ASystematicReviewoftheHematopoieticAcuteRadiationSyndrome(H-ARS)inCaninesandNon-humanPrimates:AcuteMixedNeutron/Gammavs.ReferenceQualityRadiations

Thomas J. MacVittie1, Ann M. Farese1, William E. Jackson III2

(1. University of Maryland School of Medicine, Baltimore, MD;2. Statistician, Rockville, MD)

Abstract:A systematic review of relevant studies that determined the dose response relationship (DRR) for the hematopoietic (H) acute radiation syndrome (ARS) in the canine relative to radiation quality of mixed neutron:gamma radiations, dose rate, and exposure uniformity relative to selected reference radiation exposure has not been performed. The datasets for rhesus macaques exposure to mixed neutron:gamma radiation are used herein as a species comparative reference to the canine database. The selection of data cohorts was made from the following sources: Ovid Medline (1957-present), PubMed (1954-present), AGRICOLA (1976-present), Web of Science (1954-present), and US HHS RePORT (2002-present). The total number of hits across all search sites was 3,077. Several referenced, unpublished, non-peer reviewed government reports were unavailable for review. Primary published studies using canines, beagles, and mongrels were evaluated to provide an informative and consistent review of mixed neutron:gamma radiation effects to establish the DRRs for the H-ARS. Secondary and tertiary studies provided additional information on the hematologic response or the effects on hematopoietic progenitor cells, radiation dosimetry, absorbed dose, and organ dose. TheLD50/30values varied with neutron quality, exposure aspect, and mixed neutron:gamma ratio. The reference radiation quality varied from 250 kVp or 1-2 MeV X radiation and60Co gamma radiation. A summary of a published review of a data set describing the DRR in rhesus macaques for mixed neutron:gamma radiation exposure in the H-ARS is included for a comparative reference to the canine dataset. The available evidence provided a reliable and extensive database that characterized the DRR for the H-ARS in canines and young rhesus macaques exposed to mixed neutron:gamma radiations of variable energy relative to 250 kVp, 1-2 MeV X radiation and60Co gamma, and uniform and non-uniform total-body irradiation without the benefit of medical management. The mixed neutron:gamma radiation showed an energy-dependent RBE of ～ 1.0 to 2.0 relative to reference radiation exposure within both species. A marginal database described the DRR for the gastrointestinal (GI)-ARS. Medical management showed benefit in both species relative to the mixed neutron:gamma as well as exposure to reference radiation. The DRR for the H-ARS was characterized by steep slopes and relativeLD50/30values that reflected the radiation quality, exposure aspect, and dose rate over a range in time from 1956-2012.

Keywords: dogs; gamma rays; radiation damage; X rays

Health Phys. 119(5):527-558; 2020

AcuteRadiation-inducedLungInjuryintheNon-humanPrimate:AReviewandComparisonofMortalityandCo-morbiditiesUsingModelsofPartial-bodyIrradiationwithMarginalBoneMarrowSparingandWholeThoraxLungIrradiation

Thomas J. MacVittie1, Ann M. Farese1, George A. Parker2, Alexander W. Bennett3, William E. Jackson III4

(1.University of Maryland School of Medicine, Baltimore, MD;2. Charles River Laboratories, Durham, NC;3. Louisville, KY, formerly at University of Maryland School of Medicine, Baltimore, MD;4. Rockville, MD)

