Exposure of healthy tissue to ionizing radiation (IR) occurs due to nuclear accidents and terrorism, as well as radiotherapy. The vascular endothelium is a key target of IR, and microvascular endothelial cells (ECs) are particularly vulnerable to radiation. IR induces EC activation leading to endothelial cell injury. Human ghrelin is a stomach-derived peptide with pleiotropic effects, including protection against inflammation. We hypothesize that human ghrelin improves survival in total body irradiation (TBI) and that ghrelin’s protective effect could be mediated by attenuating endothelial cell injury. To test this, mice were exposed to TBI and after 24 h were treated subcutaneously with human ghrelin once daily for 4 days and monitored for 30 days. The survival rate of the human ghrelin-treated group was significantly higher than that of the vehicle group. Subsequently, human ghrelin treatment showed an effective dose modification factor of 1.0681. On day 4 after TBI, human ghrelin significantly attenuated EC permeability in the lungs and improved tight junction protein ZO-1 expression. Human ghrelin also improved ZO-1 and Claudin5 expression in primary mouse lung vascular endothelial cells. Taken together, these results indicate that human ghrelin improves survival after TBI, and its survival benefit is in part due to the attenuation of EC permeability and microvascular barrier dysfunction.

Acute kidney injury (AKI) from renal ischemia–reperfusion (IR) injury is a major cause of acute organ dysfunction with significant morbidity and mortality. Critically, up to 50

Background: Radiation-induced sepsis resulting from intestinal inflammation and injury is a severe complication of high-dose radiation exposure. High-dose radiation compromises the integrity of the intestinal lining, causing bacterial translocation, systemic inflammation, and sepsis. Extracellular cold-inducible RNA-binding protein (eCIRP), a damage-associated molecular pattern, plays a pivotal role in the pathogenesis of inflammatory diseases and contributes to organ injury. C23, a small molecular peptide antagonist of eCIRP, has demonstrated anti-inflammatory and organ-protective effects during organ-injury indications. However, its role in radiation-induced inflammation and injury is not known. The objective of this study is to evaluate whether C23 mitigates inflammation and injury, leading to improved survival in a murine model of partial-body irradiation (PBI) combined with sepsis. Methods: Mice were exposed to 10 Gy PBI, and at 48 h, they were subjected to cecal ligation and puncture (CLP), a well-established model of sepsis. C23 (8 mg/kg body weight [BW]) or vehicle (saline) was administered subcutaneously at 24 h and 48 h after PBI. Blood and intestinal tissue samples were collected 20 h after CLP (i.e., 68 h post-PBI) for various analyses. In another set of mice after PBI–sepsis, intestinal permeability was also assessed. In an additional cohort of mice subjected to the same experimental procedure, 10-day survival was monitored. Results: Our findings demonstrate that administration of C23 attenuated systemic inflammatory responses, intestinal permeability, intestinal injury, and apoptosis; increased crypt cell proliferation and improved survival after PBI–sepsis. Conclusions: These results reveal a novel protective role of the eCIRP antagonist, C23, in mitigating PBI–sepsis-induced inflammation and injury and represents a promising therapeutic candidate for the treatment of radiation combined injury with sepsis.

Cranial radiotherapy is associated with neuroinflammation, cognitive dysfunction, and dementia. Tau pathology plays a key role in Alzheimer’s disease (AD) but has not been studied in the context of radiation injury. Extracellular cold-inducible RNA-binding protein (eCIRP) is a novel neuroinflammatory mediator that activates neuronal IL6Rα/p25/Cdk5, leading to tau hyperphosphorylation. We hypothesized that radiation promotes pathological tau phosphorylation via the eCIRP/IL6Rα/p25/Cdk5 pathway, which can be mitigated by the eCIRP inhibitor C23. Adult C57BL/6 and CIRP−/− mice were exposed to a single dose of 10-Gy X-rays at a dose-rate of 1 Gy/min. Hippocampal tissue lysates were analyzed for p25, phosphorylated tau, and CIRP using Western blotting. Cdk5 activity was assessed utilizing a modified Cdk5/p35 kinase enzyme system. Serum and cerebrospinal fluid (CSF) eCIRP was quantified using ELISA. eCIRP (1 µg) was injected intracerebroventricularly (icv) and C23 (8 mg/kg BW) retro-orbitally. N2a and primary neurons were pre-treated with 3 μg/ml IL-6Rα neutralizing Abs or IgG control, or 25 µg/mL C23 and stimulated with up to 2.5 μg/mL eCIRP for 48 h. Irradiation significantly increased hippocampal p25 expression, Cdk5 activity, and tau phosphorylation [AT-8 (Ser202/Thr205/Ser208), and combined Ser199/Ser202 and Ser396 p-tau antibodies] at 48 h post-irradiation. eCIRP increased by 1.2-fold in the serum and 2.6-fold in the CSF, and CIRP increased by 65

