Radiation Combined Injury: Models, Mechanisms, and Countermeasures
April 5, 2012, symposium sponsored by:
The Armed Forces Radiobiology Research Institute (AFRRI) at the Uniformed Services University of the Health Sciences (USU) in Bethesda, Maryland. To learn more about the symposium, please contact symposium chairman, Dr. Juliann G. Kiang (e-mail or 301-295-1076), or symposium officer-in-charge, LT Joshua M. Swift (e-mail or 301-295-1947).
Past, Present, and Future in Radiation Combined Injury
G. David Ledney, PhD
Retired, formerly with the Combined Injury Program, Armed Forces Radiobiology Research Institute
The condition identified as combined injury (CI) is not a new phenomenon but rather, was described more than 9 decades ago. Due to recent concerns relating to the high risk of nuclear weapon use, attention has been focused on establishing useful animal systems for evaluating the consequences of exposure to radiation in conjunction with injuries associated with nuclear weapon detonation. Casualties are expected to overwhelm healthcare facilities and thus it is imperative to determine (1) the physiologic changes resultant from radiation, tissue injury, and CI that lead to morbidity and mortality and (2) countermeasures useful for mass-casualty applications. In our CI-studies, mice received various doses and qualities of x-rays, 60Co-γ-photons, and reactor-produced mixed field (n + γ-photons) radiation given at 0.4 Gy/min. Compared to x-irradiation and 60Co-γ-photon irradiation, as the Dn/Dt increased the RBE increased and the LD50/30 decreased. The addition of a standard-sized wound or thermal burn in irradiated mice further increased the RBE and further decreased the LD50/30. In all cases, wounding subsequent to irradiation resulted in greater 30-day mortality and protracted wound healing compared to thermal injury. Compared to wounding after irradiation, wounding prior to irradiation decreased mortality. While wounding, compared to thermal injury, had a greater impact on survival, both wounding and thermal burns resulted in serum increases of C-reactive protein, C3, and PGE2. Decreased IG production along with an early rise in corticosterone followed by a subsequent decrease was noted for each CI situation. Many challenges face investigators studying CI. Perhaps the most important are (1) finding effective countermeasures promoting short-term survival, (2) evaluating the consequences of partial-body irradiation along with tissue injuries, and (3) development of a thermal burn situation for testing in animals. (PDF of abstract)
Infection Complications of Combined Injury and Antimicrobial Countermeasures
Thomas B. Elliott, PhD
Senior Principal Investigator, Armed Forces Radiobiology Research Institute
Traumatic injury inflicted after irradiation increases mortality. Ionizing radiation increases susceptibility to infection. Radio-sensitive proliferative cells are damaged in bone marrow and intestinal epithelium. Hematopoiesis diminishes and innate immune responses are depressed. Bacterial infections are a major cause of morbidity and mortality after whole-body doses of ionizing radiation, which cause hematopoietic or intestinal failure. After lethal doses of radiation, polymicrobial sepsis develops from endogenous facultative bacteria. Species of Gram-positive Enterococcus, Streptococcus, Staphylococcus and Gram-negative Enterobacteriaceae cause polymicrobial sepsis in combined injured (CI) laboratory animals. Infection and sepsis occur earlier after CI. Traumatic injury further complicates infection management after irradiation. The wound is an additional portal of entry for environmental microorganisms. Elevated doses of antimicrobial agents may eradicate infections but other therapeutic interventions are needed to promote recovery of innate immune responses and intestinal epithelium, reduce bacterial translocation, as well as eliminate potentially pathogenic microorganisms in intestines in order to enhance survival. Understanding changes in pharmacokinetics after irradiation improves effective use of antimicrobial agents after irradiation. Quinolones may be the optimal agents to use because they may be delivered orally, particularly with mass casualties. Antimicrobial therapy with levofloxacin was more effective than other quinolones after irradiation. Combination therapy of an immunomodulator together with antimicrobial agents improved survival following lethal irradiation. The combination was effective in sublethally irradiated CI mice but only extended survival time a few days in lethally irradiated CI mice. Topical silvadene and gentamicin creams protected sublethally irradiated CI mice. Therapy for sepsis and recovery of radiation-injured tissues needs improvement, particularly after CI. (PDF of abstract)
Mathematical Modeling of Combined Injury: Radiation and Burns
