Hematopoietic Acute Radiation Syndrome Research Articles

Interleukin-12 sustained release system promotes hematopoietic recovery after radiation injury.

The continuous production of mature blood cell lineages is maintained by hematopoietic stem cells but they are highly susceptible to damage by ionizing radiation (IR) that induces death. Thus, devising therapeutic strategies that can mitigate hematopoietic toxicity caused by IR would benefit acute radiation syndrome (ARS) victims and patients receiving radiotherapy. Herein, we describe the preparation of an injectable hydrogel formulation based on Arg-Gly-Asp-alginate (RGD-Alg) and Laponite using a simple mixing method that ensured a slow and sustained release of interleukin-12 (IL-12) (RGD-Alg/Laponite@IL-12). The local administration of RGD-Alg/Laponite@IL-12 increased survival rates and promoted the hematopoietic recovery of mice who had received sublethal-dose irradiation. Local intra-bone marrow (intra-BM) injection of RGD-Alg/Laponite@IL-12 hydrogel effectively stimulated IL12 receptor-phosphoinositide 3-kinase/protein kinase B (IL-12R-PI3K/AKT) signaling axis, which promoted proliferation and hematopoietic growth factors secretion of BM mesenchymal stem/stromal cells. This signaling axis facilitates the repair of the hematopoietic microenvironment and plays a pivotal role in hematopoietic reconstitution. In conclusion, we describe a biomaterial-sustained release of IL-12 for the treatment of irradiated hematopoietic injury and provide a new therapeutic strategy for hematopoietic ARS.

Mitigating the Effects of 1-Palmitoyl-2-linoleoyl-3-acetyl-rac-glycerol on Gastrointestinal Acute Radiation Syndrome after Total-Body Irradiation in Mice.

Total-body irradiation (TBI) with gamma rays can damage organisms in various unexpected ways and trigger several organ dysfunction syndromes, such as acute radiation syndrome (ARS). Hematopoietic cells and enterocytes are particularly sensitive to radiation due to their self-renewal ability and rapid division, which leads to hematopoietic ARS (H-ARS) and gastrointestinal ARS (GI-ARS). We previously showed that a lipid-based small molecule, 1-palmitoyl-2-linoleoyl-3-acetyl-rac-glycerol (PLAG), improved 30-day survival and alleviated H-ARS symptoms in BALB/c mice after a lethal dose (LD70/30) of gamma-ray TBI. In this study, we investigated the mitigating effects of PLAG on radiation-induced GI damage that occurs under the same conditions as H-ARS in BALB/c mice. Our study showed that PLAG facilitated the structural restoration of intestinal tissues by increasing villus height, crypt depth, crypt number, mucin-producing goblet cells, and proliferating cell nuclear antigen (PCNA)-positive crypt cells. PLAG significantly improved intestinal absorptive capacity and reduced intestinal injury-induced bacterial translocation. In addition, PLAG effectively inhibited radiation-induced necroptosis signaling activation in the intestinal crypt cells, which was responsible for sustained tissue damage and the release of high mobility group box 1 (HMGB1), a typical damage-associated molecular pattern. Overall, our findings support the radiation-mitigating potential of PLAG against GI-ARS after accidental radiation exposure.

Anti-Ceramide ScFv Prophylaxis for First Responders to a Limited Nuclear Attack.

After 9/11, multiple government agencies instituted programs aimed at developing medical radiation countermeasures (MRCs) for two syndromes lethal within weeks of a limited nuclear attack; the hematopoietic-acute radiation syndrome (H-ARS) and the higher-dose gastrointestinal-acute radiation syndrome (GI-ARS). While re-purposing drugs that enhance marrow repopulation treats H-ARS, no mitigator protects GI tract. We recently reported anti-ceramide 6B5 single-chain variable fragment (scFv) pre-treatment abrogates ongoing small intestinal endothelial apoptosis to rescue Lgr5+ stem cells, preventing GI-ARS lethality in C57B/L6J mice. Here, with US Department of Defense support, we provide evidence that humanized anti-ceramide scFv (CX-01) is a promising prophylactic MRC for first responders, who risk exposure upon entering a radiation-contaminated site. CX-01, when delivered up to 90 min before irradiation, is highly-effective in preventing small intestinal endothelial apoptosis in mice and lethality in both sexes. Unexpectedly, females require an ~2-fold higher CX-01 dose than males for full protection. CX-01 is effective subcutaneously and intramuscularly, a property critical for battlefield use. Increasing the maximally-effective dose 5-fold does not extend duration of bioeffectiveness. While CX-01 prevents GI-ARS lethality, structural modification to extend half-life may be necessary to optimize first responder prophylaxis.

