Immediate effects of acute Mars mission equivalent doses of SEP and GCR radiation on the murine gastrointestinal system-protective effects of curcumin-loaded nanolipoprotein particles (cNLPs).

Introduction: Missions beyond low Earth orbit (LEO) will expose astronauts to ionizing radiation (IR) in the form of solar energetic particles (SEP) and galactic cosmic rays (GCR) including high atomic number and energy (HZE) nuclei. The gastrointestinal (GI) system is documented to be highly radiosensitive with even relatively low dose IR exposures capable of inducing mucosal lesions and disrupting epithelial barrier function. IR is also an established risk factor for colorectal cancer (CRC) with several studies examining long-term GI effects of SEP/GCR exposure using tumor-prone APC mouse models. Studies of acute short-term effects of modeled space radiation exposures in wildtype mouse models are more limited and necessary to better define charged particle-induced GI pathologies and test novel medical countermeasures (MCMs) to promote astronaut safety. Methods: In this study, we performed ground-based studies where male and female C57BL/6J mice were exposed to γ-rays, 50 MeV protons, or 1 GeV/n Fe-56 ions at the NASA Space Radiation Laboratory (NSRL) with histology and immunohistochemistry endpoints measured in the first 24 h post-irradiation to define immediate SEP/GCR-induced GI alterations. Results: Our data show that unlike matched γ-ray controls, acute exposures to protons and iron ions disrupts intestinal function and induces mucosal lesions, vascular congestion, epithelial barrier breakdown, and marked enlargement of mucosa-associated lymphoid tissue. We also measured kinetics of DNA double-strand break (DSB) repair using gamma-H2AX- specific antibodies and apoptosis via TUNEL labeling, noting the induction and disappearance of extranuclear cytoplasmic DNA marked by gamma-H2AX only in the charged particle-irradiated samples. We show that 18 h pre-treatment with curcumin-loaded nanolipoprotein particles (cNLPs) delivered via IV injection reduces DSB-associated foci levels and apoptosis and restore crypt villi lengths. Discussion: These data improve our understanding of physiological alterations in the GI tract immediately following exposures to modeled space radiations and demonstrates effectiveness of a promising space radiation MCM.

Heavy Ion Minibeam Therapy: Side Effects in Normal Brain.

The purpose of this work was to investigate whether minibeam therapy with heavy ions might offer improvements of the therapeutic ratio for the treatment of human brain cancers. To assess neurotoxicity, we irradiated normal juvenile rats using 120 MeV lithium-7 ions at an absorbed integral dose of 20 Gy. Beams were configured either as a solid parallel circular beam or as an array of planar parallel minibeams having 300-micron width and 1-mm center-to-center spacing within a circular array. We followed animals for 6 months after treatment and utilized behavioral testing and immunohistochemical studies to investigate the resulting cognitive impairment and chronic pathologic changes. We found both solid-beam therapy and minibeam therapy to result in cognitive impairment compared with sham controls, with no apparent reduction in neurotoxicity using heavy ion minibeams instead of solid beams under the conditions of this study.

NASA's first ground-based Galactic Cosmic Ray Simulator: Enabling a new era in space radiobiology research.