Abstract:The nonhuman primate, rhesus macaque, is a relevant animal model that has been used to determine the efficacy of medical countermeasures to mitigate major signs of morbidity and mortality of radiation-induced lung injury. Herein, a literature review of published studies showing the evolution of lethal lung injury characteristic of the delayed effects of acute radiation exposure between the two significantly different exposure protocols, whole thorax lung irradiation and partial-body irradiation with bone marrow sparing in the nonhuman primate, is provided. The selection of published data was made from the open literature. The primary studies conducted at two research sites benefitted from the similarity of major variables; namely, both sites used rhesus macaques of approximate age and body weight and radiation exposure by LINAC-derived 6 MV photons at dose rates of 0.80 Gy min-1and 1.00 Gy min-1delivered to the midline tissue via bilateral, anterior/posterior, posterior/anterior geometry. An advantage relative to sex difference resulted from the use of male and female macaques by the Maryland and the Washington sites, respectively. Subject-based medical management was used for all macaques. The primary studies (6) provided adequate data to establish dose response relationships within 180 d for the radiation-induced lung injury consequent to whole thorax lung irradiation (male vs. female) and partial-body irradiation with bone marrow sparing exposure protocols (male). The dose response relationships established by probit analyses vs. linear dose relationships were characterized by two main parameters or dependent variables, a slope andLD50/180. RespectiveLD50/180values for the primary studies that used whole thorax lung irradiation for respective male and female nonhuman primates were 10.24 Gy [9.87, 10.52] (n=76, male) and 10.28 Gy [9.68, 10.92] (n= 40, female) at two different research sites. The respective slopes were steep at 1.73 [0.841, 2.604] and 1.15 [0.65, 1.65] probits per linear dose. TheLD50/180value and slope derived from the dose response relationships for the partial-body irradiation with bone marrow sparing exposure was 9.94 Gy [9.35, 10.29] (n=87) and 1.21 [0.70, 1.73] probits per linear dose. A secondary study (1) provided data on limited control cohort of nonhuman primates exposed to whole thorax lung irradiation. The data supported the incidence of clinical, radiographic, and histological indices of the dose-dependent lung injury in the nonhuman primates. Tertiary studies (6) provided data derived from collaboration with the noted primary and secondary studies on control cohorts of nonhuman primates exposed to whole thorax lung irradiation and partial-body irradiation with bone marrow sparing exposure. These studies provided a summary of histological evidence of fibrosis, inflammation and reactive/proliferative changes in pneumonocytes characteristic of lung injury and data on biomarkers for radiation-induced lung injury based on matrix-assisted laser desorption ionization-mass spectrometry imaging and gene expression approaches. The available database in young rhesus macaques exposed to whole thorax lung irradiation or partial-body irradiation with bone marrow sparing using 6 MV LINAC-derived radiation with medical management showed that the dose response relationships were equivalent relative to the primary endpoint all-cause mortality. Additionally, the latency, incidence, severity, and progression of the clinical, radiographic, and histological indices of lung injury were comparable. However, the differences between the exposure protocols are remarkable relative to the demonstrated time course between the multiple organ injury of the acute radiation syndrome and that of the delayed effects of acute radiation exposure, respectively.

Keywords: exposure, radiation; health effects; laboratory animals; radiation damage

Health Phys. 119(5):559-587; 2020

LackofCellularInflammationinaNon-humanPrimateModelofRadiationNephropathy

Eric P. Cohen1, Ann M. Farese2, George A. Parker3, Maureen A. Kane4, Thomas J. MacVittie2

(1.Departments of Medicine, University of Maryland, School of Medicine, Baltimore, MD;2. Radiation Oncology, University of Maryland, School of Medicine, Baltimore, MD;3. Charles River Laboratories, Durham, NC;4. Department of Pharmaceutical Sciences, University of Maryland School of Pharmacy, Baltimore, MD)

Abstract:Inflammation is commonly cited as a mechanism of delayed effects of acute radiation exposure (DEARE). Confirmation of its presence could provide significant insight to targeted use of treatments or mitigators of DEARE. We sought to quantify the presence of cellular inflammation in kidneys of non-human primates that developed acute and chronic kidney injury after a partial body irradiation exposure. We show herein that cellular inflammation is not found as a component of either acute or chronic kidney injury. Other mechanistic pathways of injury must be sought.

Keywords: kidneys; radiation damage; tissue, body; whole body irradiation

Health Phys. 119(5):588-593; 2020

EvaluationofPlasmaBiomarkerUtilityfortheGastrointestinalAcuteRadiationSyndromeinNon-humanPrimatesafterPartialBodyIrradiationwithMinimalBoneMarrowSparingthroughCorrelationwithTissueandHistologicalAnalyses

Praveen Kumar1, Pengcheng Wang1, Gregory Tudor2, Catherine Booth2, Ann M. Farese3, Thomas J. MacVittie3, Maureen A. Kane1

(1.University of Maryland, School of Pharmacy, Department of Pharmaceutical Sciences, Baltimore, MD;2. Epistem Ltd., Manchester, UK;3. University of Maryland, School of Medicine, Department of Radiation Oncology, Baltimore, MD 21201)