BIO 300: A Prophylactic Radiation Countermeasure for Acute Radiation Syndrome.

Exposure to high doses of ionizing radiation can result in hematopoietic acute radiation syndrome. Currently, there is no radiation medical countermeasure approved by the U.S. FDA which can be used before radiation exposure to protect exposed individuals. Here we aimed to evaluate the therapeutic potential of an aqueous suspension of synthetic genistein nanoparticles (BIO 300) as a radioprotectant in a pilot efficacy study using a nonhuman primate model of total body irradiation. Eight rhesus macaques were divided into two groups; four received vehicle and four received BIO 300 Injectable Suspension 24 h before 5.8 Gy total-body irradiation. Survival, blood cell counts, blood chemistry, and clinical parameters were monitored over the 60 days of the study. Tissues were collected at necropsy 60 days post-irradiation or from animals that met unscheduled euthanasia criteria and subjected to histopathological analysis. Tissues analyzed included the duodenum, jejunum, ileum, sternum, lung, heart, liver, kidney, spleen, gut-associated lymphoid tissue, and urinary bladder. In this pilot study, all BIO 300 Injectable Suspension treated animals survived to day 60, while only 50% of the vehicle-treated animals survived. We found that BIO 300 Injectable Suspension did not mediate an improvement in blood cell counts (e.g., neutrophils, platelets, white blood cells). However, BIO 300 Injectable Suspension treated animals had a lower incidence of fever and febrile neutropenia, were able to better maintain their body weight post radiation exposure, and exhibited less anemia and faster recovery from anemia. Histopathological analysis revealed that BIO 300-treated animals had less irradiation-induced damage to the sternum and other tissues compared to vehicle controls. BIO 300's mechanism of action is complex and protection against irradiation is attainable without much improvement in the complete blood count (CBC) profile. BIO 300's mechanism for radioprotection involves multiple biological pathways and systems.

Biomarkers for Radiation Biodosimetry and Correlation with Hematopoietic Injury in a Humanized Mouse Model.

After a large-scale radiological or nuclear event, hundreds of thousands of people may be exposed to ionizing radiation and require subsequent medical management. Acute exposure to moderate doses (2-6 Gy) of radiation can lead to the hematopoietic acute radiation syndrome, in which the bone marrow (BM) is severely compromised, and severe hemorrhage and infection are common. Previously, we have developed a panel of intracellular protein markers (FDXR, ACTN1, DDB2, BAX, p53 and TSPYL2), designed to reconstruct absorbed radiation dose from human peripheral blood (PB) leukocyte samples in humanized mice up to 3 days after exposure. The objective of this work was to continue to use the humanized mouse model to evaluate biomarker dose-/time- kinetics in human PB leukocytes invivo, at an earlier (day 2) and later (day 7) time point, after exposure to total-body irradiation (TBI) doses of 0 to 2 Gy of X rays. In addition, to assess hematological sensitivity and radiation-induced injury, PB leukocyte cell counts, human BM hematopoietic stem cell (HSC) and progenitor cell [multipotent progenitor (MPP), common myeloid progenitor (CMP), granulocyte myeloid progenitor (GMP), megakaryocyte/erythrocyte progenitor (MEP) and multi-lymphoid progenitor (MLP)] levels were measured, and their correlation was also examined as the BM damages are difficult to assess by routine tests. Peripheral blood B-cells were significantly lower after TBI doses of 0.5 Gy on day 2 and 2 Gy on days 2 and 7; T-cells were significantly reduced only on day 2 after 2 Gy TBI. Bone marrow HSCs and MPP cells showed a dose-dependent depletion after irradiation with 0.5 Gy and 2 Gy on day 2, and after 1 Gy and 2 Gy on day 7. Circulating B cells correlated with HSCs, MPP and MLP cells on day 2, whereas T cells correlated with MPP, and myeloid cells correlated with MLP cells. On day 7, B cells correlated with MPP, CMP, GMP and MEP, while myeloid cells correlated with CMP, GMP and MEP. The intracellular leukocyte biomarkers were able to discriminate unirradiated and irradiated samples at different time points calculated by receiver operating characteristic (ROC) curve. Using machine learning algorithm methods, combining ACTN1, p53, TSPYL2 and PB-T cell and PB-B cell counts served as a strong predictor (area under the ROC >0.8) to distinguish unirradiated and irradiated samples independent of the days after TBI. The results further validated our biomarker-based triage assay and additionally evaluated the radiation sensitivity of the hematopoietic system after TBI exposures.

