Geometrical Properties of the Nucleus and Chromosome Intermingling Are Possible Major Parameters of Chromosome Aberration Formation.

Ionizing radiation causes chromosome aberrations, which are possible biomarkers to assess space radiation cancer risks. Using the Monte Carlo codes Relativistic Ion Tracks (RITRACKS) and Radiation-Induced Tracks, Chromosome Aberrations, Repair and Damage (RITCARD), we investigated how geometrical properties of the cell nucleus, irradiated with ion beams of linear energy transfer (LET) ranging from 0.22 keV/µm to 195 keV/µm, influence the yield of simple and complex exchanges. We focused on the effect of (1) nuclear volume by considering spherical nuclei of varying radii; (2) nuclear shape by considering ellipsoidal nuclei of varying thicknesses; (3) beam orientation; and (4) chromosome intermingling by constraining or not constraining chromosomes in non-overlapping domains. In general, small nuclear volumes yield a higher number of complex exchanges, as compared to larger nuclear volumes, and a higher number of simple exchanges for LET < 40 keV/µm. Nuclear flattening reduces complex exchanges for high-LET beams when irradiated along the flattened axis. The beam orientation also affects yields for ellipsoidal nuclei. Reducing chromosome intermingling decreases both simple and complex exchanges. Our results suggest that the beam orientation, the geometry of the cell nucleus, and the organization of the chromosomes within are important parameters for the formation of aberrations that must be considered to model and translate in vitro results to in vivo risks.

Cancer incidence and mortality in the USA Astronaut Corps, 1959-2017.

OBJECTIVES: Cancer incidence and mortality are important outcomes in the surveillance of long-term astronaut health. We compare cancer incidence rates, cancer-specific mortality rates, and cancer case-fatality ratios in US astronauts with those in the US general population. METHODS: We use standardised incidence ratios (SIRs) and standardised mortality ratios (SMRs) to index the incidence and mortality of various cancers against rates in the US general population, from the US astronaut cohort inception in April 1959 through 31 December 2017. We compare the lethality of these cancers using the relative case-fatality ratio. RESULTS: Overall cancer incidence and mortality were slightly lower than expected from national rates with SIR 82 (95% CI 63 to 104) and SMR 72 (95% CI 44 to 111) with a modest 14% reduction in case-fatality ratio. Prostate cancer and melanoma skin cancer had significant increases in incidence, with SIR of 162 (95% CI 109 to 232) and 252 (95% CI 126 to 452), respectively, though only melanoma had a significant increase in mortality, with SMR 508 (95% CI 105 to 1485). Lung cancer had a significant deficit of both cases and deaths, while colon cancer had sizeable (but not significant) reductions in incidence and mortality. CONCLUSIONS: The increase in incidence of melanoma is consistent with that observed in aircraft pilots, suggesting this may be associated with ultraviolet radiation or lifestyle factors rather than any astronaut-specific exposure. Reductions in lung cancer incidence and mortality, and trends towards such reductions in colon cancer, may be explained in part by healthy lifestyle, as well as differential screening among astronauts.

Considering clonal hematopoiesis of indeterminate potential in space radiation risk analysis for hematologic cancers and cardiovascular disease.

BACKGROUND: Expanding human presence in space through long-duration exploration missions and commercial space operations warrants improvements in approaches for quantifying crew space radiation health risks. Currently, risk assessment models for radiogenic cancer and cardiovascular disease consider age, sex, and tobacco use, but do not incorporate other modifiable (e.g., body weight, physical activity, diet, environment) and non-modifiable individual risk factors (e.g., genetics, medical history, race/ethnicity, family history) that may greatly influence crew health both in-mission and long-term. For example, clonal hematopoiesis of indeterminate potential (CHIP) is a relatively common age-related condition that is an emerging risk factor for a variety of diseases including cardiovascular disease and cancer. CHIP carrier status may therefore exacerbate health risks associated with space radiation exposure. METHODS: In the present study, published CHIP hazard ratios were used to modify background hazard rates for coronary heart disease, stroke, and hematologic cancers in the National Aeronautics and Space Administration space radiation risk assessment model. The risk of radiation exposure-induced death for these endpoints was projected for a future Mars exploration mission scenario. RESULTS: Here we show appreciable increases in the lifetime risk of exposure-induced death for hematologic malignancies, coronary heart disease, and stroke, which are observed as a function of age after radiation exposure for male and female crew members that are directly attributable to the elevated health risks for CHIP carriers. CONCLUSIONS: We discuss the importance of evaluating individual risk factors such as CHIP as part of a comprehensive space radiation risk assessment strategy aimed at effective risk communication and disease surveillance for astronauts embarking on future exploration missions.