Abstract:Exposure to total- and partial-body irradiation following a nuclear or radiological incident result in the potentially lethal acute radiation syndromes of the gastrointestinal and hematopoietic systems in a dose- and time-dependent manner. Radiation-induced damage to the gastrointestinal tract is observed within days to weeks post-irradiation. Our objective in this study was to evaluate plasma biomarker utility for the gastrointestinal acute radiation syndrome in non-human primates after partial body irradiation with minimal bone marrow sparing through correlation with tissue and histological analyses. Plasma and jejunum samples from non-human primates exposed to partial body irradiation of 12 Gy with bone marrow sparing of 2.5% were evaluated at various time points from day 0 to day 21 as part of a natural history study. Additionally, longitudinal plasma samples from non-human primates exposed to 10 Gy partial body irradiation with 2.5% bone marrow sparing were evaluated at timepoints out to 180 d post-irradiation. Plasma and jejunum metabolites were quantified via liquid chromatography-tandem mass spectrometry and histological analysis consisted of corrected crypt number, an established metric to assess radiation-induced gastrointestinal damage. A positive correlation of metabolite levels in jejunum and plasma was observed for citrulline, serotonin, acylcarnitine, and multiple species of phosphatidylcholines. Citrulline levels also correlated with injury and regeneration of crypts in the small intestine. These results expand the characterization of the natural history of gastrointestinal acute radiation syndrome in non-human primates exposed to partial body irradiation with minimal bone marrow sparing and also provide additional data toward the correlation of citrulline with histological endpoints.

Keywords: biological indicators; gastrointestinal tract; radiation damage; radiation, ionizing

Health Phys. 119(5):594-603; 2020

ProteomicEvaluationoftheNaturalHistoryoftheAcuteRadiationSyndromeoftheGastrointestinalTractinaNon-humanPrimateModelofPartial-bodyIrradiationwithMinimalBoneMarrowSparingIncludesDysregulationoftheRetinoidPathway

Weiliang Huang1, Jianshi Yu1, Tian Liu1, Gregory Tudor2, Amy E. Defnet1, Stephanie Zalesak1, Praveen Kumar1, Catherine Booth2, Ann M. Farese3, Thomas J. MacVittie3, Maureen A. Kane1

(1.University of Maryland, School of Pharmacy, Department of Pharmaceutical Sciences, Baltimore, MD;2. Epistem Ltd., Manchester, UK;3. University of Maryland, School of Medicine, Department of Radiation Oncology, Baltimore, MD 21201)

Abstract:Exposure to ionizing radiation results in injuries of the hematopoietic, gastrointestinal, and respiratory systems, which are the leading causes responsible for morbidity and mortality. Gastrointestinal injury occurs as an acute radiation syndrome. To help inform on the natural history of the radiation-induced injury of the partial body irradiation model, we quantitatively profiled the proteome of jejunum from non-human primates following 12 Gy partial body irradiation with 2.5% bone marrow sparing over a time period of 3 wk. Jejunum was analyzed by liquid chromatography-tandem mass spectrometry, and pathway and gene ontology analysis were performed. A total of 3,245 unique proteins were quantified out of more than 3,700 proteins identified in this study. Also a total of 289 proteins of the quantified proteins showed significant and consistent responses across at least three time points post-irradiation, of which 263 proteins showed strong upregulations while 26 proteins showed downregulations. Bioinformatic analysis suggests significant pathway and upstream regulator perturbations post-high dose irradiation and shed light on underlying mechanisms of radiation damage. Canonical pathways altered by radiation included GP6 signaling pathway, acute phase response signaling, LXR/RXR activation, and intrinsic prothrombin activation pathway. Additionally, we observed dysregulation of proteins of the retinoid pathway and retinoic acid, an active metabolite of vitamin A, as quantified by liquid chromatography-tandem mass spectrometry. Correlation of changes in protein abundance with a well-characterized histological endpoint, corrected crypt number, was used to evaluate biomarker potential. These data further define the natural history of the gastrointestinal acute radiation syndrome in a non-human primate model of partial body irradiation with minimal bone marrow sparing.