With exciting new NASA plans for a sustainable return to the moon, astronauts will once again leave Earth's protective magnetosphere only to endure higher levels of radiation from galactic cosmic radiation (GCR) and the possibility of a large solar particle event (SPE). Gateway, lunar landers, and surface habitats will be designed to protect crew against SPEs with vehicle optimization, storm shelter concepts, and/or active dosimetry; however, the ever penetrating GCR will continue to pose the most significant health risks especially as lunar missions increase in duration and as NASA sets its aspirations on Mars. The primary risks of concern include carcinogenesis, central nervous system (CNS) effects resulting in potential in-mission cognitive or behavioral impairment and/or late neurological disorders, degenerative tissue effects including circulatory and heart disease, as well as potential immune system decrements impacting multiple aspects of crew health. Characterization and mitigation of these risks requires a significant reduction in the large biological uncertainties of chronic (low-dose rate) heavy-ion exposures and the validation of countermeasures in a relevant space environment. Historically, most research on understanding space radiation-induced health risks has been performed using acute exposures of monoenergetic single-ion beams. However, the space radiation environment consists of a wide variety of ion species over a broad energy range. Using the fast beam switching and controls systems technology recently developed at the NASA Space Radiation Laboratory (NSRL) at Brookhaven National Laboratory, a new era in radiobiological research is possible. NASA has developed the "GCR Simulator" to generate a spectrum of ion beams that approximates the primary and secondary GCR field experienced at human organ locations within a deep-space vehicle. The majority of the dose is delivered from protons (approximately 65%-75%) and helium ions (approximately 10%-20%) with heavier ions (Z ≥ 3) contributing the remainder. The GCR simulator exposes state-of-the art cellular and animal model systems to 33 sequential beams including 4 proton energies plus degrader, 4 helium energies plus degrader, and the 5 heavy ions of C, O, Si, Ti, and Fe. A polyethylene degrader system is used with the 100 MeV/n H and He beams to provide a nearly continuous distribution of low-energy particles. A 500 mGy exposure, delivering doses from each of the 33 beams, requires approximately 75 minutes. To more closely simulate the low-dose rates found in space, sequential field exposures can be divided into daily fractions over 2 to 6 weeks, with individual beam fractions as low as 0.1 to 0.2 mGy. In the large beam configuration (60 × 60 cm2), 54 special housing cages can accommodate 2 to 3 mice each for an approximately 75 min duration or 15 individually housed rats. On June 15, 2018, the NSRL made a significant achievement by completing the first operational run using the new GCR simulator. This paper discusses NASA's innovative technology solution for a ground-based GCR simulator at the NSRL to accelerate our understanding and mitigation of health risks faced by astronauts. Ultimately, the GCR simulator will require validation across multiple radiogenic risks, endpoints, doses, and dose rates.

Overview of the NASA space radiation laboratory.

The NASA Space Radiation Laboratory (NSRL) is a multidisciplinary center for space radiation research funded by NASA and located at the Brookhaven National Laboratory (BNL), Upton NY. Operational since 2003, the scope of NSRL is to provide ion beams in support of the NASA Humans in Space program in radiobiology, physics and engineering to measure the risk and ameliorate the effect of radiation in space. Recently, it has also been recognized as the only facility in the U.S. currently capable of contributing to heavy ion radiotherapy research. This work contains a general overview of NSRL structure, capabilities and operation.

Reference field specification and preliminary beam selection strategy for accelerator-based GCR simulation.

The galactic cosmic ray (GCR) simulator at the NASA Space Radiation Laboratory (NSRL) is intended to deliver the broad spectrum of particles and energies encountered in deep space to biological targets in a controlled laboratory setting. In this work, certain aspects of simulating the GCR environment in the laboratory are discussed. Reference field specification and beam selection strategies at NSRL are the main focus, but the analysis presented herein may be modified for other facilities and possible biological considerations. First, comparisons are made between direct simulation of the external, free space GCR field and simulation of the induced tissue field behind shielding. It is found that upper energy constraints at NSRL limit the ability to simulate the external, free space field directly (i.e. shielding placed in the beam line in front of a biological target and exposed to a free space spectrum). Second, variation in the induced tissue field associated with shielding configuration and solar activity is addressed. It is found that the observed variation is likely within the uncertainty associated with representing any GCR reference field with discrete ion beams in the laboratory, given current facility constraints. A single reference field for deep space missions is subsequently identified. Third, a preliminary approach for selecting beams at NSRL to simulate the designated reference field is presented. This approach is not a final design for the GCR simulator, but rather a single step within a broader design strategy. It is shown that the beam selection methodology is tied directly to the reference environment, allows facility constraints to be incorporated, and may be adjusted to account for additional constraints imposed by biological or animal care considerations. The major biology questions are not addressed herein but are discussed in a companion paper published in the present issue of this journal. Drawbacks of the proposed methodology are discussed and weighed against alternative simulation strategies.

Galactic cosmic ray simulation at the NASA Space Radiation Laboratory.