An Overview of Radiation Countermeasure Development in Radiation Research from 1954 to 2024.

Preparation for medical responses to major radiation accidents, further driven by increases in the threat of nuclear warfare, has led to a pressing need to understand the underlying mechanisms of radiation injury (RI) alone or in combination with other trauma (combined injury, CI). The identification of these mechanisms suggests molecules and signaling pathways that can be targeted to develop radiation medical countermeasures. Thus far, the United States Food and Drug Administration (U.S. FDA) has approved seven countermeasures to mitigate hematopoietic acute radiation syndrome (H-ARS), but no drugs are available for prophylaxis and no agents have been approved to combat the other sub-syndromes of ARS, let alone delayed effects of acute radiation exposure or the effects of combined injury. From its inception, Radiation Research has significantly contributed to the understanding of the underlying mechanisms of radiation injury and combined injury, and to the development of radiation medical countermeasures for these indications through the publication of peer-reviewed research and review articles.

Cell Therapies for Acute Radiation Syndrome.

The risks of severe ionizing radiation exposure are increasing due to the involvement of nuclear powers in combat operations, the increasing use of nuclear power, and the existence of terrorist threats. Exposure to a whole-body radiation dose above about 0.7 Gy results in H-ARS (hematopoietic acute radiation syndrome), which is characterized by damage to the hematopoietic system; higher doses result in further damage to the gastrointestinal and nervous systems. Only a few medical countermeasures for ARS are currently available and approved for use, although others are in development. Cell therapies (cells or products produced by cells) are complex therapeutics that show promise for the treatment of radiation injury and have been shown to reduce mortality and morbidity in animal models. Since clinical trials for ARS cannot be ethically conducted, animal testing is extremely important. Here, we describe cell therapies that have been tested in animal models. Both cells and cell products appear to promote survival and lessen tissue damage after whole-body irradiation, although the mechanisms are not clear. Because radiation exposure often occurs in conjunction with other traumatic injuries, animal models of combined injury involving radiation and future countermeasure testing for these complex medical problems are also discussed.

Effects of radiation mitigating amino acid mixture on mice of different sexes.

To date, few FDA-approved medical countermeasures are available for addressing hematopoietic acute radiation syndrome (H-ARS). In this study, we present our latest research findings focusing on the evaluation of a novel radiation mitigator known as the mitigating amino acid mixture (MAAM). MAAM is composed of five amino acids as the recently reported amino acid-based oral rehydration solution for mitigating gastrointestinal (GI)-ARS. CD2F1 male and female mice were exposed to 60Co-γ total body irradiation (TBI) at 9.0 or 9.5 Gy. Following irradiation, mice were orally administered with MAAM or a saline vehicle control once daily for a duration of 14 days, commencing 24 h after TBI. Mouse survival and body weight change were monitored for 30 days after irradiation. Complete blood counts (CBCs), bone marrow (BM) stem and progenitor cell survival (clonogenicity), and a serum cytokine antibody array were analyzed using samples from day 30 surviving mice. Our data revealed that MAAM treatment significantly enhanced survival rates in irradiated male CD2F1 mice, and the survival rate increased from 25% in the vehicle control group to 60% in the MAAM-treated group (p < 0.05) after 9.0 Gy TBI. The number of BM colonies significantly increased from 41.8 ± 6.4 /104 cells (in the vehicle group) to 78.5 ± 17.0 /104 cells (in the MAAM group) following 9.0 Gy TBI. Furthermore, MAAM treatment led to a decrease in the levels of six cytokines/proteins [cluster of differentiation 40 (CD40), interleukin (IL)-17A, C-X-C motif chemokine 10 (CXCL10/CRG-2), cutaneous T cell-attracting chemokine (CTACK), macrophage inflammatory protein (MIP)-3β, and IL-1β] and an increase in the levels of five other cytokines/proteins [IL-3Rβ, IL-5, leptin, IL-6, and stem cell factor (SCF)] in mouse serum compared to the vehicle group after 9.0 Gy TBI. However, similar alleviating effects of MAAM were not observed in the irradiated CD2F1 female mice. The serum cytokine profile in the irradiated female mice was different compared to the irradiated male mice. In summary, our data suggest that the beneficial effects of the mitigative amino acid combination treatment after radiation exposure may depend on sex.

Pharmacokinetic and Metabolomic Studies with a Promising Radiation Countermeasure, BBT-059 (PEGylated interleukin-11), in Rhesus Nonhuman Primates.