Space radiation exposure is a major hazard of spaceflight that may increase cancer and cardiovascular disease risks for future astronauts exploring the moon and Mars. There is a need for accurate risk assessment that considers individual risk factors to support informed consent and medical management of these risks. Clonal hematopoiesis of indeterminate potential (CHIP) is a condition that occurs when copies of variant cells accumulate in the blood of otherwise healthy individuals. CHIP is an emerging risk factor linked with blood cancers and cardiovascular disease. We evaluated how CHIP can alter space radiation health risks in astronauts for a Mars exploration mission scenario. We find large increases in lifetime risk of space radiation exposure-induced death for hematologic malignancies and cardiovascular disease in CHIP carriers. These results suggest that increased screening may help facilitate better management of radiation risks.

Galactic cosmic ray simulation at the NASA space radiation laboratory - Progress, challenges and recommendations on mixed-field effects.

For missions beyond low Earth orbit to the moon or Mars, space explorers will encounter a complex radiation field composed of various ion species with a broad range of energies. Such missions pose significant radiation protection challenges that need to be solved in order to minimize exposures and associated health risks. An innovative galactic cosmic ray simulator (GCRsim) was recently developed at the NASA Space Radiation Laboratory (NSRL) at Brookhaven National Laboratory (BNL). The GCRsim technology is intended to represent major components of the space radiation environment in a ground analog laboratory setting where it can be used to improve understanding of biological risks and serve as a testbed for countermeasure development and validation. The current GCRsim consists of 33 energetic ion beams that collectively simulate the primary and secondary GCR field encountered by humans in space over the broad range of particle types, energies, and linear energy transfer (LET) of interest to health effects. A virtual workshop was held in December 2020 to assess the status of the NASA baseline GCRsim. Workshop attendees examined various aspects of simulator design, with a particular emphasis on beam selection strategies. Experimental results, modeling approaches, areas of consensus, and questions of concern were also discussed in detail. This report includes a summary of the GCRsim workshop and a description of the current status of the GCRsim. This information is important for future advancements and applications in space radiobiology.

Medical Countermeasure Requirements to Meet NASA's Space Radiation Permissible Exposure Limits for a Mars Mission Scenario.

ABSTRACT: The space radiation environment consists of a complex mixture of ionizing particles that pose significant health risks to crew members. NASA currently requires that an astronaut's career Risk of Exposure Induced Death (REID) for cancer mortality should not exceed 3% at the upper 95% confidence level. This career radiation limit is likely to be exceeded for even the shortest round-trip mission scenario to Mars. As such, NASA has begun to pursue more vigorously approaches to directly reduce radiation risks, despite the large uncertainties associated with such projections. A recent study considered cohort studies of aspirin and warfarin as possible medical countermeasures (MCMs) acting to reduce background cancer mortality rates used in astronaut risk projections. It was shown that such MCMs can reduce the REID for specific tissues in restricted time intervals over which the drugs were administered; however, the cumulative effect on total lifetime REID was minimal. As an extension, the present work addresses more general MCM requirements that would be needed to meet current NASA radiation limits for a Mars mission scenario. A sensitivity analysis is performed within the major components of the NASA cancer risk model that would likely be modified by MCM interventions. This includes the background cancer incidence and mortality rates, epidemiologically based hazard rates derived from acute terrestrial exposures, and radiation quality factors used to translate terrestrial exposures to space radiation. Relationships between possible MCMs and each of these components are discussed. Results from this study provide important information regarding MCM requirements needed to meet NASA limits for planned Mars missions. Insight into the types of countermeasures expected to yield greatest reductions in crew risk is also gained.

Cardiovascular Disease Risk Modeling for Astronauts: Making the Leap From Earth to Space.