Critical care research focuses on life-threatening conditions such as sepsis, trauma, hemorrhage, and burn injury, which account for millions of hospitalizations and hundreds of thousands of deaths annually in the United States alone. Recent policy initiatives by the US Food and Drug Administration and the National Institutes of Health have promoted new approach methodologies, including organoids, organ-on-chip platforms, and computational models, as alternatives to animal research. While new approach methodologies offer valuable tools for mechanistic investigation and screening applications, this review examines whether current new approach methodology technologies can adequately replace animal models in critical care research. Critical illness involves the whole organism, including dynamic organ-organ interactions, immune-microbiome crosstalk, and adaptive systemic feedback loops. By examining major domains in critical care research and targeted organ injuries, it becomes clear that while new approach methodologies excel at interrogating isolated subsystems, they cannot currently replicate integrated physiological responses. Animal models remain essential for questions requiring assessment of multiorgan dysfunction, therapeutic safety evaluation, and clinically relevant disease trajectories. Premature policy shifts away from animal research will impede advances in critical care medicine. This review proposes a "methodological pluralism" approach that integrates new approach methodologies with appropriately designed animal studies through harmonized endpoints and reverse-translation frameworks. Recommendations include continued refinement of animal models to better represent the heterogeneity of human populations, adoption of quality standards for preclinical research, and strategic deployment of both new approach methodologies and animal models based on fit-for-purpose criteria. We believe this approach will satisfy ethical considerations, scientific rigor, and public health needs in critical care research.

Introduction:The mechanism by which extracellular cold-inducible RNA-binding protein (eCIRP) aggravates renal ischemia/reperfusion (RIR) injury leading to acute kidney injury (AKI) is poorly understood. The natural killer group 2D (NKG2D) receptor and its ligand MULT-1 are key immunoregulatory mechanisms promoting responses to damaged and inflamed cells. Methods:We subjected wild-type and CIRP-/- mice to RIR. We then used immunohistochemistry (IHC), flow cytometry, and Western blotting to assess NKG2D and MULT-1 in kidney tissues, macrophages, and renal tubular epithelial cells (RTECs), and ELISA to assess TNFa and IL-6. Results:The expression levels of NKG2D and its ligand MULT-1 were significantly elevated in wild-type mice subjected to RIR compared with sham. In contrast, CIRP-/- mice exhibited markedly reduced expression of both NKG2D and MULT-1 after RIR compared to wild-type mice. In vitro, eCIRP stimulated the expression of NKG2D in peritoneal macrophages and of MULT-1 in RTECs. Treatment of eCIRP-stimulated peritoneal macrophage and RTEC co-cultures with an NKG2D-neutralizing antibody significantly and markedly downregulated supernatant levels of TNFa and IL-6. Discussion:In conclusion, eCIRP induces NKG2D+ macrophages and MULT-1+ RTECs, and their interaction further increases the inflammatory response. Targeting the NKG2D/MULT-1 may reduce RIR-induced inflammation and thus attenuate AKI.

