to irradiation. Exposure to 56Fe caused significant neurochemical changes in hippocampus and frontal cortex, the two critical regions of the brain involved in cognitive function. BB supplementation significantly attenuated protein carbonylation, which was significantly increased by exposure to 56Fe in the hippocampus and frontal cortex. Moreover, BB supplementation significantly reduced radiation-induced elevations in NADPH-oxidoreductase-2 (NOX2) and cyclooxygenase-2 (COX-2), and upregulated nuclear factor erythroid 2-related factor 2 (Nrf2) in the hippocampus and frontal cortex. Overall results indicate that 56Fe particles may induce their toxic effects on hippocampus and frontal cortex by reactive oxygen species (ROS) overload, which can cause alterations in the neuronal environment, eventually leading to hippocampal neuronal death and subsequent impairment of cognitive function. Blueberry supplementation provides an effective preventative measure to reduce the ROS load on the CNS in an event of acute HZE exposure.

Radiation damage to neuronal cells: Simulating the energy deposition and water radiolysis in a small neural network

Radiation damage to the central nervous system (CNS) has been an on-going challenge for the last decades primarily due to the issues of brain radiotherapy and radiation protection for astronauts during space travel. Although recent findings revealed a number of molecular mechanisms associated with radiation-induced impairments in behaviour and cognition, some uncertainties exist in the initial neuronal cell injury leading to the further development of CNS malfunction. The present study is focused on the investigation of early biological damage induced by ionizing radiations in a sample neural network by means of modelling physico-chemical processes occurring in the medium after exposure. For this purpose, the stochastic simulation of incident particle tracks and water radiation chemistry was performed in realistic neuron phantoms constructed using experimental data on cell morphology. The applied simulation technique is based on using Monte-Carlo processes of the Geant4-DNA toolkit. The calculations were made for proton, 12C, and 56Fe particles of different energy within a relatively wide range of linear energy transfer values from a few to hundreds of keV/μm. The results indicate that the neuron morphology is an important factor determining the accumulation of microscopic radiation dose and water radiolysis products in neurons. The estimation of the radiolytic yields in neuronal cells suggests that the observed enhancement in the levels of reactive oxygen species may potentially lead to oxidative damage to neuronal components disrupting the normal communication between cells of the neural network.

Performance in hippocampus- and PFC-dependent cognitive domains are not concomitantly impaired in rats exposed to 20 cGy of 1 GeV/n 56Fe particles

NASA is currently conducting ground based experiments to determine whether the radiation environment that astronauts will encounter on deep space missions will have an impact on their long-term health and their ability to complete the various tasks during the mission. Emerging data suggest that exposure of rodents to mission-relevant HZE radiation doses does result in the impairment of various neurocognitive processes. An essential part of mission planning is a probabilistic risk assessment process that takes into account the likely incidence and severity of a problem. To date few studies have reported the impact of space radiation in a format that is amenable to PRA, and those that have only reported data for a single cognitive process. This study has established the ability of individual male Wistar rats to conduct a hippocampus-dependent (spatial memory) task and a cortex-dependent (attentional set shifting task) 90 days after exposure to 20cGy 1GeV/n 56Fe particles. Radiation-induced impairment of performance in one cognitive domain was not consistently associated with impaired performance in the other domain. Thus sole reliance upon a single measure of cognitive performance may substantially under-estimate the risk of cognitive impairment, and ultimately it may be necessary to establish the likelihood that mission-relevant HZE doses will impair performance in the three or four cognitive domains that NASA considers to be most critical for mission success, and build a PRA using the composite data from such studies.

Irradiation of Neurons with High-Energy Charged Particles: An In Silico Modeling Approach.

In this work, a stochastic computational model of microscopic energy deposition events is used to study for the first time damage to irradiated neuronal cells of the mouse hippocampus. An extensive library of radiation tracks for different particle types is created to score energy deposition in small voxels and volume segments describing a neuron’s morphology that later are sampled for given particle fluence or dose. Methods included the construction of in silico mouse hippocampal granule cells from neuromorpho.org with spine and filopodia segments stochastically distributed along the dendritic branches. The model is tested with high-energy 56Fe, 12C, and 1H particles and electrons. Results indicate that the tree-like structure of the neuronal morphology and the microscopic dose deposition of distinct particles may lead to different outcomes when cellular injury is assessed, leading to differences in structural damage for the same absorbed dose. The significance of the microscopic dose in neuron components is to introduce specific local and global modes of cellular injury that likely contribute to spine, filopodia, and dendrite pruning, impacting cognition and possibly the collapse of the neuron. Results show that the heterogeneity of heavy particle tracks at low doses, compared to the more uniform dose distribution of electrons, juxtaposed with neuron morphology make it necessary to model the spatial dose painting for specific neuronal components. Going forward, this work can directly support the development of biophysical models of the modifications of spine and dendritic morphology observed after low dose charged particle irradiation by providing accurate descriptions of the underlying physical insults to complex neuron structures at the nano-meter scale.