NASA has recently completed several long-duration missions to the International Space Station and is solidifying plans to return to the Moon, with an eye toward Mars and beyond. As NASA pushes the boundaries of human space exploration, the hazards of spaceflight, including space radiation, levy an increasing burden on astronaut health and performance. The cardiovascular system may be especially vulnerable due to the combined impacts of space radiation exposure, lack of gravity, and other spaceflight hazards. On Earth, the risk for cardiovascular disease (CVD) following moderate to high radiation doses is well-established from clinical, environmental, and occupational exposures (largely from gamma- and x-rays). Less is known about CVD risks associated with high-energy charged ions found in space and increasingly used in radiotherapy applications on Earth, making this a critical area of investigation for occupational radiation protection. Assessing CVD risk is complicated by its multifactorial nature, where an individual's risk is strongly influenced by factors such as family history, blood pressure, and lipid profiles. These known risk factors provide the basis for development of a variety of clinical risk prediction models (CPMs) that inform the likelihood of medical outcomes over a defined period. These tools improve clinical decision-making, personalize care, and support primary prevention of CVD. They may also be useful for individualizing risk estimates for CVD following radiation exposure both in the clinic and in space. In this review, we summarize unique aspects of radiation risk assessment for astronauts, and we evaluate the most widely used CVD CPMs for their use in NASA radiation risk assessment applications. We describe a comprehensive dual-use risk assessment framework that supports both clinical care and operational management of space radiation health risks using quantitative metrics. This approach is a first step in using personalized medicine for radiation risk assessment to support safe and productive spaceflight and long-term quality of life for NASA astronauts.

Ionizing radiation, cerebrovascular disease, and consequent dementia: A review and proposed framework relevant to space radiation exposure.

Space exploration requires the characterization and management or mitigation of a variety of human health risks. Exposure to space radiation is one of the main health concerns because it has the potential to increase the risk of cancer, cardiovascular disease, and both acute and late neurodegeneration. Space radiation-induced decrements to the vascular system may impact the risk for cerebrovascular disease and consequent dementia. These risks may be independent or synergistic with direct damage to central nervous system tissues. The purpose of this work is to review epidemiological and experimental data regarding the impact of low-to-moderate dose ionizing radiation on the central nervous system and the cerebrovascular system. A proposed framework outlines how space radiation-induced effects on the vasculature may increase risk for both cerebrovascular dysfunction and neural and cognitive adverse outcomes. The results of this work suggest that there are multiple processes by which ionizing radiation exposure may impact cerebrovascular function including increases in oxidative stress, neuroinflammation, endothelial cell dysfunction, arterial stiffening, atherosclerosis, and cerebral amyloid angiopathy. Cerebrovascular adverse outcomes may also promote neural and cognitive adverse outcomes. However, there are many gaps in both the human and preclinical evidence base regarding the long-term impact of ionizing radiation exposure on brain health due to heterogeneity in both exposures and outcomes. The unique composition of the space radiation environment makes the translation of the evidence base from terrestrial exposures to space exposures difficult. Additional investigation and understanding of the impact of low-to-moderate doses of ionizing radiation including high (H) atomic number (Z) and energy (E) (HZE) ions on the cerebrovascular system is needed. Furthermore, investigation of how decrements in vascular systems may contribute to development of neurodegenerative diseases in independent or synergistic pathways is important for protecting the long-term health of astronauts.

AT cells are not radiosensitive for simple chromosomal exchanges at low dose.

Cells deficient in ATM (product of the gene that is mutated in ataxia telangiectasia patients) or NBS (product of the gene mutated in the Nijmegen breakage syndrome) show increased yields of both simple and complex chromosomal aberrations after high doses (>0.5Gy) of ionizing radiation (X-rays or γ-rays), however less is known on how these cells respond at low dose. Previously we had shown that the increased chromosome aberrations in ATM and NBS defective lines was due to a significantly larger quadratic dose-response term compared to normal fibroblasts for both simple and complex exchanges. The linear dose-response term for simple exchanges was significantly higher in NBS cells compared to wild type cells, but not for AT cells. However, AT cells have a high background level of exchanges compared to wild type or NBS cells that confounds the understanding of low dose responses. To understand the sensitivity differences for high to low doses, chromosomal aberration analysis was first performed at low dose-rates (0.5Gy/d), and results provided further evidence for the lack of sensitivity for exchanges in AT cells below doses of 1Gy. Normal lung fibroblast cells treated with KU-55933, a specific ATM kinase inhibitor, showed increased numbers of exchanges at a dose of 1Gy and higher, but were similar to wild type cells at 0.5Gy or below. These results were confirmed using siRNA knockdown of ATM. The present study provides evidence that the increased radiation sensitivity of AT cells for chromosomal exchanges found at high dose does not occur at low dose.