Daniela L. Stricklin, PhD
Principal Scientist, Applied Research Associates in Arlington, Virginia
Health effect models that mathematically describe pathophysiological mechanisms of radiation combined injury can facilitate integration of modern medical expertise and cutting-edge research into practical tools for estimating casualties and resource requirements by emergency response planners. Applied Research Associates, Inc. (ARA), under contract with the Defense Threat Reduction Agency (DTRA), has developed a software tool, Radiation Induced Performance Decrement (RIPD), using physiologically based radiation injury models. The models in RIPD are used to predict probability of lethality, time to lethality, time-dependent severity of the signs and symptoms of acute radiation syndrome, and performance decrement after untreated acute or protracted radiation exposures. RIPD is being updated with models that predict additional clinical parameters and expanded to include models for additional injury types such as burn, trauma, and radiation combined injury. A brief overview of current radiation and burn injury models, their associated pathophysiological bases, and points of synergistic interaction will be provided. Models that describe effects at different levels (i.e., from whole organism response to molecular-level response) will be discussed. Their outputs from higher-level models can be used to improve casualty estimates and provide insight on resource requirements. More detailed physiologically based models can help explain synergistic effects, provide projections on time to presentation of clinically relevant symptoms, and help identify targets for treatment in combined injury. Tools incorporating physiologically-based models can be used by planners to gain insight on patient-flow and potentially the impact of different medical countermeasures. Physiologically based mathematical models allow the translation of complex biomedical research into practical information for application in emergency response planning. (PDF of abstract)
Wound Trauma Combined With Ionizing Irradiation Affects Biodosimetric Assessment
Juliann G. Kiang, PhD
Program Advisor, Radiation Combined Injury Program, Armed Forces Radiobiology Research Institute;
Professor, Department of Radiation Biology, Uniformed Services University of the Health Sciences
Wounding following whole-body γ-irradiation (radiation combined injury, RCI) increases mortality. Wounding-induced increases in radiation mortality are triggered by sustained activation of inducible nitric oxide synthase pathways, persistent alteration of cytokine homeostasis, and increased susceptibility to bacterial infection. Among these factors, cytokines along with other biomarkers have been adopted for biodosimetric evaluation and assessment of radiation dose and injury. Therefore, wounding could complicate biodosimetric assessments. In our laboratory, such confounding effects were addressed. Mice were given 60Co γ-photon radiation followed by 15% total-body surface area skin wounding. Wound trauma exacerbated radiation-induced mortality, bodyweight loss, and wound healing. Analyses of DNA damage in bone-marrow cells and peripheral blood mononuclear cells (PBMCs), changes in hematology and cytokine profiles, and fundamental clinical signs were evaluated. Early biomarkers (l d after RCI) vs. irradiation alone included enhanced increases in γ-H2AX formation in Lin+ bone marrow cells and increases in IL-1ß, IL-6, IL-8, and G-CSF concentrations in blood, and concomitant decreases in γ-H2AX formation in PBMCs and decreases in numbers of splenocytes, lymphocytes, and neutrophils. Intermediate biomarkers (7–10 d after RCI) included continuously decreased γ-H2AX formation in PBMCs and enhanced increases in IL-1ß, IL-6, IL-8, and G-CSF concentrations in blood. The clinical signs evaluated after RCI were increased water consumption, decreased body weight, and decreased wound healing rate and survival rate. Late clinical signs (30 d after RCI) included poor survival and wound healing. Results suggest that confounding factors such as wounding complicate biodosimetric assessment and agents inhibiting these responses may prove therapeutic for RCI and reduce related mortality. [The views expressed here are those of the author; no endorsement by the U.S. Department of Defense or the U.S. Government has been given or inferred. Supported by National Institutes of Health contract YI-AI5045-04 and grants R21/33 AI080553] (PDF of abstract)
Effect of Ionizing Radiation on Combined Skin Wound Injury
A. Lopez1, E.B. Olasz1, J.E. Moulder2, S.R. Doctrow3, Z. Lazarova1, MD
Departments of Dermatology1 and Radiology2, Medical College of Wisconsin, Milwaukee; Department of Medicine3, Boston University
Aim: Assess the effect of radiation on combined skin and wound injury.