BBT-059, a long-acting PEGylated interleukin-11 (IL-11) analog that is believed to have hematopoietic promoting and anti-apoptotic properties, is being developed as a potential radiation medical countermeasure (MCM) for hematopoietic acute radiation syndrome (H-ARS). This agent has been shown to improve survival in lethally irradiated mice. To further evaluate the drug's toxicity and safety profile, 12 naïve nonhuman primates (NHPs, rhesus macaques) were administered one of three doses of BBT-059 subcutaneously and were monitored for the next 21 days. Blood samples were collected throughout the study to assess the pharmacokinetics (PK) and pharmacodynamics (PD) of the drug as well as its effects on complete blood counts, cytokines, vital signs, and to conduct metabolomic studies. No adverse effects were detected in any treatment group during the study. Short-term changes in metabolomic profiles were present in all groups treated with BBT-059 beginning immediately after drug administration and reverting to near normal levels by the end of the study period. Several pathways and metabolites, particularly those related to inflammation and steroid hormone biosynthesis, were activated by BBT-059 administration. Taken together, these observations suggest that BBT-059 has a good safety profile for further development as a radiation MCM for regulatory approval for human use.

Ferroptosis, Inflammation, and Microbiome Alterations in the Intestine in the Göttingen Minipig Model of Hematopoietic-Acute Radiation Syndrome.

Hematopoietic acute radiation syndrome (H-ARS) involves injury to multiple organ systems following total body irradiation (TBI). Our laboratory demonstrated that captopril, an angiotensin-converting enzyme inhibitor, mitigates H-ARS in Göttingen minipigs, with improved survival and hematopoietic recovery, as well as the suppression of acute inflammation. However, the effects of captopril on the gastrointestinal (GI) system after TBI are not well known. We used a Göttingen minipig H-ARS model to investigate captopril's effects on the GI following TBI (60Co 1.79 or 1.80 Gy, 0.42-0.48 Gy/min), with endpoints at 6 or 35 days. The vehicle or captopril (0.96 mg/kg) was administered orally twice daily for 12 days, starting 4 h post-irradiation. Ilea were harvested for histological, protein, and RNA analyses. TBI increased congestion and mucosa erosion and hemorrhage, which were modulated by captopril. GPX-4 and SLC7A11 were downregulated post-irradiation, consistent with ferroptosis at 6 and 35 days post-irradiation in all groups. Interestingly, p21/waf1 increased at 6 days in vehicle-treated but not captopril-treated animals. An RT-qPCR analysis showed that radiation increased the gene expression of inflammatory cytokines IL1B, TNFA, CCL2, IL18, and CXCL8, and the inflammasome component NLRP3. Captopril suppressed radiation-induced IL1B and TNFA. Rectal microbiome analysis showed that 1 day of captopril treatment with radiation decreased overall diversity, with increased Proteobacteria phyla and Escherichia genera. By 6 days, captopril increased the relative abundance of Enterococcus, previously associated with improved H-ARS survival in mice. Our data suggest that captopril mitigates senescence, some inflammation, and microbiome alterations, but not ferroptosis markers in the intestine following TBI.

Large-scale bioreactor production of extracellular vesicles from mesenchymal stromal cells for treatment of acute radiation syndrome

BackgroundHematopoietic acute radiation syndrome (H-ARS) occurring after exposure to ionizing radiation damages bone marrow causing cytopenias, increasing susceptibility to infections and death. We and others have shown that cellular therapies like human mesenchymal stromal cells (MSCs), or monocytes/macrophages educated ex-vivo with extracellular vesicles (EVs) from MSCs were effective in a lethal H-ARS mouse model. However, given the complexity of generating cellular therapies and the potential risks of using allogeneic products, development of an “off-the-shelf” cell-free alternative like EVs may have utility in conditions like H-ARS that require rapid deployment of available therapeutics. The purpose of this study was to determine the feasibility of producing MSC-derived EVs at large scale using a bioreactor and assess critical quality control attributes like identity, sterility, and potency in educating monocytes and promoting survival in a lethal H-ARS mouse model.MethodsEVs were isolated by ultracentrifugation from unprimed and lipopolysaccharide (LPS)-primed MSCs grown at large scale using a hollow fiber bioreactor and compared to a small scale system using flasks. The physical identity of EVs included a time course assessment of particle diameter, yield, protein content and surface marker profile by flow-cytometry. Comparison of the RNA cargo in EVs was determined by RNA-seq. Capacity of EVs to generate exosome educated monocytes (EEMos) was determined by qPCR and flow cytometry, and potency was assessed in vivo using a lethal ARS model with NSG mice.ResultsPhysical identity of EVs at both scales were similar but yields by volume were up to 38-fold more using a large-scale bioreactor system. RNA-seq indicated that flask EVs showed upregulated let-7 family and miR-143 micro-RNAs. EEMos educated with LPS-EVs at each scale were similar, showing increased gene expression of IL-6, IDO, FGF-2, IL-7, IL-10, and IL-15 and immunophenotyping consistent with a PD-L1 high, CD16 low, and CD86 low cell surface expression. Treatment with LPS-EVs manufactured at both scales were effective in the ARS model, improving survival and clinical scores through improved hematopoietic recovery. EVs from unprimed MSCs were less effective than LPS-EVs, with flask EVs providing some improved survival while bioreactor EVs provide no survival benefit.ConclusionsLPS-EVs as an effective treatment for H-ARS can be produced using a scale-up development manufacturing process, representing an attractive off-the-shelf, cell-free therapy.