Earth-Based Research Analogs to Investigate Space-Based Health Risks.

During spaceflight, astronauts are exposed to a variety of unique hazards, including altered gravity fields, long periods of isolation and confinement, living in a closed environment at increasing distances from Earth, and exposure to higher levels of hazardous ionizing radiation. Preserving human health and performance in the face of these relentless hazards becomes progressively more difficult as missions increase in length and extend beyond low Earth orbit. Finding solutions is a significant challenge that is further complicated by logistical issues associated with studying these unique hazards. Although research studies using space-based platforms are the gold standard, these are not without limitations. Factors such as the small sample size of the available astronaut crew, high expense, and time constraints all add to the logistical challenge. To overcome these limitations, a wide variety of Earth-based analogs, from polar research outposts to an undersea laboratory, are available to augment space-based studies. Each analog simulates unique physiological and behavioral effects associated with spaceflight and, therefore, for any given study, the choice of an appropriate platform is closely linked to the phenomena under investigation as well as the characteristics of the analog. There are pros and cons to each type of analog and each actual facility, but overall they provide a reasonable means to overcome the barriers associated with conducting experimental research in space. Analogs, by definition, will never be perfect, but they are a useful component of an integrated effort to understand the human risks of living and working in space. They are a necessary resource for pushing the frontier of human spaceflight, both for astronauts and for commercial space activities. In this review, we describe the use of analogs here on Earth to replicate specific aspects of the spaceflight environment and highlight how analog studies support future human endeavors in space.

Expert consultation is vital for adverse outcome pathway development: a case example of cardiovascular effects of ionizing radiation.

BACKGROUND: The circulatory system distributes nutrients, signaling molecules, and immune cells to vital organs and soft tissues. Epidemiological, animal, and in vitro cellular mechanistic studies have highlighted that exposure to ionizing radiation (IR) can induce molecular changes in cellular and subcellular milieus leading to long-term health impacts, particularly on the circulatory system. Although the mechanisms for the pathologies are not fully elucidated, endothelial dysfunction is proven to be a critical event via radiation-induced oxidative stress mediators. To delineate connectivities of events specifically to cardiovascular disease (CVD) initiation and progression, the adverse outcome pathway (AOP) approach was used with consultation from field experts. AOPs are a means to organize information around a disease of interest to a regulatory question. An AOP begins with a molecular initiating event and ends in an adverse outcome via sequential linkages of key event relationships that are supported by evidence in the form of the modified Bradford-Hill criteria. Detailed guidelines on building AOPs are provided by the Organisation for Economic Cooperation and Development (OECD) AOP program. Here, we report on the questions and discussions needed to develop an AOP for CVD resulting from IR exposure. A recent workshop jointly organized by the MELODI (Multidisciplinary European Low Dose Initiative) and the ALLIANCE (European Radioecology Alliance) associations brought together experts from the OECD to present the AOP approach and tools with examples from the toxicology field. As part of this workshop, four working groups were formed to discuss the identification of adverse outcomes relevant to radiation exposures and development of potential AOPs, one of which was focused on IR-induced cardiovascular effects. Each working group comprised subject matter experts and radiation researchers interested in the specific disease area and included an AOP coach. CONCLUSION: The CVD working group identified the critical questions of interest for AOP development, including the exposure scenario that would inform the evidence, the mechanisms of toxicity, the initiating event, intermediate key events/relationships, and the type of data currently available. This commentary describes the four-day discussion of the CVD working group, its outcomes, and demonstrates how collaboration and expert consultation is vital to informing AOP construction.

Nucleus-mitochondria positive feedback loop formed by ERK5 S496 phosphorylation-mediated poly (ADP-ribose) polymerase activation provokes persistent pro-inflammatory senescent phenotype and accelerates coronary atherosclerosis after chemo-radiation.