Background: In the event of a radiological terrorist attack, skin injury may be combined with wounds thus impacting prognosis of affected individuals. Laminin 332, an epidermal basement membrane protein, plays an important role in wound healing. We investigated the effect of radiation on laminin 332 expression. Chronic oxidative stress has been suggested as a contributor to the progression of radiation-induced skin injury. To examine the role of oxidative stress in radiation-induced skin injury, we tested the effects of a synthetic superoxide dismutase/catalase mimetic, EUK-207, in a rat model of combined skin irradiation and wound injury.
Methods: Using a soft x-ray beam, a single dose of ionizing radiation with a steep dose gradient (0–40 Gy) was delivered to the skin. Skin was wounded, wounds were measured, and tissue samples were evaluated for laminin 332 and matrix metalloproteinase-2 (MMP-2) expression. In a separate set of experiments, EUK-207 or vehicle was administered systemically 48 hr following irradiation/wounding. Skin injury and wounds were measured. Tissue samples were evaluated for oxidative stress markers.
Results: Radiation significantly delayed wound healing, elevated expression of laminin 332 genes and decreased deposition of laminin 332 protein in the skin. Elevated laminin 332 gene expression was paralleled with elevated gene and protein expression of MMP-2, suggesting that the reduced amount of laminin 332 in irradiated skin is due to an imbalance between laminin 332 secretion and processing by elevated tissue metalloproteinases. EUK-207 administration mitigated radiation dermatitis, suppressed indicators of tissue oxidative stress, and enhanced wound healing in irradiated skin.
Conclusion: Laminin 332 deposition is inhibited by ionizing radiation, which contributes to the delayed wound healing of irradiated skin. In addition, these results support the critical role of oxidative stress in the progression of radiation-induced skin injury, and that antioxidant compounds, administered after exposure, can mitigate radiation-induced skin injury. (PDF of abstract)
Redirecting the Immune System as a Medical Countermeasure to Radiation Combined Injury
Veit M. Stoecklein, Akinori Osuka, and James Lederer, PhD
Brigham and Women’s Hospital and Harvard Medical School in Boston, Massachusetts
Background, Aim, and Rationale: The complexity of a radio-nuclear event would be immense due to varying levels of radiation exposures and injuries caused by blast-associated trauma. With this scenario in mind, we developed a mouse model to mimic as closely as possible the possible consequences of radiation injury and radiation combined injury (RCI). Immune response phenotyping data revealed that RCI disrupted immune system homeostasis and predisposed mice to sepsis and associated complications. This RCI mouse model was established to develop and test novel countermeasure approaches for protecting individuals from the immunological complications of radiation and RCI. We tested the general hypothesis that early post-injury treatments will redirect the immune response to RCI to help restore immune system homeostasis and function. In this presentation, we will report our findings on using CpG oligodeoxynucleotides (ODNs) as highly-effective immune response-modifying treatments that restore immune system homeostasis and protect RCI mice from sepsis and its complications.
Methods: Mice were exposed to 1, 4, or 6 Gy whole-body radiation and underwent near-simultaneous 25% surface-area full-thickness burn injury. At one day after injury, mice were given class A, B, or C CpG ODNs, non-CpG ODNs, or saline. Seven days later, mice were challenged by cecal ligation and puncture (CLP) to cause polymicrobial sepsis and survival was recorded. Immune system phenotyping studies were performed to measure immune cell subset changes and cytokine production profiles. Finally, fluorescently-labeled CpG ODNs were used to track the in vivo uptake of CpG ODNs in immune cell subsets following treatment.