Protective effects of blueberry- and strawberry diets on neuronal stress following exposure to 56Fe particles

Particles of high energy and charge (HZE particles), which are abundant outside the magnetic field of the Earth, have been shown to disrupt the functioning of neuronal communication in critical regions of the brain. Previous studies with HZE particles, have shown that irradiation produces enhanced indices of oxidative stress and inflammation as well as altered neuronal function that are similar to those seen in aging. Feeding animals antioxidant-rich berry diets, specifically blueberries and strawberries, countered the deleterious effects of irradiation by reducing oxidative stress and inflammation, thereby improving neuronal signaling. In the current study, we examined the effects of exposure to 56Fe particles in critical regions of brain involved in cognitive function, both 36h and 30 days post irradiation. We also studied the effects of antioxidant-rich berry diets, specifically a 2% blueberry or strawberry diet, fed for 8 weeks prior to radiation as well as 30 days post irradiation. 56Fe exposure caused significant differential, neurochemical changes in critical regions of the brain, such as hippocampus, striatum, frontal cortex, and cerebellum, through increased inflammation, and increased oxidative stress protein markers. 56Fe exposure altered the autophagy markers, and antioxidant-rich berry diets significantly reduced the accumulation of p62 in hippocampus, a scaffold protein that co-localizes with ubiquitinated protein at the 30 days post irradiation time-point. Exposure to 56Fe particles increased the accumulation of disease-related proteins such as PHF-tau in the hippocampus of animals fed the control diet, but not in the irradiated animals fed the blueberry diet. These results indicate the potential protective effects of antioxidant-rich berry diets on neuronal functioning following exposure to HZE particles.

Effect of radiation quality on mutagenic joining of enzymatically-induced DNA double-strand breaks in previously irradiated human cells.

Previous work has shown that high charge and energy particle irradiation of human cells evokes a mutagenic repair phenotype, defined by increased mutagenic repair of new double-strand breaks that are introduced enzymatically, days or weeks after the initial irradiation. The effect was seen originally with 600 MeV/u (56)Fe particles, which have a linear energy transfer (LET) value of 174 keV/μm, but not with X rays or γ rays (LET ≤ 2 keV/μm). To better define the radiation quality dependence of the phenomenon, we tested two ions with intermediate LET values, 1,000 MeV/u (48)Ti (LET = 108 keV/μm) and 300 MeV/u (28)Si (LET = 69 keV/μm). These experiments used a previously validated assay, where a rare-cutting nuclease introduces double-strand breaks in two reporter transgene cassettes, which are located on different chromosomes. Deletions of a block of sequence in one of the cassettes, or translocations between cassettes, are measured independently using a multicolor fluorescence assay. The results showed that (48)Ti was a potent, but transient, inducer of mutagenic repair, based on increased frequency of nuclease-induced translocations. The (48)Ti ions did not affect the frequency of nuclease-induced deletions. The (28)Si ions had no measurable effect on either endpoint. There was a close correlation between the induction of the mutagenic repair phenomenon and the frequency of micronuclei in the targeted population (R(2) = 0.74), whereas there was no apparent correlation with radiation-induced cell inactivation. Together, these results better define the radiation quality dependence of the mutagenic repair phenomenon and establish its correlation, or lack of correlation, with other endpoints.

Iron Distribution in Size‐Resolved Aerosols Generated by UV‐Femtosecond Laser Ablation: Influence of Cell Geometry and Implications for In Situ Isotopic Determination by LA‐MC‐ICP‐MS