The incidence of cardiovascular disease (CVD) is higher in cancer survivors than in the general population. Several cancer treatments are recognized as risk factors for CVD, but specific therapies are unavailable. Many cancer treatments activate shared signaling events, which reprogram myeloid cells (MCs) towards persistent senescence-associated secretory phenotype (SASP) and consequently CVD, but the exact mechanisms remain unclear. This study aimed to provide mechanistic insights and potential treatments by investigating how chemo-radiation can induce persistent SASP. We generated ERK5 S496A knock-in mice and determined SASP in myeloid cells (MCs) by evaluating their efferocytotic ability, antioxidation-related molecule expression, telomere length, and inflammatory gene expression. Candidate SASP inducers were identified by high-throughput screening, using the ERK5 transcriptional activity reporter cell system. Various chemotherapy agents and ionizing radiation (IR) up-regulated p90RSK-mediated ERK5 S496 phosphorylation. Doxorubicin and IR caused metabolic changes with nicotinamide adenine dinucleotide depletion and ensuing mitochondrial stunning (reversible mitochondria dysfunction without showing any cell death under ATP depletion) via p90RSK-ERK5 modulation and poly (ADP-ribose) polymerase (PARP) activation, which formed a nucleus-mitochondria positive feedback loop. This feedback loop reprogramed MCs to induce a sustained SASP state, and ultimately primed MCs to be more sensitive to reactive oxygen species. This priming was also detected in circulating monocytes from cancer patients after IR. When PARP activity was transiently inhibited at the time of IR, mitochondrial stunning, priming, macrophage infiltration, and coronary atherosclerosis were all eradicated. The p90RSK-ERK5 module plays a crucial role in SASP-mediated mitochondrial stunning via regulating PARP activation. Our data show for the first time that the nucleus-mitochondria positive feedback loop formed by p90RSK-ERK5 S496 phosphorylation-mediated PARP activation plays a crucial role of persistent SASP state, and also provide preclinical evidence supporting that transient inhibition of PARP activation only at the time of radiation therapy can prevent future CVD in cancer survivors.

Determination of Chromosome Aberrations in Human Fibroblasts Irradiated by Mixed Fields Generated with Shielding.

To better study biological effects of space radiation using ground-based facilities, the NASA Space Radiation Laboratory (NSRL) at the Brookhaven National Laboratory has been upgraded to rapidly switch ions and energies. This has allowed investigators to design irradiation protocols comprising a mixture of ions and energies more indicative of the galactic cosmic ray (GCR) environment. Despite these advancements, beam selection and delivery schemes should be optimized against facility and experimental constraints and validated to ensure such irradiations are a suitable representation of the space environment. Importantly, since experiments are time consuming and expensive, models capable of predicting biological outcomes over a range of irradiation conditions (single ion, sequential multi ion or mixed fields) are needed to support such efforts. In this work, human fibroblasts were placed behind 20 g/cm2 aluminum and 10.345 g/cm2 polyethylene and irradiated separately by 344 MeV hydrogen, 344 MeV/n helium, 450 MeV/n oxygen and 950 MeV/n iron ions at various doses. The fluorescence in situ hybridization (FISH) whole chromosome painting technique was then used to assess the cells for chromosome aberrations (CAs), notably simple exchanges. A multi-scale modeling approach was also developed to predict the formation of chromosome aberrations in these experiments. The Geant4 simulation toolkit was used to determine the spectra of particles and energies produced by interactions between the incident beams and shielding. The simulated mixed field generated by shielding was then transferred into the track structure code, RITRACKS (relativistic ion tracks), to generate three-dimensional (3D) voxelized dose maps at the nanometer scale. Finally, these voxel dose maps were input into the new damage and repair model, RITCARD (radiation-induced tracks, chromosome aberrations, repair and damage), to predict the formation of various CAs. The multi-scale model described herein is a significant advancement for the computational tools used to predict biological outcomes in cells exposed to highly complex, mixed ion fields related to the GCR environment. Results show that the simulation and experimental data are in good agreement for the complex radiation fields generated by all ions incident on shielding for most data points. The differences between model predictions and measurements are discussed. Although improvements are needed, the model extends current capabilities for evaluating beam selection and delivery schemes at the NSRL ground-based GCR simulator and for informing NASA risk projection models in the future.

Red risks for a journey to the red planet: The highest priority human health risks for a mission to Mars.