Delayed effects of radiation exposure in a C57L/J mouse model of partial body irradiation with ~2.5% bone marrow shielding.

Mouse models of radiation injury are critical to the development of medical countermeasures (MCMs) against radiation. Now that MCMs against hematopoietic acute radiation syndrome (H-ARS) have achieved regulatory approval, attention is shifting to develop MCMs against the adverse effects of gastrointestinal acute radiation syndrome (GI-ARS) and delayed effects of acute radiation exposure (DEARE). The C57L/J mouse model of partial body irradiation (PBI) with 2.5% bone marrow shielding (BM2.5) is being leveraged to examine both GI-ARS and DEARE effects. Within days of PBI, mice may develop H- and GI-ARS followed several months later by DEARE as a multi-organ injury, which typically involves the lung and kidney (L- and K-DEARE, respectively). The objective of this manuscript is to describe the dose response relationship and progression of radiation injury in the C57L/J mouse and to evaluate its suitability for use in DEARE MCM testing. In two separate studies conducted over 2 years, male and female C57L/J mice were exposed to PBI BM2.5 with one hindlimb shielded from radiation, representing ~2.5% bone marrow shielding/sparing. Mice were X-ray irradiated at doses ranging from 9 to 13 Gy at 10 to 12 weeks of age for the purposes of assessing ARS survival at 30 days and DEARE survival at 182 days post-irradiation. Clinical indicators of ARS and DEARE were determined by clinical observations, body weights, hematology, clinical chemistry, magnetic resonance imaging (MRI) of lung, and histopathology of selected tissues. C57L/J mice developed canonical ARS responses of hematopoietic atrophy and gastrointestinal injury resulting in dose dependent mortality at doses ≥11 Gy between 1- and 15-days post-irradiation. In animals that survived ARS, DEARE associated mortality occurred in dose dependent fashion at ≥9 Gy for both sexes between 60- and 159-days post-irradiation with histopathology examinations indicating lung injury as the primary cause of death in moribund animals. The PBI BM2.5 C57L/J mouse model reliably produced known H- and GI-ARS effects at doses greater than those resulting in DEARE effects. Because of this, the C57L/J mouse can be used to test MCMs against L-DEARE injury, while avoiding ARS associated mortality.

Effects of Hemorrhage on Hematopoietic Cell Depletion after a Combined Injury with Radiation: Role of White Blood Cells and Red Blood Cells as Biomarkers.

Combined radiation with hemorrhage (combined injury, CI) exacerbates hematopoietic acute radiation syndrome and mortality compared to radiation alone (RI). We evaluated the effects of RI or CI on blood cell depletion as a biomarker to differentiate the two. Male CD2F1 mice were exposed to 8.75 Gy γ-radiation (60Co). Within 2 h of RI, animals were bled under anesthesia 0% (RI) or 20% (CI) of total blood volume. Blood samples were collected at 4-5 h and days 1, 2, 3, 7, and 15 after RI. CI decreased WBC at 4-5 h and continued to decrease it until day 3; counts then stayed at the nadir up to day 15. CI decreased neutrophils, lymphocytes, monocytes, eosinophils, and basophils more than RI on day 1 or day 2. CI decreased RBCs, hemoglobin, and hematocrit on days 7 and 15 more than RI, whereas hemorrhage alone returned to the baseline on days 7 and 15. RBCs depleted after CI faster than post-RI. Hemorrhage alone increased platelet counts on days 2, 3, and 7, which returned to the baseline on day 15. Our data suggest that WBC depletion may be a potential biomarker within 2 days post-RI and post-CI and RBC depletion after 3 days post-RI and post-CI. For hemorrhage alone, neutrophil counts at 4-5 h and platelets for day 2 through day 7 can be used as a tool for confirmation.