Gastrointestinal acute radiation syndrome (GI-ARS) is a deadly consequence of radiation exposure. We hypothesized that the peptide ghrelin is enteroprotective after radiation injury, and that ghrelin promotes intestinal stem cell regeneration via the vagus nerve. We subjected mice to 12-Gy partial body irradiation (PBI) with 5% bone marrow sparing. Some mice were vagotomized prior to PBI. We then injected the mice with human ghrelin (6 nmol/mouse) or vehicle at 24, 48, and 72 h post-irradiation, and collected blood and tissues at 96 h. PBI caused an 80% reduction in plasma citrulline, 32% shorter villi, 59% fewer crypts, a 14-fold increase in TUNEL+ cells, a 9-fold increase in intestinal permeability (FD4), and 4- to 30-fold increases in bacterial translocation (16S rRNA) to the liver and mesentery. Ghrelin significantly improved all these parameters. Remarkably, vagotomy attenuated ghrelin's protective effects by 22-58%. Mechanistically, ghrelin increased proliferating crypt cells by 2.3-fold, Lgr5+ active stem cells by 2.6-fold, Clu+ revival stem cells by 3.3-fold (immunofluorescence), and Clu mRNA by 1.6-fold compared to PBI alone, and all these effects were significantly diminished by vagotomy. Thus, ghrelin mitigates GI-ARS through vagus nerve-dependent activation of Clu+ revival stem cells and Lgr5+ stem cells, identifying a novel vagal-dependent neuroenteric pathway that regulates intestinal crypt regeneration after radiation injury.

Traumatic brain injury (TBI) frequently precipitates chronic psychological disorder, particularly anxiety, depression, post-traumatic stress disorder (PTSD), and increased suicide risk, that worsen with injury severity yet remain understudied relative to sensorimotor and cognitive deficits. As TBI severity increases, conventional interventions (pharmacotherapy, cognitive-behavioral therapy, exercise) lose efficacy, revealing an urgent need for novel treatments targeting affective and stress-related disorders. Neuromodulatory approaches such as transcranial magnetic stimulation (TMS), transcranial direct current stimulation (tDCS), deep brain stimulation (DBS), photobiomodulation (PBM), vagus nerve stimulation (VNS) and trigeminal nerve stimulation (TNS), offer a mechanism-based strategy to recalibrate dysfunctional neural circuits across the full spectrum of TBI severity. In this review, we present the first comprehensive review of 45 preclinical and clinical studies examining all six modalities in TBI. We delineate intervention-specific mechanisms encompassing enhancement of synaptic plasticity, restoration of large-scale network connectivity, normalization of neurotransmitter homeostasis, augmentation of cerebral perfusion, and attenuation of neuroinflammatory cascades, correlating these neurobiological changes with quantifiable improvements in affective symptoms and stress adaptation. Through rigorous mechanistic-to-clinical mapping, this analysis establishes an evidence-based framework for developing precision neuromodulation protocols and mechanism-targeted pharmacological interventions targeting TBI-associated psychological disorders.

Activation of nuclear factor erythroid 2-related factor 2 (Nrf2) is a powerful neuroprotective strategy, yet clinical translation to conditions of high stress remains limited by the paradoxical depletion of glutathione and increase of reactive oxygen species observed with conventional electrophilic activation. Here, we identify the diving reflex (DR) as a potent, non-electrophilic physiological activator of Nrf2 in the brain that circumvents this potential liability. Using a validated voluntary diving model in rats with acute (single session) and chronic (4-week) paradigms, we show that DR triggers calcitonin gene-related peptide (CGRP) release from trigeminal afferents, which engages parallel KEAP1-dependent (p62/Ser351-mediated autophagic degradation) and KEAP1-independent (AMPK/SIRT1/PI3K) signaling cascades to drive Nrf2 nuclear translocation and phosphorylation across multiple brain regions. Anti-CGRP antibody blockade abolished these effects, establishing CGRP as the essential upstream mediator. Unbiased proteomic profiling of 2388 brain proteins confirmed upregulation of Nrf2-regulated antioxidant enzymes with concurrent downregulation of oxidative stress markers. Critically, DR preserved cellular glutathione pools, elevated the GSH/GSSG ratio, and suppressed lipid peroxidation and protein nitration, a redox-sparing profile divergent from electrophilic activation. DR also exhibited a unique biphasic response: acute exposure produced rapid transcriptional upregulation of antioxidant genes, while chronic exposure generated a sustained extranuclear reserve of phosphorylated Nrf2, conferring long-term resilience. In a vascular dementia model of chronic cerebral hypoperfusion, chronic DR rescued hippocampal Nrf2 depletion, restored glutathione homeostasis, and preserved working memory. These findings establish the DR-CGRP-Nrf2 axis as a druggable, non-electrophilic pathway for neuroprotection that achieves pharmacological-level potency without redox compromise.