NASA's plans for space exploration include a return to the Moon to stay-boots back on the lunar surface with an orbital outpost. This station will be a launch point for voyages to destinations further away in our solar system, including journeys to the red planet Mars. To ensure success of these missions, health and performance risks associated with the unique hazards of spaceflight must be adequately controlled. These hazards-space radiation, altered gravity fields, isolation and confinement, closed environments, and distance from Earth-are linked with over 30 human health risks as documented by NASA's Human Research Program. The programmatic goal is to develop the tools and technologies to adequately mitigate, control, or accept these risks. The risks ranked as "red" have the highest priority based on both the likelihood of occurrence and the severity of their impact on human health, performance in mission, and long-term quality of life. These include: (1) space radiation health effects of cancer, cardiovascular disease, and cognitive decrements (2) Spaceflight-Associated Neuro-ocular Syndrome (3) behavioral health and performance decrements, and (4) inadequate food and nutrition. Evaluation of the hazards and risks in terms of the space exposome-the total sum of spaceflight and lifetime exposures and how they relate to genetics and determine the whole-body outcome-will provide a comprehensive picture of risk profiles for individual astronauts. In this review, we provide a primer on these "red" risks for the research community. The aim is to inform the development of studies and projects with high potential for generating both new knowledge and technologies to assist with mitigating multisystem risks to crew health during exploratory missions.

Coronary Artery Dose-Volume Parameters Predict Risk of Calcification After Radiation Therapy.

BACKGROUND: Radiation exposure increases the risk of coronary artery disease (CAD). We explored the association of CAD with coronary artery dose-volume parameters in patients treated with 3D-planned radiation therapy (RT). METHODS: Patients who received thoracic RT and were evaluated by cardiac computed tomography ≥ 1 year later were included. Demographic data and cardiac risk factors were retrospectively collected. Dosimetric data (mean heart dose, dmax, dmean, V50 - V5) were collected for the whole heart and for each coronary artery. A coronary artery calcium (CAC) Agatston score was calculated on a per-coronary basis and as a total score. Multivariable generalized linear mixed models were generated. The predicted probabilities were used for receiver operating characteristic analyses. RESULTS: Twenty patients with a median age of 53 years at the time of RT were included. Nine patients (45%) had ≥ 3/6 conventional cardiac risk factors. Patients received RT for breast cancer (10, 50%), lung cancer (6, 30%), or lymphoma/myeloma (4, 20%) with a median dose of 60 Gy. CAC scans were performed a median of 32 months after RT. CAC score was significantly associated with radiation dose and presence of diabetes. In a multivariable model adjusted for diabetes, segmental coronary artery dosimetric parameters (dmax, dmean, V50, V40 V30, V20, V10, and V5) were significantly associated with CAC score > 0. V50 had the highest area under the ROC curve (0.89, 95% confidence interval, 0.80-0.97). CONCLUSIONS: Coronary artery radiation exposure is strongly correlated with subsequent segmental CAC score. Coronary calcification may occur soon after RT and in individuals with conventional cardiac risk factors.

Dual delivery of an angiogenic and an osteogenic growth factor for bone regeneration in a critical size defect model.

This study investigated the effects of dual delivery of vascular endothelial growth factor (VEGF) and bone morphogenetic protein-2 (BMP-2) for bone regeneration in a rat cranial critical size defect. Four groups of scaffolds were generated with VEGF (12 microg), BMP-2 (2 mug), both VEGF (12 microg) and BMP-2 (2 microg), or no growth factor released from gelatin microparticles incorporated within the scaffold pores. These scaffolds were implanted within an 8 mm rat cranial critical size defect (n=8-9 for each group). At 4 and 12 weeks, implants were retrieved and evaluated by microcomputed tomography (microCT) and histological scoring analysis. Additionally, 4 week animals were perfused with a radiopaque material to visualize and quantify blood vessel formation. Histological analysis revealed that for all groups at 4 weeks, a majority of the porous scaffold volume was filled with vascularized fibrous tissue; however, bone formation appeared most abundant in the dual release group at this time. At 12 weeks, both dual release and BMP-2 groups showed large amounts of bone formation within the scaffold pores and along the outer surfaces of the scaffold; osteoid secretion and mineralization were apparent, and new bone was often in close or direct contact with the scaffold interface. MicroCT results showed no significant difference among groups for blood vessel formation at 4 weeks (<4% blood vessel volume); however, the dual release group showed significantly higher bone formation (16.1+/-9.2% bone volume) than other groups at this time. At 12 weeks, dual release and BMP-2 groups exhibited significantly higher bone formation (39.7+/-14.1% and 37.4+/-18.8% bone volume, respectively) than either the VEGF group or blank scaffolds (6.3+/-4.8% and 7.8+/-7.1% bone volume, respectively). This work indicates a synergistic effect of the dual delivery of VEGF and BMP-2 on bone formation at 4 weeks and suggests an interplay between these growth factors for early bone regeneration. For the doses investigated, the results show that the addition of VEGF does not affect the amount of bone formation achieved by BMP-2 at 12 weeks; however, they also indicate that delivery of both growth factors may enhance bone bridging and union of the critical size defect compared to delivery of BMP-2 alone.