The influence of ablation cell geometry (Frames single‐ and HelEx two‐volume cells) and laser wavelength (198 and 266 nm) on aerosols produced by femtosecond laser ablation (fs‐LA) were evaluated. Morphologies, iron mass distribution (IMD) and 56Fe/54Fe ratios of particles generated from magnetite, pyrite, haematite and siderite were studied. The following two morphologies were identified: spherules (10–200 nm) and agglomerates (5–10 nm). Similarity in IMD and ablation rate at 198 and 266 nm indicates similar ablation mechanisms. 56Fe/54Fe ratios increased with aerodynamic particle size as a result of kinetic fractionation during laser plasma plume expansion, cooling and aerosol condensation. The HelEx cell produces smaller particles with a larger range of 56Fe/54Fe ratios (1.85‰) than particles from the Frames cell (1.16‰), but the bulk aerosol matches the bulk substrate for both cells, demonstrating stoichiometric fs‐LA sampling. IMD differences are the result of faster wash out of the HelEx cell allowing less time for agglomeration of small, low‐δ 56Fe particles with larger, high‐δ 56Fe particles in the cell. Even with a shorter ablation time, half the total Fe ion intensity, and half the ablation volume, the HelEx cell produced Fe isotope determinations for magnetite that were as precise as the Frames cell, even when the latter included an aerosol‐homogenising mixing chamber. The HelEx cell delivered a more constant stream of small particles to the ICP, producing a more stable Fe ion signal (0.7% vs. 1.5% RSE for 56Fe in a forty‐cycle single analysis), constant instrumental mass bias and thus a more precise measurement.

Neuronal stress following exposure to 56Fe particles and the effects of antioxidant-rich diets

Exposing young rats to particles of high energy and charge (HZE particles), a ground-based model for exposure to cosmic rays, enhances indices of oxidative stress and inflammation and disrupts the functioning of neuronal communication in critical regions of the brain [ 1]. These changes in neuronal function are similar to those seen in ageing [ 2, 3]. Although there is some recovery of function after exposure to 56Fe particles, particularly in changes observed 36 h following irradiation, long-term changes (75 days) have been observed, suggesting subcellular damage. Consequently, oxidative stress and inflammation induced by radiation could affect downstream events, such as changes in behavior and gene expression. Therefore, berry fruits high in antioxidant and anti-inflammatory activity, such as blueberries and strawberries, may prevent the occurrence of neurochemical and behavioral changes that occur if fed prior to radiation [ 4].Rats were exposed to 56Fe (1000 MeV/n; 1.5 Gy) particles at the NASA Space Radiation Laboratory at Brookhaven National Laboratory; other rats served as non-irradiated controls. The animals were fed either a control or a 2% blueberry or strawberry diet 8 weeks prior to radiation. Rats were then either euthanized at 36 h (short term) or 30 days following irradiation (long term). Before and after the irradiation, the animals were housed at USDA Human Nutrition Research Center on Aging at Tufts University, Boston.The results of the experiments indicate that: (1) 56Fe exposure caused significant differential, neurochemical changes in critical regions of the brain, such as hippocampus, striatum, frontal cortex and cerebellum, particularly long term. (2) Neurochemical changes resulted in the disruption of autophagy, increased inflammation and increased oxidative stress protein markers. (3) Antioxidant-rich berry diets significantly reduced the accumulation of toxic cellular debris in critical regions of the brain, primarily at the 30 days post-irradiation time-point. (4) Susceptibility to inflammation, autophagy dysregulation, and oxidative stress were proportional to the levels of antioxidant enzymes in the respective brain regions. (5) Exposure to 56Fe radiation may cause the accumulation of disease-related proteins such as PHF-Tau, which has been implicated in the pathogenesis of Alzheimer's disease.Irradiation with 56Fe, which causes substantial build-up of toxic cellular debris in critical regions of the brain, may overwhelm the innate antioxidant enzyme defense system [ 5]. Therefore, berry diets high in antioxidants may be used to counter these damaging effects by reducing oxidative stress and inflammation, and activating neuronal housekeeping, in addition to boosting endogenous antioxidant enzymes.This paper was presented at the NASA Session at Heavy Ion in Therapy and Space Radiation Symposium 2013.