Most accelerator-based space radiation experiments have been performed with single ion beams at fixed energies. However, the space radiation environment consists of a wide variety of ion species with a continuous range of energies. Due to recent developments in beam switching technology implemented at the NASA Space Radiation Laboratory (NSRL) at Brookhaven National Laboratory (BNL), it is now possible to rapidly switch ion species and energies, allowing for the possibility to more realistically simulate the actual radiation environment found in space. The present paper discusses a variety of issues related to implementation of galactic cosmic ray (GCR) simulation at NSRL, especially for experiments in radiobiology. Advantages and disadvantages of different approaches to developing a GCR simulator are presented. In addition, issues common to both GCR simulation and single beam experiments are compared to issues unique to GCR simulation studies. A set of conclusions is presented as well as a discussion of the technical implementation of GCR simulation.

Charged Particle Therapy with Mini-Segmented Beams.

One of the fundamental attributes of proton therapy and carbon ion therapy is the ability of these charged particles to spare tissue distal to the targeted tumor. This significantly reduces normal tissue toxicity and has the potential to translate to a wider therapeutic index. Although, in general, particle therapy also reduces dose to the proximal tissues, particularly in the vicinity of the target, dose to the skin and to other very superficial tissues tends to be higher than that of megavoltage x-rays. The methods presented here, namely, "interleaved carbon minibeams" and "radiosurgery with arrays of proton and light ion minibeams," both utilize beams segmented into arrays of parallel "minibeams" of about 0.3 mm incident-beam size. These minibeam arrays spare tissues, as demonstrated by synchrotron x-ray experiments. An additional feature of particle minibeams is their gradual broadening due to multiple Coulomb scattering as they penetrate tissues. In the case of interleaved carbon minibeams, which do not broaden much, two arrays of planar carbon minibeams that remain parallel at target depth, are aimed at the target from 90° angles and made to "interleave" at the target to produce a solid radiation field within the target. As a result, the surrounding tissues are exposed only to individual carbon minibeam arrays and are therefore spared. The method was used in four-directional geometry at the NASA Space Radiation Laboratory to ablate a 6.5-mm target in a rabbit brain at a single exposure with 40 Gy physical absorbed dose. Contrast-enhanced magnetic resonance imaging and histology 6-month later showed very focal target necrosis with nearly no damage to the surrounding brain. As for minibeams of protons and light ions, for which the minibeam broadening is substantial, measurements at MD Anderson Cancer Center in Houston, TX, USA; and Monte Carlo simulations showed that the broadening minibeams will merge with their neighbors at a certain tissue depth to produce a solid beam to treat the target. The resulting sparing of proximal normal tissue allows radiosurgical ablative treatments with smaller impact on the skin and shallow tissues. This report describes these two methods and discusses their potential clinical applications.

Issues for Simulation of Galactic Cosmic Ray Exposures for Radiobiological Research at Ground-Based Accelerators.

For radiobiology research on the health risks of galactic cosmic rays (GCR) ground-based accelerators have been used with mono-energetic beams of single high charge, Z and energy, E (HZE) particles. In this paper, we consider the pros and cons of a GCR reference field at a particle accelerator. At the NASA Space Radiation Laboratory (NSRL), we have proposed a GCR simulator, which implements a new rapid switching mode and higher energy beam extraction to 1.5 GeV/u, in order to integrate multiple ions into a single simulation within hours or longer for chronic exposures. After considering the GCR environment and energy limitations of NSRL, we performed extensive simulation studies using the stochastic transport code, GERMcode (GCR Event Risk Model) to define a GCR reference field using 9 HZE particle beam-energy combinations each with a unique absorber thickness to provide fragmentation and 10 or more energies of proton and (4)He beams. The reference field is shown to well represent the charge dependence of GCR dose in several energy bins behind shielding compared to a simulated GCR environment. However, a more significant challenge for space radiobiology research is to consider chronic GCR exposure of up to 3 years in relation to simulations with animal models of human risks. We discuss issues in approaches to map important biological time scales in experimental models using ground-based simulation, with extended exposure of up to a few weeks using chronic or fractionation exposures. A kinetics model of HZE particle hit probabilities suggests that experimental simulations of several weeks will be needed to avoid high fluence rate artifacts, which places limitations on the experiments to be performed. Ultimately risk estimates are limited by theoretical understanding, and focus on improving knowledge of mechanisms and development of experimental models to improve this understanding should remain the highest priority for space radiobiology research.