FSL-1: A Synthetic Peptide Increases Survival in a Murine Model of Hematopoietic Acute Radiation Syndrome.

In the current geopolitical climate there is an unmet need to identify and develop prophylactic radiation countermeasures, particularly to ensure the well-being of warfighters and first responders that may be required to perform on radiation-contaminated fields for operational or rescue missions. Currently, no countermeasures have been approved by the U.S. FDA for prophylactic administration. Here we report on the efficacious nature of FSL-1 (toll-like receptor 2/6 agonist) and the protection from acute radiation syndrome (ARS) in a murine total-body irradiation (TBI) model. A single dose of FSL-1 was administered subcutaneously in mice. The safety of the compound was assessed in non-irradiated animals, the efficacy of the compound was assessed in animals exposed to TBI in the AFRRI Co-60 facility, the dose of FSL-1 was optimized, and common hematological parameters [complete blood cell (CBC), cytokines, and bone marrow progenitor cells] were assessed. Animals were monitored up to 60 days after exposure and radiation-induced damage was evaluated. FSL-1 was shown to be non-toxic when administered to non-irradiated mice at doses up to 3 mg/kg. The window of efficacy was determined to be 24 h prior to 24 h after TBI. FSL-1 administration resulted in significantly increased survival when administered either 24 h prior to or 24 h after exposure to supralethal doses of TBI. The optimal dose of FSL-1 administration was determined to be 1.5 mg/kg when administered prior to irradiation. Finally, FSL-1 protected the hematopoietic system (recovery of CBC and bone marrow CFU). Taken together, the effects of increased survival and accelerated recovery of hematological parameters suggests that FSL-1 should be developed as a novel radiation countermeasure for soldiers and civilians, which can be used either before or after irradiation in the aftermath of a radiological or nuclear event.

Sequential administration of delta-tocotrienol ameliorates radiation-induced myelosuppression in mice and non-human primates through inducing G-CSF production

To date only four recombinant growth factors, including Filgrastim (rhG-CSF), have been approved by FDA as radiomitigators to ameliorate hematopoietic acute radiation syndrome (H-ARS). These approved agents are not stable under room-temperature, needing to be stored at 2–8 °C, and would not be feasible in a mass casualty scenario where rapid and cost-effective intervention is crucial. Delta-tocotrienol (δ-T3H), the most potent G–CSF–inducing agent among vitamin E isoforms, exhibited efficiency and selectivity on G-CSF production in comparison with TLR and STING agonists in mice. Five-dose δ-T3H was utilized as the optimal therapeutic regimen due to long-term G-CSF production and the best peripheral blood (PB) recovery of irradiated mice. Comparable with rhG-CSF, sequential administration of δ-T3H post-irradiation improved hematologic recovery and accelerated the regeneration of hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs) in the bone marrow (BM) and spleen of 6.5Gy irradiated mice; and consistently enhanced repopulation of BM-HSCs. In 4.0Gy irradiated nonhuman primates, δ-T3H exhibited comparable efficacy as rhG-CSF to promote PB recovery and colony-formation of BM-HPCs. Altogether, we demonstrated that sequential administration of delta-tocotrienol ameliorates radiation-induced myelosuppression in mice and non-human primates through inducing G-CSF production, indicated δ-T3H as a promising radiomitigator for the management of H-ARS, particularly in a mass casualty scenario.

Fluacrypyrim Protects Hematopoietic Stem and Progenitor Cells against Irradiation via Apoptosis Prevention.

Ionizing radiation (IR)-induced hematopoietic injury has become a global concern in the past decade. The underlying cause of this condition is a compromised hematopoietic reserve, and this kind of hematopoietic injury could result in infection or bleeding, in addition to lethal mishaps. Therefore, developing an effective treatment for this condition is imperative. Fluacrypyrim (FAPM) is a recognized effective inhibitor of STAT3, which exhibits anti-inflammation and anti-tumor effects in hematopoietic disorders. In this context, the present study aimed to determine whether FAPM could serve as a curative agent in hematopoietic-acute radiation syndrome (H-ARS) after total body irradiation (TBI). The results revealed that the peritoneally injection of FAPM could effectively promote mice survival after lethal dose irradiation. In addition, promising recovery of peripheral blood, bone marrow (BM) cell counts, hematopoietic stem cell (HSC) cellularity, BM colony-forming ability, and HSC reconstituting ability upon FAPM treatment after sublethal dose irradiation was noted. Furthermore, FAPM could reduce IR-induced apoptosis in hematopoietic stem and progenitor cells (HSPCs) both in vitro and in vivo. Specifically, FAPM could downregulate the expressions of p53-PUMA pathway target genes, such as Puma, Bax, and Noxa. These results suggested that FAPM played a protective role in IR-induced hematopoietic damage and that the possible underlying mechanism was the modulation of apoptotic activities in HSCs.