Results: Treatment with class A CpG ODN significantly increased survival after RCI with 1 Gy, whereas treatment with class B or C did not. Importantly, class A CpG ODN treatment also conferred a significant survival benefit following RCI with 4 and 6 Gy. Furthermore, we found that inflammatory cytokine production was decreased with class A treatment but not with class B or C. We identified that a CD4+ T cell subset called regulatory T cells (Tregs) were most affected by class A CpG ODN and found that depleting Tregs in CpG ODN-treated mice eliminated the sepsis-survival benefit. The importance of Tregs was further supported by the finding that Tregs showed increased uptake of class A CpG as compared to class B or C.
Conclusions: We show that early interventional treatment with class A CpG ODN helped redirect the immune system response in RCI to effectively enhance host defense mechanisms in mice subjected to mild and severe RCI in animals that would otherwise have succumbed to infection and associated complications. Furthermore, these findings directly demonstrate that early treatment with immune response-modifying agents can functionally restore immune system homeostasis to provide significant protection from the immunological complications of radiation and radiation combined injury. (PDF of abstract)
Ghrelin as a Novel Therapy for Radiation Combined Injury
Ping Wang, MD
Professor of Surgery, Albert Einstein College of Medicine; Vice Chairman for Research, Department of Surgery, North Shore University Hospital & Long Island Jewish Medical Center in New York
Aim: The purpose of this study was to determine whether ghrelin is protective in a rat model of radiation combined injury (RCI) and to elucidate its potential mechanism.
Background: Radiation exposure due to nuclear terrorism and the most recent radiation leak at the Fukushima nuclear plant resulted in the continued awareness of the danger in whole-body irradiation (WBI). In these scenarios, radiation itself may not cause immediate illnesses or large casualties, but radiation victims will likely develop additional complications such as infection and sepsis.
Rationale: Ghrelin, a gastric hormone, has been shown to ameliorate sepsis-induced organ injury and mortality. Ghrelin’s effect on RCI has not been previously elucidated.
Methods: Adult male rats were exposed to 5-Gy WBI followed by cecal ligation and puncture (CLP) sepsis 48 hr thereafter to model RCI. Immediately after WBI, human ghrelin (2 nmol/rat) was given iv bolus, followed by continuous infusion for 68 hr (26 nmol) with another iv bolus at the time of CLP totaling 30 nmol/rat. A 10-day survival study also was conducted. To assess the mechanism, vagotomy was performed in RCI animals immediately prior to ghrelin administration.
Results: After RCI, serum levels of ghrelin and its gene expression in the stomach were dramatically decreased. Marked increases in liver enzymes (AST, ALT), lactate, LDH and creatinine, cytokine levels (TNF-α, IL-6), myeloperoxidase activities in the lungs, gut and kidneys were also observed. Human ghrelin treatment significantly reduced these levels by 40–60%. Treatment improved survival rate significantly to 69% from 38% (vehicle-treated). Norepinephrine, a sympathetic neurotransmitter, was markedly increased in RCI while ghrelin treatment decreased those levels by 35%. Vagotomy prior to RCI completely abolished ghrelin’s inhibitory effect.