Keywords: biological indicators; gastrointestinal tract; radiation damage; radiation, ionizing

Health Phys. 119(5):604-620; 2020

ProteomicsofNon-humanPrimatePlasmaafterPartial-bodyRadiationwithMinimalBoneMarrowSparing

Weiliang Huang1, Jianshi Yu1, Tian Liu1, Amy E. Defnet1, Stephanie Zalesak1, Ann M. Farese2, Thomas J. MacVittie2, Maureen A. Kane1

(1.University of Maryland, School of Pharmacy, Department of Pharmaceutical Sciences, Baltimore, MD;2. University of Maryland, School of Medicine, Department of Radiation Oncology, Baltimore, MD)

Abstract:High-dose radiation exposure results in organ-specific sequelae that occurs in a time- and dose-dependent manner. The partial body irradiation with minimal bone marrow sparing model was developed to mimic intentional or accidental radiation exposures in humans where bone marrow sparing is likely and permits the concurrent analysis of coincident short- and long-term damage to organ systems. To help inform on the natural history of the radiation-induced injury of the partial body irradiation model, we quantitatively profiled the plasma proteome of non-human primates following 12 Gy partial body irradiation with 2.5% bone marrow sparing with 6 MV LINAC-derived photons at 0.80 Gy min-1over a time period of 3 wk. The plasma proteome was analyzed by liquid chromatography-tandem mass spectrometry. A number of trends were identified in the proteomic data including pronounced protein changes as well as protein changes that were consistently upregulated or downregulated at all time points and dose levels interrogated. Pathway and gene ontology analysis were performed; bioinformatic analysis revealed significant pathway and biological process perturbations post high-dose irradiation and shed light on underlying mechanisms of radiation damage. Additionally, proteins were identified that had the greatest potential to serve as biomarkers for radiation exposure.

Keywords: biological indicators; health effects; partial body radiation; radiation damage

Health Phys. 119(5):621-632; 2020

DevelopmentofaModeloftheAcuteandDelayedEffectsofHighDoseRadiationExposureinJacksonDiversityOutbredMice;ComparisontoInbredC57BL/6Mice

Andrea M. Patterson1, P. Artur Plett1, Hui Lin Chua1, Carol H. Sampson1, Alexa Fisher1, Hailin Feng1, Joseph L. Unthank2, Steven J. Miller2, Barry P. Katz3, Thomas J. MacVittie4, Christie M. Orschell1

(1.Department of Medicine, Indiana University School of Medicine, Indianapolis, IN;2. Department of Surgery, Indiana University School of Medicine, Indianapolis, IN;3. Department of Biostatistics, Indiana University School of Medicine, Indianapolis, IN;4. Department of Radiation Oncology, University of Maryland School of Medicine, Baltimore, MD)

Abstract:Development of medical countermeasures against radiation relies on robust animal models for efficacy testing. Mouse models have advantages over larger species due to economics, ease of conducting aging studies, existence of historical databases, and research tools allowing for sophisticated mechanistic studies. However, the radiation dose-response relationship of inbred strains is inherently steep and sensitive to experimental variables, and inbred models have been criticized for lacking genetic diversity. Jackson Diversity Outbred (JDO) mice are the most genetically diverse strain available, developed by the Collaborative Cross Consortium using eight founder strains, and may represent a more accurate model of humans than inbred strains. Herein, models of the Hematopoietic-Acute Radiation Syndrome and the Delayed Effects of Acute Radiation Exposure were developed in JDO mice and compared to inbred C57BL/6. The dose response relationship curve in JDO mice mirrored the more shallow curves of primates and humans, characteristic of genetic diversity. JDO mice were more radioresistant than C57BL/6 and differed in sensitivity to antibiotic countermeasures. The model was validated with pegylated-G-CSF, which provided significantly enhanced 30-d survival and accelerated blood recovery. Long-term JDO survivors exhibited increased recovery of blood cells and functional bone marrow hematopoietic progenitors compared to C57BL/6. While JDO hematopoietic stem cells declined more in number, they maintained a greater degree of quiescence compared to C57BL/6, which is essential for maintaining function. These JDO radiation models offer many of the advantages of small animals with the genetic diversity of large animals, providing an attractive alternative to currently available radiation animal models.