DAMPs and radiation injury.

The heightened risk of ionizing radiation exposure, stemming from radiation accidents and potential acts of terrorism, has spurred growing interests in devising effective countermeasures against radiation injury. High-dose ionizing radiation exposure triggers acute radiation syndrome (ARS), manifesting as hematopoietic, gastrointestinal, and neurovascular ARS. Hematopoietic ARS typically presents with neutropenia and thrombocytopenia, while gastrointestinal ARS results in intestinal mucosal injury, often culminating in lethal sepsis and gastrointestinal bleeding. This deleterious impact can be attributed to radiation-induced DNA damage and oxidative stress, leading to various forms of cell death, such as apoptosis, necrosis and ferroptosis. Damage-associated molecular patterns (DAMPs) are intrinsic molecules released by cells undergoing injury or in the process of dying, either through passive or active pathways. These molecules then interact with pattern recognition receptors, triggering inflammatory responses. Such a cascade of events ultimately results in further tissue and organ damage, contributing to the elevated mortality rate. Notably, infection and sepsis often develop in ARS cases, further increasing the release of DAMPs. Given that lethal sepsis stands as a major contributor to the mortality in ARS, DAMPs hold the potential to function as mediators, exacerbating radiation-induced organ injury and consequently worsening overall survival. This review describes the intricate mechanisms underlying radiation-induced release of DAMPs. Furthermore, it discusses the detrimental effects of DAMPs on the immune system and explores potential DAMP-targeting therapeutic strategies to alleviate radiation-induced injury.

Effects of combined ciprofloxacin and Neulasta therapy on intestinal pathology and gut microbiota after high-dose irradiation in mice.

Treatments that currently exist in the strategic national stockpile for acute radiation syndrome (ARS) focus on the hematopoietic subsyndrome, with no treatments on gastrointestinal (GI)-ARS. While the gut microbiota helps maintain host homeostasis by mediating GI epithelial and mucosal integrity, radiation exposure can alter gut commensal microbiota which may leave the host susceptible to opportunistic pathogens and serious sequelae such as sepsis. To mitigate the effects of hematopoietic ARS irradiation, currently approved treatments exist in the form of colony stimulating factors and antibiotics: however, there are few studies examining how these therapeutics affect GI-ARS and the gut microbiota. The aim of our study was to examine the longitudinal effects of Neulasta and/or ciprofloxacin treatment on the gut microbiota after exposure to 9.5 Gy 60Co gamma-radiation in mice. The gut microbiota of vehicle and drug-treated mice exposed to sham or gamma-radiation was characterized by shotgun sequencing with alpha diversity, beta diversity, and taxonomy analyzed on days 2, 4, 9, and 15 post-irradiation. No significant alpha diversity differences were observed following radiation, while beta diversity shifts and taxonomic profiles revealed significant alterations in Akkermansia, Bacteroides, and Lactobacillus. Ciprofloxacin generally led to lower Shannon diversity and Bacteroides prevalence with increases in Akkermansia and Lactobacillus compared to vehicle treated and irradiated mice. While Neulasta increased Shannon diversity and by day 9 had more similar taxonomic profiles to sham than ciprofloxacin-or vehicle-treated irradiated animals. Combined therapy of Neulasta and ciprofloxacin induced a decrease in Shannon diversity and resulted in unique taxonomic profiles early post-irradiation, returning closer to vehicle-treated levels over time, but persistent increases in Akkermansia and Bacteroides compared to Neulasta alone. This study provides a framework for the identification of microbial elements that may influence radiosensitivity, biodosimetry and the efficacy of potential therapeutics. Moreover, increased survival from H-ARS using these therapeutics may affect the symptoms and appearance of what may have been subclinical GI-ARS.

The TLR2/TLR6 ligand FSL-1 mitigates radiation-induced hematopoietic injury in mice and nonhuman primates.