Acute Hematological Effects in Mice Exposed to the Expected Doses, Dose-rates, and Energies of Solar Particle Event-like Proton Radiation.

NASA has funded several projects that have provided evidence for the radiation risk in space. One radiation concern arises from solar particle event (SPE) radiation, which is composed of energetic electrons, protons, alpha particles and heavier particles. SPEs are unpredictable and the accompanying SPE radiation can place astronauts at risk of blood cell death, contributing to a weakened immune system and increased susceptibility to infection. The doses, dose rates, and energies of the proton radiation expected to occur during a SPE have been simulated at the NASA Space Radiation Laboratory, Brookhaven National Laboratory, delivering total body doses to mice. Hematological values were evaluated at acute time points, up to 24 hrs. post-radiation exposure.

Interleaved carbon minibeams: an experimental radiosurgery method with clinical potential.

PURPOSE: To evaluate the efficacy of "interleaved carbon minibeams" for ablating a 6.5-mm target in a rabbit brain with little damage to the surrounding brain. The method is based on the well-established tissue-sparing effect of arrays of thin planes of radiation. METHODS AND MATERIALS: Broad carbon beams from the National Aeronautics and Space Agency Space Radiation Facility at Brookhaven National Laboratory were segmented into arrays of parallel, horizontal, 0.3-mm-thick planar beams (minibeams). The minibeams' gradual broadening in tissues resulted in 0.525-mm beam thickness at the target's proximal side in the spread-out Bragg peak. Interleaving was therefore implemented by choosing a 1.05 mm beam spacing on-center. The anesthetized rabbit, positioned vertically on a stage capable of rotating about a vertical axis, was exposed to arrays from four 90° angles, with the stage moving up by 0.525 mm in between. This produced a solid radiation field at the target while exposing the nontargeted tissues to single minibeam arrays. The target "physical" absorbed dose was 40.2 Gy. RESULTS: The rabbit behaved normally during the 6-month observation period. Contrast magnetic resonance imaging and hematoxylin and eosin histology at 6 months showed substantial focal target damage with little damage to the surrounding brain. CONCLUSION: We plan to evaluate the method's therapeutic efficacy by comparing it with broad-beam carbon therapy in animal models. The method's merits would combine those of carbon therapy (i.e., tight target dose because of the carbon's Bragg-peak, sharp dose falloff, and high relative biological effectiveness at the target), together with the method's low impact on the nontargeted tissues. The method's smaller impact on the nontargeted brain might allow carbon therapy at higher target doses and/or lower normal tissue impact, thus leading to a more effective treatment of radioresistant tumors. It should also make the method more amenable to administration in either a single dose fraction or in a small number of fractions.

Analyses of the secondary particle radiation and the DNA damage it causes to human keratinocytes.

High-energy protons, and high mass and energy ions, along with the secondary particles they produce, are the main contributors to the radiation hazard during space explorations. Skin, particularly the epidermis, consisting mainly of keratinocytes with potential for proliferation and malignant transformation, absorbs the majority of the radiation dose. Therefore, we used normal human keratinocytes to investigate and quantify the DNA damage caused by secondary radiation. Its manifestation depends on the presence of retinol in the serum-free media, and is regulated by phosphatidylinositol 3-kinases. We simulated the generation of secondary radiation after the impact of protons and iron ions on an aluminum shield. We also measured the intensity and the type of the resulting secondary particles at two sample locations; our findings agreed well with our predictions. We showed that secondary particles inflict DNA damage to different extents, depending on the type of primary radiation. Low-energy protons produce fewer secondary particles and cause less DNA damage than do high-energy protons. However, both generate fewer secondary particles and inflict less DNA damage than do high mass and energy ions. The majority of cells repaired the initial damage, as denoted by the presence of 53BPI foci, within the first 24 hours after exposure, but some cells maintained the 53BP1 foci longer.

Dose-rate effects of protons on in vivo activation of nuclear factor-kappa B and cytokines in mouse bone marrow cells.