Conclusion: Human ghrelin is beneficial in RCI through the regulation of the dysregulated sympathetic/parasympathetic nervous systems. (PDF of abstract)
Ciprofloxacin Increases Survival after Ionizing Radiation Combined with Wound Trauma
Risaku Fukumoto, PhD
Research Scientist, Armed Forces Radiobiology Research Institute
It is evident that irradiation combined with non-lethal wound trauma leads to greater mortality than irradiation alone. The LD50/30 for wounding/irradiation combined injury (CI) is 8.95 Gy but is 9.65 Gy for irradiation alone (RI). A spectrum of specific, time-dependent pathophysiological changes is observed in this CI model. Of these changes, the massive release of pro-inflammatory cytokines, severe hematologic losses, and bacterial sepsis could prove to be especially useful treatment targets to improve survival after CI. In this study, we investigated whether ciprofloxacin (CIP) could ameliorate pathophysiological changes after CI. In addition to its antimicrobial activity, CIP is a type II topoisomerase inhibitor and hematopoietic stimulator; also, it is already part of the emergency national drug stock pile. We treated B6D2F1/J mice with CIP within 2h after CI and following daily oral route for 3 weeks. Within 10 days, CIP treatment not only significantly reduced pro-inflammatory cytokine concentrations but it also enhanced erythrocyte production. Treatment reduced the amount of body weight loss and accelerated wound healing during the first 30 days after CI. At day 30, surviving mice treated with CIP displayed a greater repopulation of bone marrow cells. CIP treatment led to better survival in CI-mice compared to RI-mice despite the fact that CIP effectively removed target microorganisms in both groups. Given the multiple beneficial activities of CIP shown in our experiments, CIP may prove to be a useful treatment for CI. (PDF of abstract)
Ciprofloxacin Inhibits Radiation Combined Wound Trauma-Induced ATP Loss by Preserving Pyruvate Dehydrogenase
Joshua M. Swift, PhD; Joan T. Smith; and Juliann G. Kiang, PhD
Radiation Combined Injury Program, Armed Forces Radiobiology Research Institute
Background: Ionizing radiation combined with wound injury (RCI) increases animal mortality more than ionizing radiation alone. Ciprofloxacin (CIP) is a fluroquinolone, a synthetic antibiotic, found in the national stockpile for emergency use and known to inhibit bacterial sepsis. The purpose of this study was to evaluate the efficacy of CIP as a countermeasure to RCI mortality and determine the signaling proteins involved in energy machinery.
Methods: B6D2F1/J female mice were randomly assigned to receive either 9.75 Gy Co-60 gamma radiation followed by skin wounding (RCI) or sham procedures (SHAM). Either CIP (90 mg/kg q.d.) or vehicle (VEH; water) were administered orally to these mice starting 1 hr after wounding and thereafter daily for 10 days. Determination of ileum ATP was conducted, and immunoblotting for signaling proteins involved in ATP machinery was conducted.
Results: RCI resulted in 60% survival after 10 days as compared to 100% survival in the SHAM group. Furthermore, RCI caused significant reductions in ileum ATP concentration (–82%) as compared to SHAM. CIP administration after RCI resulted in 100% survival and increased ATP (+5-fold) as compared to RCI. Protein levels of heat shock protein 70 kDa (HSP70; a chaperone protein involved in ATP synthesis) and pyruvate dehydrogenase (PDH; an enzyme complex crucial to conversion of pyruvate to acetyl CoA for entrance into TCA cycle) were significantly lower in RCI group (vs. SHAM). Using immunoprecipitation and immunoblotting, HSP70-PDH complex was found to be present in the ileum tissue of RCI mice treated with CIP.
Conclusion: These data suggest that CIP administration following RCI may increase animal survival by maintaining ileum ATP synthesis by preserving PDH. Furthermore, our findings imply that CIP treatment may be a valuable therapeutic treatment for RCI.
Potential impact to mission/warfighter: Ciprofloxacin is in the national stockpile for emergency use, making ciprofloxacin an attractive treatment for RCI.
(Supported by NIH/NIAID R33-A1080553 to JGK) (PDF of abstract)
Mesenchymal Stromal Cells as a Countermeasure for Radiation Combined Injury: Friend or Foe?