Radiation-associated cardiovascular risks for future deep-space missions

Background: During the future Moon and Mars missions, astronauts will be exposed to space radiation (IR) for extended time. The majority of space flight-associated risks identified for the cardiovascular (CV) system to date were determined shortly after low Earth orbit (LEO) short- and long-duration space flights that include: serious cardiac dysrhythmias, compromised orthostatic CV response and manifestation of previously asymptomatic CV disease. Further ground-based experiments using a surrogate model of microgravity supported the space flight data for significant cardiac remodeling due to prolonged exposure to microgravity. These symptoms were determined to be a consequence of adaptation to microgravity that could be ameliorated by a post-mission exercise program, and were not identified as risk factors that were causatively related to space IR. Long-term degenerative effects of cosmic IR during and after space flights on CV system are unknown. It was suggested that due to GCR, each cell in an astronaut's body will be traversed by 1H every 3 days, helium (2He) nuclei every few weeks and high charge and energy (HZE) nuclei (e.g. 28Si, 56Fe) every few months. Despite the fact that only 1% of GCR is composed of ions heavier than helium, ∼41% of the IR dose-equivalent is predicted to be HZE particles with 13% being from 56Fe particles, only. During an exploration-class space mission to Mars, astronauts will not have access to comprehensive healthcare services for a period of at least 2–3 years. Since the majority of experienced astronauts are middle-aged (average age is 46, and the range is 33–58 years), they are at risk for developing serious CV events which could be life-threatening for the astronaut and mission-threatening for NASA. Therefore, it is important to evaluate the effects and potential CV risks caused by space IR. We hypothesized that: (i) low-dose space IR-induced biological responses may be long-lasting and are IR type-dependent; (ii) IR may increase CV risks in the aging heart (IR + AGING model) and affect the heart recovery after an adverse CV event, such as acute myocardial infarct (IR + AGING + AMI model). Methods: Eight- to 9-month-old C57BL/6N male mice were IR once with proton (1H) 50 cGy, 1 GeV/n or iron (56Fe) 15 cGy, 1 GeV/n. We evaluated IR-induced biological tissue responses—underlying molecular mechanisms, calcium handling, signal transduction, gene expression and cardiac fibrosis. Cardiac function was assessed by echocardiography (ECHO) and hemodynamic measurements (HEMO) as detailed in Fig. ​Fig.1.1. AMI was induced by ligation of left anterior descending coronary artery 1 and 3 months post-IR as detailed in Fig. ​Fig.22. Fig. 1. Radiation + aging model. Fig. 2. Radiation + aging model + adverse CV event model. Results: In the IR + AGING model, cardiac function was not different among the control and 1H-IR group, whereas left ventricular end-diastolic pressure (LVEDP) was significantly increased in 56Fe mice 1 and 3 months post-IR. There was a small but statistically significant (P 200% increases, P < 0.02) and 400% decreases in p-p38 MAPK (P < 0.05), suggesting activation of compensatory mechanisms in [Ca2+]i handling in these hearts. By 3 months, compared with control, 1H- and 56Fe-IR hearts had 200–500% (P < 0.02) decreases in SERCA2a and more than 200% decreases in p-Creb-1 (P < 0.02), suggesting reduced capacity in intracellular [Ca2+]i handling. These data suggest that dysfunction in [Ca2+]i handling combined with LVEDP increase after 56Fe-IR may arise from the excessive demand on the heart due to prolonged activation of compensatory mechanisms that lead to changes in SERCA2a and p-Creb1 levels. This may represent a possible intracellular mechanism of heart failure in development in 56Fe-IR hearts. In the IR + AGING + AMI model, no mortality was observed among three different groups 1 or 3 months post-IR and up to 28 days post-AMI. However, 1 month post-IR and 28 days post-AMI, the infarct size was significantly smaller in 56Fe-IR (p 10-fold increase in p-p38 MAPK level in 56Fe vs control and 1H-IR-AMI hearts, suggesting continuous decreases in the survival, proliferation and angiogenesis signaling (p-Akt and p-S6k) and increase in the apoptotic signaling (p-p38 MAPK) up to Day 7 post-AMI in 56Fe-IR-AMI mice. In summary, our results revealed that by 1 and 3 months post-IR in IR + AGING, 56Fe-IR but not 1H-IR mice had worse cardiac function. Further, a single 1H-IR 3 months prior to AMI improved, whereas 56Fe-IR worsened, recovery from AMI recovery. Our data in the IR + AGING and IR + AGING + AMI groups strongly suggest that low-dose HZE particle IR (56Fe) have long-lasting negative effect on heart homeostasis during normal aging, and present a significant CV risk for recovery after adverse CV event, such as AMI.

Mutagenic joining of enzymatically induced DNA double-strand breaks, accompanied by persistent unrepaired DNA damage and a secretory protein phenotype, in HZE-exposed human cells