The objective of this study was to determine the kinetics of nuclear factor-kappa B (NF-kappaB) activation and cytokine expression in bone marrow (BM) cells of exposed mice as a function of the dose rate of protons. The cytokines included in this study are pro-inflammatory [i.e., tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), and IL-6] and anti-inflammatory cytokines (i.e., IL-4 and IL-10). We gave male BALB/cJ mice a whole-body exposure to 0 (sham-controls) or 1.0 Gy of 100 MeV protons, delivered at 5 or 10 mGy min(-1), the dose and dose rates found during solar particle events in space. As a reference radiation, groups of mice were exposed to 0 (sham-controls) or 1 Gy of (137)Cs gamma rays (10 mGy min(-1)). After irradiation, BM cells were collected at 1.5, 3, 24 h, and 1 month for analyses (five mice per treatment group per harvest time). The results indicated that the in vivo time course of effects induced by a single dose of 1 Gy of 100 MeV protons or (137)Cs gamma rays, delivered at 10 mGy min(-1), was similar. Although statistically significant levels of NF-kappaB activation and pro-inflammatory cytokines in BM cells of exposed mice when compared to those in the corresponding sham controls (Student's t-test, p < 0.05 or <0.01) were induced by either dose rate, these levels varied over time for each protein. Further, only a dose rate of 5 mGy min(-1) induced significant levels of anti-inflammatory cytokines. The results indicate dose-rate effects of protons.

DNA damage intensity in fibroblasts in a 3-dimensional collagen matrix correlates with the Bragg curve energy distribution of a high LET particle.

PURPOSE: The DNA double-strand break (DSB) damage response induced by high energy charged particles on lung fibroblast cells embedded in a 3-dimensional (3-D) collagen tissue equivalents was investigated using antibodies to the DNA damage response proteins gamma-histone 2AX (gamma-H2AX) and phosphorylated DNA-PKcs (p-DNA-PKcs). MATERIALS AND METHODS: 3-D tissue equivalents were irradiated in positions across the linear distribution of the Bragg curve profiles of 307.7 MeV/nucleon, 556.9 MeV/nucleon, or 967.0 MeV/nucleon (56)Fe ions at a dose of 0.30 Gy. RESULTS: Patterns of discrete DNA damage streaks across nuclei or saturated nuclear damage were observed, with saturated nuclear damage being more predominant as samples were positioned closer to the physical Bragg peak. Quantification of the DNA damage signal intensities at each distance for each of the examined energies revealed a biological Bragg curve profile with a pattern of DNA damage intensity similar to the physical Bragg curve for the particular energy. Deconvolution microscopy of nuclei with streaked or saturated nuclear damage pattern revealed more details of the damage, with evidence of double-strand breaks radially distributed from the main particle track as well as multiple discrete tracks within saturated damage nuclei. CONCLUSIONS: These 3-D culture systems can be used as a biological substrate to better understand the interaction of heavy charged particles of different energies with tissue and could serve as a basis to model space-radiation-induced cancer initiation and progression.

Effects of estrogen and gender on cataractogenesis induced by high-LET radiation.

Planning for long-duration manned lunar and interplanetary missions requires an understanding of radiation-induced cataractogenesis. Previously, it was demonstrated that low-linear energy transfer (LET) irradiation with 10 Gy of (60)Co gamma rays resulted in an increased incidence of cataracts in male rats compared to female rats. This gender difference was not due to differences in estrogen, since male rats treated with the major secreted estrogen 17-beta-estradiol (E2) showed an identical increase compared to untreated males. We now compare the incidence and rate of progression of cataracts induced by high-LET radiation in male and female Sprague-Dawley rats. Rats received a single dose of 1 Gy of 600 MeV (56)Fe ions. Lens opacification was measured at 2-4-week intervals with a slit lamp. The incidence and rate of progression of radiation-induced cataracts was significantly increased in the animals in which estrogen was available from endogenous or exogenous sources. Male rats with E2 capsules implanted had significantly higher rates of progression compared to male rats with empty capsules implanted (P = 0.025) but not compared to the intact female rats. These results contrast with data obtained after low-LET irradiation and suggest the possibility that the different types of damage caused by high- and low-LET radiation may be influenced differentially by steroid sex hormones.