Nikolai V. Gorbunov, PhD
Research Scientist, Armed Forces Radiobiology Research Institute
Radiation combined injury (RCI) is a pathophysiological condition due to ionizing irradiation combined with trauma or other insults that complicate systemic responses and exacerbate the acute radiation syndrome. Considering the growing threat from potential radiological and nuclear incidents, development of effective CI countermeasures is important to military and civilian personnel. Tissue injury is followed by recruitment of lymphoid, myeloid, and mesenchymal components at the sites of injury in order to reconstitute tissue barriers. Ionizing irradiation can inhibit this effect by damaging bone marrow—a major source of leukocytes and mesenchymal stromal cells (MSCs). Recent discovery of immunomodulatory and antibacterial function of MSCs provides new insight into the role of stroma in tissue barriers and opens new perspectives for management of the traumatic tissue injury and bacterial infection combined with ionizing irradiation. This communication is focused on assessment of survival of mice engrafted with bone marrow-derived MSCs after irradiation of mice combined with a penetrating wound (RCI). MSCs were obtained from B6D2F1/Jfemale mice and cultivated in hypoxic conditions for 4–12 weeks in MesenCult medium. Phenotype and cell proliferative activity were analyzed with flow cytometry and immunofluorescence imaging; the cells were identified by the presence of MSC-positive markers (CD44, and SCA1) and an absence of MSC-negative marker (CD34). RCI was induced in B6D2F1/J female mice by 9.25 Gy 6oCo-photon irradiation followed by a 15% total-body, surface skin-wound trauma within 1 hour. Twenty-four hours later, animals received 0.4 ml of Dulbecco's Modified Eagle Medium (DMEM) or 3 × 106 MSCs in DMEM vehicle. Animal survival, body weight, water consumption, and wound closure were monitored for 30 days. Data were analyzed by a Kaplan-Meier survival curve. Statistical significance was determined using one-way ANOVA followed by post-hoc analysis with pair-wise comparison by Tukey-Kramer test. Transplantation with 4-week old MSCs mitigated RCI-induced hematopoietic depression yielding a 20–30% increase in survival. That was accompanied by a substantial improvement in wound healing. These effects were diminished when transfused MSCs were composed from 4-week-old and 12-week-old cultures. The data support the contention that prediction of outcomes of the MSC therapy requires further investigation of the biological properties and mechanisms of action of the transplanted cells.
(Supported by NIH/NIAID YI-AI-5045-04) (PDF of abstract)
Role of Hematopoietic Cells in Radiation Combined Injury
Division of Cellular Therapy/Bone Marrow Transplantation, Department of Medicine: Divino Deoliveira*, Yiqun Jiao*, Wei Huang, Dunhua Zhou, Joel R. Ross, Kayla Corbin, Qizhen Xiao, Benny J. Chen*, and Nelson J. Chao* (*contributing equally)
Duke Cancer Institute: Benny J. Chen, Nelson J. Chao
Department of Radiation Safety: Greta Toncheva, Colin Anderson-Evans
Department of Radiation Safety and Radiology: Terry T. Yoshizumi
Department of Pathology and Immunology: Nelson J. Chao
Duke University Medical Center in Durham, North Carolina
Aim: To determine the role of hematopoietic cells in recovery after radiation and wound combined injury.
Background and Rationale: Radiation and wound combined injury represents a major clinical challenge because of the synergistic interactions that lead to higher morbidity and mortality. It is not completely clear whether and how hematopoietic cells participate in wound healing.
Methods: The role of hematopoietic cells in radiation and wound combined injury was studied using a newly developed ear-punch model. Wound healing after local and systemic irradiation was compared clinically and histologically. The role of hematopoietic cells was further studied after bone marrow transplantation.
Results: Using the newly established ear-punch model, we first demonstrated that local radiation to the wound area significantly delayed the healing of ear-punch wounds in a dose-dependent fashion. The addition of sublethal whole-body irradiation further delayed the healing of ear-punch wounds. Hematopoietic cell transplantation partially reversed the delay in wound healing after local and systemic irradiation.
Conclusion: These data indicate that an intact hematopoietic system is important for wound repair and regeneration after radiation and wound combined injury. The underlying mechanisms can be further studied by high resolution in vivo imaging using newly available genetic models and two-photon microscopy. (PDF of abstract)
Role of Radiation-Induced Skin Damage as an Essential Aspect in Combined Injuries
COL Viktor Meineke, MD, MC, German Army
Professor and Director Bundeswehr Institute of Radiobiology affiliated to the University of Ulm Neuherbergstr in Munich, Germany
The skin organ plays a crucial role in accidental radiation exposure, both in case of whole-body and significant partial-body exposure, but also in context with localized radiation injuries. After whole-body exposure, depending on radiation type and dose, the skin is one of the affected organs and multi-organ interaction is the predominant pathophysiological principle determining the outcome of radiation victims. However, extensive skin affection always is a sign for a very bad prognosis. In contrast, in localized radiation injuries the skin is the prominent affected organ and interactions with other organ systems play a minor role. These pathophysiological considerations are extremely important in order to adjust appropriate diagnostic procedures to estimate the degree of radiation damage as well as to decide upon therapeutical interventions.