Extracellular cold-inducible RNA-binding protein (eCIRP) was discovered as a potent damage-associated molecular pattern (DAMP). It has been shown that eCIRP is linked to various types of programmed cell death and acute inflammation. However, the role of eCIRP in chronic inflammation and renal fibrosis has not been elucidated. Accumulating evidence indicates that renal tubular epithelial cells (RTECs) play a significant role in renal fibrosis. C23, a small molecular peptide inhibitor of eCIRP, has been implicated as a therapeutic agent in the context of acute inflammation and tissue injury. PANoptosis or synchronized cell death is observed as simultaneous triggering of apoptosis, pyroptosis, and necroptosis. However, its role in renal fibrosis is not known. We therefore hypothesize that eCIRP induced-chronic inflammation and injury in RTECs are mediated by PANoptosis and that inhibition of eCIRP by C23 decreases RTEC PANoptosis and attenuates renal injury and fibrosis in a mouse model of unilateral ureter obstruction (UUO) injury. By using primary RTECs, we demonstrated that eCIRP induces inflammatory cytokines, Z-DNA-binding protein-1, and other PANoptosome markers and markers of apoptosis, pyroptosis, and necroptosis. We then substantiated that C23 downregulated proinflammatory cytokines and inhibited PANoptosis in the RTECs. Using the UUO mouse model, we demonstrated renal cell PANoptosis and renal fibrosis 7 days after UUO. Importantly, treatment with C23 effectively inhibited PANoptosis and concurrently ameliorated renal fibrosis. Taken together, eCIRP induces inflammation and PANoptosis in RTECs, whereas C23 inhibits PANoptosis in these cells and attenuates renal fibrosis in UUO mice.NEW & NOTEWORTHY Renal fibrosis is a common pathological manifestation of chronic kidney disease (CKD). Extracellular cold-inducible RNA-binding protein (eCIRP) was discovered as a potent damage-associated molecular pattern (DAMP). eCIRP is linked to various types of programmed cell death. PANoptosis or synchronized cell death is observed as simultaneous triggering of apoptosis, pyroptosis, and necroptosis. Inhibiting eCIRP by C23, a small molecular peptide inhibitor of eCIRP, attenuated PANoptosis and renal fibrosis in CKD.

Gastrointestinal acute radiation syndrome (GI-ARS) is characterized by disruption of the intestinal barrier function, leading to bacterial translocation and sepsis. Intestinal stem cells are highly radiosensitive and dramatically reduced after radiation injury. Clusterin (Clu)-positive revival stem cells contribute to the restoration of intestinal stem cells. Ghrelin, a gastric peptide hormone, has been shown to improve intestinal integrity in models of inflammatory enteropathy. In this study, we investigated the effects of ghrelin on intestinal stem cell recovery and its potential to mitigate radiation-induced intestinal injury. Mice were subjected to 12 Gy partial body irradiation (PBI). Ghrelin at the doses of 2 to 6 nmol per mouse was administered daily for 4 consecutive days, starting at 24 h post-PBI, and survival was monitored for 30 days. To assess intestinal histology, cell proliferation, and intestinal stem cell markers, mice were treated with 6 nmol of ghrelin on days 1, 2, and 3 post-PBI, and on day 4 jejunal samples were collected for qPCR, immunofluorescence, and microcolony assays. Intestinal permeability was assessed in vivo by the leakage of gavage-fed 4-kDa FITC-dextran into the circulation. Ghrelin administration significantly improved 30-day survival rate following 12-Gy PBI in a dose-dependent manner. Treatment with ghrelin restored villus length and enhanced intestinal barrier integrity. Ghrelin also significantly increased the expression of proliferation markers in the jejunum. Microcolony assays revealed that ghrelin reversed the decrease in BrdU-positive cells following PBI. The mRNA and protein expression of intestinal stem cell markers was decreased after PBI but was restored by ghrelin treatment. Finally, ghrelin significantly increased the population of Clu+ population following irradiation. These findings indicate that ghrelin mitigates radiation-induced intestinal injury by promoting the expansion of Clu+ revival stem cells and the recovery of intestinal stem cells. This study highlights the therapeutic potential and identifies the mechanism of action of ghrelin as a medical countermeasure against GI-ARS.