Thrombocytopenia, hemorrhage, anemia, and infection are life-threatening issues following accidental or intentional radiation exposure. Since few therapeutics are available, safe and efficacious small molecules to mitigate radiation-induced injury need to be developed. Our previous study showed the synthetic TLR2/TLR6 ligand fibroblast stimulating lipopeptide (FSL-1) prolonged survival and provided MyD88-dependent mitigation of hematopoietic acute radiation syndrome (H-ARS) in mice. Although mice and humans differ in TLR number, expression, and function, nonhuman primate (NHP) TLRs are like those of humans; therefore, studying both animal models is critical for drug development. The objectives of this study were to determine the efficacy of FSL-1 on hematopoietic recovery in small and large animal models subjected to sublethal total body irradiation and investigate its mechanism of action. In mice, we demonstrate a lack of adverse effects, an easy route of delivery (subcutaneous) and efficacy in promoting hematopoietic progenitor cell proliferation by FSL-1. NHP given radiation, followed a day later with a single subcutaneous administration of FSL-1, displayed no adversity but showed elevated hematopoietic cells. Our analyses revealed that FSL-1 promoted red blood cell development and induced soluble effectors following radiation exposure. Cytologic analysis of bone marrow aspirates revealed a striking enhancement of mononuclear progenitor cells in FSL-1-treated NHP. Combining the efficacy of FSL-1 in promoting hematopoietic cell recovery with the lack of adverse effects induced by a single administration supports the application of FSL-1 as a viable countermeasure against H-ARS.

Pharmacokinetics and Biodistribution of 16,16 dimethyl Prostaglandin E2 in Non-Irradiated and Irradiated Mice and Non-Irradiated Non-Human Primates.

Exposure to high-dose ionizing radiation can lead to life-threatening injuries and mortality. Bone marrow is the most sensitive organ to radiation damage, resulting in the hematopoietic acute radiation syndrome (H-ARS) with the potential sequelae of infection, hemorrhage, anemia, and death if untreated. The development of medical countermeasures (MCMs) to protect or mitigate radiation injury is a medical necessity. In our well-established murine model of H-ARS we have demonstrated that the prostaglandin E2 (PGE2) analog 16,16 dimethyl-PGE2 (dmPGE2) has survival efficacy as both a radioprotectant and radiomitigator. The purpose of this study was to investigate the pharmacokinetics (PK) and biodistribution of dmPGE2 when used as a radioprotector in irradiated and non-irradiated inbred C57BL/6J mice, PK in irradiated and non-irradiated Jackson Diversity Outbred (JDO) mice, and the PK profile of dmPGE2 in non-irradiated non-human primates (NHPs). The C57BL/6J and JDO mice each received a single subcutaneous (SC) dose of 35 ug of dmPGE2 and were randomized to either receive radiation 30 min later or remain non-irradiated. Plasma and tissue PK profiles were established. The NHP were dosed with 0.1 mg/kg by SC administration and the PK profile in plasma was established. The concentration time profiles were analyzed by standard non-compartmental analysis and the metrics of AUC0-Inf, AUC60-480 (AUC from 60-480 min), Cmax, and t1/2 were evaluated. AUC60-480 represents the postirradiation time frame and was used to assess radiation effect. Overall, AUC0-Inf, Cmax, and t1/2 were numerically similar between strains (C57BL/6J and JDO) when combined, regardless of exposure status (AUC0-Inf: 112.50 ng·h/ml and 114.48 ng·h/ml, Cmax: 44.53 ng/ml and 63.96 ng/ml; t1/2: 1.8 h and 1.1 h, respectively). PK metrics were numerically lower in irradiated C57BL/6J mice than in non-irradiated mice [irradiation ratio: irradiated values/non-irradiated values = 0.71 for AUC60-480 (i.e., 29% lower), and 0.6 for t1/2]. In JDO mice, the radiation ratio was 0.53 for AUC60-480 (i.e., 47% lower), and 1.7 h for t1/2. The AUC0-Inf, Cmax, and t1/2 of the NHPs were 29.20 ng·h/ml, 7.68 ng/ml, and 3.26 h, respectively. Despite the numerical differences seen between irradiated and non-irradiated groups in PK parameters, the effect of radiation on PK can be considered minimal based on current data. The biodistribution in C57BL/6J mice showed that dmPGE2 per gram of tissue was highest in the lungs, regardless of exposure status. The radiation ratio for the different tissue AUC60-480 in C57BL/6J mice ranged between 0.5-1.1 (50% lower to 10% higher). Spleen, liver and bone marrow showed close to twice lower exposures after irradiation, whereas heart had a 10% higher exposure. Based on the clearance values from mice and NHP, the estimated allometric scaling coefficient was 0.81 (95% CI: 0.75, 0.86). While slightly higher than the current literature estimates of 0.75, this scaling coefficient can be considered a reasonable estimate and can be used to scale dmPGE2 dosing from animals to humans for future trials.