Since the early studies in the 1940s and 50s, it is well known that additional trauma to the skin, such as burns and/or conventional injuries in the classical sense of a combined injury, exponentially worsens the outcome of irradiated victims. In the scenario of an improvised nuclear device (IND), a combination of thermal and conventional injury will have to be noticed, whereas in the case of a pure external gamma irradiation, there will only be a minor burn component to be taken into account. Thermal injury and radiation injury share similarities concerning the clinical picture, but the underlying pathophysiological principles are totally different, due to different energies and penetrations into the skin, as well as different effects on the dermal stem cell compartment. Recent studies have provided more and more insight into the molecular mechanisms involved in combined injuries. Nevertheless, the main therapeutic principle in the case of combined injuries still is that needed surgical procedures, such as wound debridement or wound closures, have to be done as early as possible but no later than three days after radiation exposure. (PDF of abstract)
Macrophage Response to Ionizing Radiation and Virus Infection: Puzzling the Pathways
R. Joel Lowy, PhD
Program Advisor, Radiation Neutralization, Armed Forces Radiobiology Research Institute
The pathophysiology of combined injury due to ionizing radiation (IRAD) and infectious disease agents remains poorly defined. Infectious disease exposure concurrent with IRAD can occur from endemic microorganisms, as well as from wounds or endogenous sources. Influenza virus (FLUA) circulates yearly, causes significant mortality and morbidly and like IRAD is immune suppressive. Ionizing radiation and viral infections both elicit production of host defense factors including cytokines (CK) and chemokines (CHK), whose production is controlled by several well known cell signaling mechanisms. These include pathways resulting in activation of Nuclear Factor kappa B (NFκB), Mitogen Activated Protein Kinase (MAPK), Interferon Response Factors (IRF), and NOD Like Receptor (NLR). The responses of macrophages to combined IRAD and FLUA are particularly ill-defined, despite these cells being central to innate immune response and important for host survival. The murine macrophage cultured cells J774.1 and RAW 267.1 were exposed to FLUA and 6OCo gamma photons IRAD singly and in combination. The production of CK/CHK and the response of the NFκB and MAPK cell signaling pathways were assayed. FLUA stimulates CK/CHK, as expected, whereas IRAD can be suppressive or stimulatory. Interestingly IRAD modulates production of some but not all CK/CHKs produced in response to FLUA. Surprisingly, activation of the NFκB-dependent pathway does not appear to be the major pathway transducing responses in these cells; in contrast MAPKases appear to have a major role. (PDF of abstract)
Current Gaps in Radiation Combined Injury Research
Glen I. Reeves, MD
Principal Scientist, Applied Research Associates, Inc., in Arlington, Virginia
The pathophysiology and medical management of burns and trauma in humans have been studied for centuries. Local and systemic radiation exposure injuries also have been well researched. We currently have a good grasp on what gaps in our knowledge of models, mechanisms, and countermeasures for these insults, considered individually, remain. However, radiation combined injury in humans has not been as well studied. In Japan, two-thirds of the casualties had burns and/or trauma in addition to significant radiation exposure.