Biodegradable gelatin microparticles as delivery systems for the controlled release of bone morphogenetic protein-2.

This work evaluated gelatin microparticles and biodegradable composite scaffolds for the controlled release of bone morphogenetic protein-2 (BMP-2) in vitro and in vivo. Gelatin crosslinking (10 and 40mM glutaraldehyde), BMP-2 dose (6 and 60ng BMP-2 per mg dry microparticles), buffer type (phosphate buffered saline (PBS) and collagenase-containing PBS), and gelatin type (acidic and basic) were investigated for their effects on BMP-2 release. Release profiles were also observed using poly(lactic-co-glycolic acid) (PLGA) microparticles with varying molecular weights (8300 and 57,500). In vitro and in vivo studies were conducted using radiolabeled BMP-2; the chloramine-T method was preferred over Bolton-Hunter reagent for radioiodination with this system. BMP-2 release from PLGA microparticles resulted in a moderate burst release followed by minimal cumulative release, while BMP-2 release from gelatin microparticles exhibited minimal burst release followed by linear release kinetics in vitro. Growth factor dose had a small effect on its normalized release kinetics probably because of an equilibrium between gelatin-bound and unbound BMP-2. Differences in release from acidic and basic gelatin microparticles may result from the different pretreatment conditions used for gelatin synthesis. The in vitro release kinetics for both gelatin microparticles alone and within composite scaffolds were dependent largely on the extent of gelatin crosslinking; varying buffer type served to confirm that controlled release relies on enzymatic degradation of the gelatin for controlled release. Finally, in vivo studies with composite scaffolds exhibited minimal burst and linear release up to 28 days. In summary, dose effects on BMP-2 release were found to be minimal while varying gelatin type and release medium can alter release kinetics. These results demonstrate that a systematic control of BMP-2 delivery from gelatin microparticles can be achieved by altering the extent of basic gelatin crosslinking.

Radiation-Induced Cardiovascular Disease: Mechanisms and Importance of Linear Energy Transfer.

Radiation therapy (RT) in the form of photons and protons is a well-established treatment for cancer. More recently, heavy charged particles have been used to treat radioresistant and high-risk cancers. Radiation treatment is known to cause cardiovascular disease (CVD) which can occur acutely during treatment or years afterward in the form of accelerated atherosclerosis. Radiation-induced cardiovascular disease (RICVD) can be a limiting factor in treatment as well as a cause of morbidity and mortality in successfully treated patients. Inflammation plays a key role in both acute and chronic RICVD, but the underling pathophysiology is complex, involving DNA damage, reactive oxygen species, and chronic inflammation. While understanding of the molecular mechanisms of RICVD has increased, the growing number of patients receiving RT warrants further research to identify individuals at risk, plans for prevention, and targets for the treatment of RICVD. Research on RICVD is also relevant to the National Aeronautics and Space Administration (NASA) due to the prevalent space radiation environment encountered by astronauts. NASA's current research on RICVD can both contribute to and benefit from concurrent work with cell and animal studies informing radiotoxicities resulting from cancer therapy. This review summarizes the types of radiation currently in clinical use, models of RICVD, current knowledge of the mechanisms by which they cause CVD, and how this knowledge might apply to those exposed to various types of radiation.

Radiation Exposure and Mortality from Cardiovascular Disease and Cancer in Early NASA Astronauts.