High charge and energy (HZE) particles are a component of galactic cosmic rays. They cause complex damage to DNA and other cellular components, leading to both direct and indirect biological effects. Here, we investigate the hypothesis that one of these indirect effects is to compromise the accuracy of the DNA repair machinery, reducing the ability to cope with subsequent genotoxic insults. To this end, we used a new human reporter cell line with single-copy integrated fluorescent reporter cassettes that allow measurement of the frequency of mutagenic repair. Introduction of the rare cutting I-SceI nuclease stimulates both translocations (joining of two I-SceI sites on different chromosomes), and deletions (joining of two I-SceI sites after deletion of an intervening fragment) [ 1]. These can be measured simultaneously in the same cell population using different color reporter genes. To test the effect of HZE exposure on the frequency of translocations and deletions in this assay, cells were exposed to 600 MeV/u 56Fe ions or 1000 MeV/u 48Ti ions at doses of 0.3 or 1.0 Gy. They were allowed to recover and challenged with I-SceI at 1, 7, 14, 21 and 28 days post-irradiation. Results showed that HZE particle irradiation significantly increased the frequency of I-SceI translocations and deletions above baseline levels. There was an increase in translocations by up to threefold, seen in both 56Fe- and 48Ti-treated populations. There was also a more modest, but significant increase in I-SceI-mediated deletions seen in a population that received the higher dose (1.0 Gy) of 56Fe particles. The increased frequency of I-SceI-induced translocations and deletions persisted for 2–3 weeks with 56Fe (but not with 48Ti). The increased frequency of translocations and deletions was not observed in populations treated with low-LET radiation at doses up 3 Gy. Thus, the phenomenon, which we term the ‘mutagenic repair phenotype’ depends on both dose and radiation quality. The mutagenic repair phenotype was associated with an elevated frequency of micronuclei and excess DNA repair foci, suggesting that persistent genomic stress might be one causative factor [ 2].To further explore the mechanism underlying the mutagenic DNA repair phenotype, we performed genome-wide expression profiling on cells that were harvested 7 days post-56Fe ion exposure. Results showed significant alterations in 234 genes. Many of the most highly induced genes encode secreted proteins that have previously been implicated in cellular senescence or pro-inflammatory processes. These findings suggest that a paracrine mechanism (induction of a set of senescence or inflammation-related proteins) may also contribute to the mutagenic repair phenotype.Altered regulation of cellular double-strand break repair, with an increased reliance on an error-prone pathway, is a novel mechanism whereby an indirect effect of HZE exposure may amplify cancer risk.

Dose- and ApoE Isoform-Dependent Cognitive Injury after Cranial56Fe Irradiation in Female Mice

Apolipoprotein E (ApoE) plays an important role in lipid metabolism and neuronal repair. In humans, there are three major apoE isoforms: apoE2, apoE3 and apoE4. Compared to apoE3, apoE4 increases the risk to develop Alzheimer's disease, particularly in women, and of developing cognitive impairments after specific environmental challenges. ApoE isoform might also be a determinant of cognitive injury after cranial ⁵⁶Fe irradiation. To assess this possibility, in this study female apoE2, apoE3 and apoE4 mice were cranially irradiated with ⁵⁶Fe particles (600 MeV, 0, 1 or 2 Gy) and behaviorally tested 3 months later. Exploratory activity and measures of anxiety were also assessed as they can affect performance on cognitive tests. There were no effects of irradiation on exploratory activity or measures of anxiety in the open field or elevated zero maze. However, there were dose- and apoE isoform-dependent effects of irradiation on novel object recognition and spatial memory retention in the water maze. Compared to apoE2 and apoE3 mice, apoE4 mice were more sensitive to 2 Gy induced impairments in hippocampus-dependent spatial memory retention in the water maze after training to locate the first hidden platform location, but less sensitive to 2 Gy induced cortical impairments in novel object recognition. Conversely, of the irradiated mice, apoE4 mice irradiated with 1 Gy were the only group of mice that showed spatial memory retention for the second platform location after reversal learning in the water maze. Together, these data show that cranial ⁵⁶Fe irradiation causes dose- and apoE isoform-dependent cognitive impairments in female mice and that anatomical specificity might contribute to the relative sensitivity of apoE4 mice to develop space radiation-induced cognitive impairments.

Changes in Gene Expression in the Rat Hippocampus Following Exposure to 56Fe Particles and Protection by Berry Diets

Exposing young rats to particles of high energy and charge, such as (56)Fe, enhances indices of oxidative stress and inflammation and disrupts behavior, including spatial learning and memory. In the present study, we examined whether gene expression in the hippocampus, an area of the brain important in memory, is affected by exposure to 1.5 Gy or 2.5 Gy of 1 GeV/n high-energy (56)Fe particles 36 hours after irradiation. We also determined if 8 weeks of pre-feeding with 2% blueberry or 2% strawberry antioxidant diets could ameliorate irradiation-induced changes in gene expression. Alterations in gene expression profile were analyzed by pathway-focused microarrays for inflammatory cytokines and genes involved in nuclear factor-kappa B (NF-κB) signal transduction pathways. We found that genes that are directly or indirectly involved in the regulation of growth and differentiation of neurons were changed following irradiation. Genes that regulate apoptosis were up-regulated whereas genes that modulate cellular proliferation were down-regulated. The brains of animals supplemented with berry diets demonstrated an up-regulation of some protective stress signal genes. Therefore, these data suggest that (56)Fe particle irradiation causes changes in gene expression in rats that are ameliorated by berry fruit diets.