Keywords: bone marrow; health effects; mice; whole body irradiation

Health Phys. 119(5):633-646; 2020

ImmuneReconstitutionandThymicInvolutionintheAcuteandDelayedHematopoieticRadiation

Syndromes

Tong Wu1, P. Artur Plett1, Hui Lin Chua1, Max Jacobsen2, George E. Sandusky2, Thomas J. MacVittie3, Christie M. Orschell1

(1. Department of Medicine, Indiana University School of Medicine, Indianapolis, IN;2. Department of Pathology and Laboratory Medicine, Indiana University School of Medicine, Indianapolis, IN;3. Department of Radiation Oncology, University of Maryland School of Medicine, Baltimore, MD)

Abstract:Lymphoid lineage recovery and involution after exposure to potentially lethal doses of ionizing radiation have not been well defined, especially the long-term effects in aged survivors and with regard to male/female differences. To examine these questions, male and female C57BL/6 mice were exposed to lethal radiation at 12 wk of age in a model of the Hematopoietic-Acute Radiation Syndrome, and bone marrow, thymus, spleen, and peripheral blood examined up to 24 mo of age for the lymphopoietic delayed effects of acute radiation exposure. Aged mice showed myeloid skewing and incomplete lymphocyte recovery in all lymphoid tissues. Spleen and peripheral blood both exhibited a monophasic recovery pattern, while thymus demonstrated a biphasic pattern. Na?ve T cells in blood and spleen and all subsets of thymocytes were decreased in aged irradiated mice compared to age-matched non-irradiated controls. Of interest, irradiated males experienced significantly improved reconstitution of thymocyte subsets and peripheral blood elements compared to females. Bone marrow from aged irradiated survivors was significantly deficient in the primitive lymphoid-primed multipotent progenitors and common lymphoid progenitors, which were only 8%-10% of levels in aged-matched non-irradiated controls. Taken together, these analyses define significant age- and sex-related deficiencies at all levels of lymphopoiesis throughout the lifespan of survivors of the Hematopoietic-Acute Radiation Syndrome and may provide a murine model suitable for assessing the efficacy of potential medical countermeasures and therapeutic strategies to alleviate the severe immune suppression that occurs after radiation exposure.

Keywords: bone marrow; health effects; mice; whole body irradiation

Health Phys. 119(5):647-658; 2020

APotentialRoleforExcessTissueIroninDevelopmentofCardiovascularDelayedEffectsofAcuteRadiationExposure

Steven J. Miller1, Supriya Chittajallu1, Carol Sampson2, Alexa Fisher2, Joseph L. Unthank1, Christie M. Orschell2

(1.Department of Surgery, Indiana University School of Medicine, Indianapolis, IN 46202-5181;2. Department of Medicine, Indiana University School of Medicine, Indianapolis, IN 46202-5181)

Abstract:Murine hematopoietic-acute radiation syndrome (H-ARS) survivors of total body radiation (TBI) have a significant loss of heart vessel endothelial cells, along with increased tissue iron, as early as 4 mo post-TBI. The goal of the current study was to determine the possible role for excess tissue iron in the loss of coronary artery endothelial cells. Experiments used the H-ARS mouse model with gamma radiation exposure of 853 cGy (LD50/30) and time points from 1 to 12 wk post-TBI. Serum iron was elevated at 1 wk post-TBI, peaked at 2 wk post-TBI, and returned to non-irradiated control values by 4 wk post-TBI. A similar trend was seen for transferrin saturation, and both results correlated inversely with red blood cell number. Perls’ Prussian Blue staining, used to detect iron deposition in heart tissue sections, showed myocardial iron was present as early as 2 wk following irradiation. Pretreatment of mice with the iron chelator deferiprone decreased tissue iron but not serum iron at 2 wk. Coronary artery endothelial cell density was significantly decreased as early as 2 wk vs. non-irradiated controls (P<0.05), and the reduced density persisted to 12 wk after irradiation. Deferiprone treatment of irradiated mice prevented the decrease in endothelial cell density at 2 and 4 wk post-TBI compared to irradiated, non-treated mice (P<0.03). Taken together, the results suggest excess tissue iron contributes to endothelial cell loss early following TBI and may be a significant event impacting the development of delayed effects of acute radiation exposure.

Keywords: mice; radiation damage; radiation dose; whole body radiation

Health Phys. 119(5):659-665; 2020