Understanding space radiation health effects is critical due to potential increased morbidity and mortality following spaceflight. We evaluated whether there is evidence for excess cardiovascular disease or cancer mortality in early NASA astronauts and if a correlation exists between space radiation exposure and mortality. Astronauts selected from 1959-1969 were included and followed until death or February 2017, with 39 of 73 individuals still alive at that time. Calculated standardized mortality rates for tested outcomes were significantly below U.S. white male population rates, including all-cardiovascular disease (n = 7, SMR = 33; 95% CI, 14-65) and all-cancer (n = 7, SMR = 43; 95% CI, 18-83), as anticipated in a healthy worker population. Space radiation doses for cohort members ranged from 0-78 mGy. No significant associations between space radiation dose and mortality were found using logistic regression with an internal reference group, adjusting for medical radiation. Statistical power of the logistic regression was <6%, remaining <12% even when expected risk level or observed deaths were assumed to be 10 times higher than currently reported. While no excess radiation-associated cardiovascular or cancer mortality risk was observed, findings must be tempered by the statistical limitations of this cohort; notwithstanding, this small unique cohort provides a foundation for assessment of astronaut health.

Developing a Reliable Mouse Model for Cancer Therapy-Induced Cardiovascular Toxicity in Cancer Patients and Survivors.

BACKGROUND: The high incidence of cardiovascular events in cancer survivors has long been noted, but the mechanistic insights of cardiovascular toxicity of cancer treatments, especially for vessel diseases, remain unclear. It is well known that atherosclerotic plaque formation begins in the area exposed to disturbed blood flow, but the relationship between cancer therapy and disturbed flow in regulating plaque formation has not been well studied. Therefore, we had two goals for this study; (1) Generate an affordable, reliable, and reproducible mouse model to recapitulate the cancer therapy-induced cardiovascular events in cancer survivors, and (2) Establish a mouse model to investigate the interplay between disturbed flow and various cancer therapies in the process of atherosclerotic plaque formation. METHODS AND RESULTS: We examined the effects of two cancer drugs and ionizing radiation (IR) on disturbed blood flow-induced plaque formation using a mouse carotid artery partial ligation (PCL) model of atherosclerosis. We found that doxorubicin and cisplatin, which are commonly used anti-cancer drugs, had no effect on plaque formation in partially ligated carotid arteries. Similarly, PCL-induced plaque formation was not affected in mice that received IR (2 Gy) and PCL surgery performed one week later. In contrast, when PCL surgery was performed 26 days after IR treatment, not only the atherosclerotic plaque formation but also the necrotic core formation was significantly enhanced. Lastly, we found a significant increase in p90RSK phosphorylation in the plaques from the IR-treated group compared to those from the non-IR treated group. CONCLUSIONS: Our results demonstrate that IR not only increases atherosclerotic events but also vulnerable plaque formation. These increases were a somewhat delayed effect of IR as they were observed in mice with PCL surgery performed 26 days, but not 10 days, after IR exposure. A proper animal model must be developed to study how to minimize the cardiovascular toxicity due to cancer treatment.

Enhanced catalytic efficiency of aminoglycoside phosphotransferase (3')-IIa achieved through protein fragmentation and reassembly.

Many monomeric proteins can be split into two fragments, yet the two fragments can associate to make an active heterodimer. However, for most locations in a protein such a conversion is not feasible, presumably due to inefficient assembly or improper folding of the fragments. For some locations, this can be overcome by fusion of the fragments to dimerization domains that facilitate correct assembly. A variety of heterodimers of aminoglycoside phosphotransferase (3')-IIa (Neo) were created in which the Neo fragments required fusion to a pair of leucine zippers for activity in vivo. However, the ability of these heterodimers to confer kanamycin resistance to Escherichia coli cells was impaired compared to wild-type Neo, primarily due to poor production of soluble protein. The mutations R177S and V198E restored the kanamycin resistance to wild-type levels while maintaining the dependence on leucine zippers for activity. These mutations restored high levels of kanamycin resistance not through an improvement in the production of soluble protein but rather by conferring a large improvement in k(cat)/K(m), surpassing that of Neo. Furthermore, whereas R177S and V198E served to improve k(cat)/K(m) 60-fold in the context of the heterodimer, the same mutations in the context of wild-type Neo had a ninefold negative effect on k(cat)/K(m). This demonstrates the possibility that enzymes with improved catalytic properties can be created through a process involving fragmentation and fusion to domains that facilitate assembly of the fragments.