Macrophage extracellular traps (METs) are a poorly understood process beneficial for infection control but detrimental in inflammation, autoimmunity and cancer. Our research shows that viable macrophages release METs even when plasma membrane lysis is blocked. We demonstrate, for the first time, that nuclear DNA is extruded directly into the cytoplasm through Gasdermin D pores on the nuclear envelope. Gasdermin D pore formation was triggered by extracellular cold-inducible RNA-binding protein, which activates the TLR4 signal transduction pathway. This DNA is processed in the cytoplasm, enters the vesicular transport system aided by autophagic flux and the Endosomal Sorting Complex. The DNA then enters the lysosomal compartment, where it undergoes histone 3 citrullination, forms nascent traps containing myeloperoxidase, and is released to the extracellular space. Our study provides valuable insights into vital MET formation and its mechanism that will enable future studies on the role of METs in health and disease.

Ionizing radiation causes immune dysfunction, increasing susceptibility to infection and mortality. Extracellular cold-inducible RNA-binding protein (eCIRP) is released from cells during irradiation. This study investigates how radiation-induced eCIRP release causes macrophage phagocytic dysfunction via ferroptosis, with a focus on the role of mitochondrial dysfunction. Peritoneal macrophages were exposed to 10-Gy irradiation. eCIRP levels in the culture supernatants were assessed post-irradiation by ELISA. Ferroptosis was assessed by measuring lipid peroxidation and glutathione peroxidase 4 (GPX4) expression. Mitochondrial function was assessed using Mito stress assay in a Seahorse metabolic analyzer. Phagocytic activity was quantified by measuring the uptake of pHrodo-labeled E. coli. Our results demonstrated that 10-Gy irradiation induced ferroptosis in peritoneal macrophages. Markers of ferroptosis, lipid peroxidation, were significantly elevated, and GPX4 was significantly downregulated in a time-dependent manner on days 3 and 5 post-irradiation. We unveiled a strong time-dependent correlation between post-irradiation eCIRP release and the increases in ferroptosis and macrophage phagocytic dysfunction at days 3 and 5. Furthermore, radiation-induced eCIRP positively correlated with mitochondrial dysfunction, evidenced by marked reductions in basal and maximal respiration and ATP production, mirroring effects of direct eCIRP treatment. Crucially, the application of MFG-E8-derived oligopeptide 3 (MOP3), a novel opsonic eCIRP inhibitor, effectively cleared eCIRP, restoring mitochondrial function, reducing ferroptosis, and improving phagocytosis in irradiated macrophages. These findings establish that radiation-induced eCIRP release drives mitochondrial dysfunction and ferroptosis, thereby impairing macrophage phagocytosis. Targeting eCIRP offers a promising therapeutic strategy to enhance host defense following radiation exposure.

While A1 astrocytes are reported in Alzheimer’s disease (AD), the underlying molecular mechanisms are complex and remain elusive. Proinflammatory extracellular cold-inducible RNA-binding protein (eCIRP) is released by microglia in response to AD-associated neuronal amyloid-β. eCIRP activates the triggering receptor expressed on myeloid cells-1 (TREM-1). Thus, we hypothesized that increased levels of eCIRP in AD due to inflammatory stress induce A1 via astrocytic TREM-1. ELISA was performed in the AD patients’ cerebrospinal fluid (CSF; in-house Alzheimer’s Disease Center repository) or plasma (Precision Biospecimen Solutions, Bethesda, MD) or hTau.P301S mice (Dr. Michel Goedert, Cambridge, UK) plasma. C8-D1a cells (ATCC) or magnetically purified primary astrocytes, from C57BL/6 mice ± small anti-CIRP peptide M3 or from TREM1-KO mice, were stimulated with eCIRP. C57BL/6 mice were injected intracerebroventricular ( icv ) with eCIRP. Expression of mRNA was quantified by qPCR, protein by immunoblotting and released levels by ELISA or R&D proteome profiler. Student’s t-test was used for two-group analysis, one-way analysis of variance (ANOVA)-SNK test for multigroup analysis, and Spearman’s-Rho analysis for correlations, with significance if p < 0.05. Levels of eCIRP were elevated in the CSF and plasma of AD patients, in hTau.P301S mice, and plasma eCIRP strongly correlated with astrocyte activation marker glial fibrillary acidic protein (GFAP) in AD patients ( Fig. 1 ). eCIRP strongly induced A1 astrocyte-specific genes and astrocyte release of proinflammatory and neurotoxic factors in C8-D1a cells, primary astrocytes and icv eCIRP-injected mice brains. In particular, eCIRP increased Complement 3, an A1 astrocytic marker involved in neuroinflammation-associated neurodegeneration. Primary astrocytes from TREM-1 knockout mice were resistant to eCIRP induction of A1 astrocytes. Moreover, astrocytes exposed to eCIRP had higher TREM-1 total and surface protein levels, increased Syk phosphorylation, and upregulated NFκB mRNA. Importantly, M3 effectively inhibited eCIRP’s induction of A1 astrocytes and their release of proinflammatory and neurotoxic mediators. Collectively, our data provides a strong correlation of eCIRP with astrocyte reactivity in AD patients, identifies a novel molecular mechanism by which eCIRP induces A1 astrocytes via TREM-1 and M3 peptide attenuation of eCIRP-induced A1 astrocytes. These findings point to a novel therapeutic opportunity targeting neurotoxic A1 astrocytes in AD.