We know that the overall effect of radiation combined injury is synergistic, not merely additive; the response to sepsis from burns or trauma is hindered by radiation-induced leukopenia. Silver-impregnated dressings, which are necessary for treatment of extensive severe burns, can occasionally induce marked neutropenia; the interaction of this therapeutic countermeasure with radiation exposure has not been well studied. The demographic responses to the different components of combined injury vary by gender and age. Vascular permeability is increased in severe burns and significant radiation exposure; however, these changes may be by different mechanisms. It has been recommended that colony stimulating factors be given at lower exposures in combined vs. radiation alone injury. Because the effects on function, location, and number of granulocytes vary depending on insult, further study is needed to refine treatment protocols. Epidermal growth factor, useful in burn treatment, is also effective in gastrointestinal injury from radiation in animal models. Clinicians facing multiple radiation combined injury casualties need to be aware how differing mechanisms of injury might modify existing treatment protocols. (PDF of abstract)
Challenges in Developing Medical Countermeasures for Combined Injury
Ronald Manning, Rodney Wallace, Marcy Grace, Mary Horner, Tom Hu, Narayan Iyer, Brian Moyer, Brian Tse, Lynne Wathen, and Richard Hatchett, MD
Biomedical Advanced Research and Development Authority, Office of the Assistant Secretary for Preparedness and Response, U.S. Department of Health and Human Services
The Biomedical Advanced Research and Development Authority (BARDA) supports the advanced research, development, and procurement of medical countermeasures (MCMs) against a variety of national security threats and in recent years has significantly expanded its investment in radiation therapeutics and diagnostics. The Public Health Emergency Medical Countermeasures Enterprise (PHEMCE), of which BARDA is a component, has also identified gaps in preparedness to provide blood products and treat thermal burns following the detonation of an improvised nuclear device, and BARDA is considering how it might support the development of MCMs to remedy these gaps. Developing MCMs to diagnose and treat radiation combined injury per se, however, poses significant conceptual, practical, and regulatory challenges. “Radiation combined injury” is loosely defined and includes a broad array and scope of injuries, which may confound novel point-of-care and high-throughput methods of dose estimation; specific clinical protocols for treating patients receiving significant absorbed doses of ionizing radiation—in the setting of non-radiation components of combined injuries have not been developed; animal models are rudimentary and difficult to extrapolate to humans; and little progress has been made in establishing regulatory pathways for MCMs seeking a radiation combined injury indication. BARDA’s current approach is to support the development of MCMs for the different components of acute radiation syndrome, thermal burns, and transfusion needs in isolation. Elucidation of the underlying mechanisms of pathogenesis of radiation combined injury and the development and validation of relevant animal models will be required before specific radiation combined injury MCMs and diagnostics can be developed. (PDF of abstract)
NIAID Radiation/Nuclear Medical Countermeasure Research & Development Program on Combined Radiation Injuries
Andrea DiCarlo, PhD; Narayani Ramakrishnam, PhD; and Bert Maidment, PhD
Division of Allergy, Immunology, & Transplantation; U.S. National Institute of Allergy & Infectious Diseases
Over the past decade, there has been increased awareness that terrorists could attack using radiation. These attacks could involve placement of concealed radiation sources, detonation of a dirty bomb, or attacks on nuclear power plants or nuclear waste facilities. The highest impact scenario would be detonation of a nuclear explosive device, which, in addition to causing damage from blast and heat, would produce an intense burst of gamma radiation and large quantities of radioactive fallout. Concomitant with the radiation exposure, other bodily damage would be expected in many of the survivors, including thermal and/or radiation burns, lacerations, fractures, infections and traumatic brain injuries. Established in 2005, NIAID’s Radiation Program is tasked with funding research and development to accelerate licensure of medical countermeasures for the different organs and systems affected by radiation exposure. Given anticipated exposure scenarios, in 2008 NIAID awarded 10 grants to fund animal model and countermeasure development, which will be needed to FDA licensure, in the area of radiation combined injuries. Although several models were previously developed at AFRRI, NIAID currently funds expansion of and countermeasures testing in these models via interagency agreement. Funded grants now encompass three projects on burns, three involving wounds, three studying infection and one looking at traumatic brain injury concomitant with radiation exposure. Now in the fourth year, researchers have validated new animal models for expected combined injuries, and have generated intriguing data on the use of several countermeasure approaches to improve survival and enhance cutaneous and other healing in the presence of radiation exposure. (PDF of abstract)