Abstract Background Chronic kidney disease (CKD) is a leading cause of death in the United States, and renal fibrosis represents a pathologic hallmark of CKD. Extracellular cold-inducible RNA-binding protein (eCIRP) is a stress response protein involved in acute inflammation, tissue injury and regulated cell death. However, the role of eCIRP in chronic inflammation and tissue injury has not been elucidated. We hypothesize that eCIRP is involved in renal ischemia/reperfusion (RIR)-induced CKD and that C23, an antagonist to eCIRP, is beneficial in attenuating renal fibrosis and ferroptosis in RIR-induced CKD. Methods C57BL/6 (WT) or CIRP−/− mice underwent renal injury with total blockage of blood perfusion by clamping bilateral renal pedicles for 28 min. In the WT mice at the time of reperfusion, they were treated with C23 (8 mg/kg) or vehicle. Blood and kidneys were harvested for further analysis at 21 days thereafter. In a separate cohort, mice underwent bilateral RIR and treatment with C23 or vehicle and were then subjected to left nephrectomy 72 h thereafter. Mice were then monitored for additional 19 days, and glomerular filtration rate (GFR) was assessed using a noninvasive transcutaneous method. Results In the RIR-induced CKD, CIRP−/− mice showed decreased collagen deposition, fibronectin staining, and renal injury as compared to the WT mice. Administration of C23 ameliorated renal fibrosis by decreasing the expression of active TGF-β1, α-SMA, collagen deposition, fibronectin and macrophage infiltration to the kidneys. Furthermore, intervention with C23 significantly decreased renal ferroptosis by reducing iron accumulation, increasing the expression of glutathione peroxidase 4 (GPX4) and lipid peroxidation in the kidneys of RIR-induced CKD mice. Treatment with C23 also attenuated BUN and creatinine. Finally, GFR was significantly decreased in RIR mice with left nephrectomy and C23 treatment partially prevented their decrease. Conclusion Our data show that eCIRP plays an important role in RIR-induced CKD. Treatment with C23 decreased renal inflammation, alleviated chronic renal injury and fibrosis, and inhibited ferroptosis in the RIR-induced CKD mice.

High‑dose ionizing radiation induces multiple types of tissue injuries, including hematopoietic dysfunction characterized by neutropenia. Neutrophil extracellular traps (NETs) released during NETosis may contribute to the neutropenia, and subsequent infection and inflammation. Triggering receptor expressed on myeloid cells‑1 (TREM‑1) is one of receptors responsible for NET formation and extracellular cold‑inducible RNA‑binding protein (eCIRP) is a ligand for the TREM‑1 receptor. The present study aimed to investigate NET formation after exposure to high‑dose ionizing radiation and to explore the underlying role of the eCIRP/TREM‑1 axis as its mechanism. Bone marrow‑derived neutrophils (BMDNs) isolated from C57BL/6 mice were exposed to 5 to 15 Gy irradiation. C57BL/6 wild‑type (WT), CIRP‑/‑ and TREM‑1‑/‑ mice were exposed to 10 Gy total body irradiation (TBI). NET formation was analyzed 24 h after irradiation using flow cytometry and fluorescence microscopy, and also after treatment with eCIRP. TREM‑1 cell surface expression on neutrophils was assessed using flow cytometry. Peptidyl arginine deiminase 4 (PAD4) protein expression levels in BMDNs were evaluated using western blotting. TREM‑1 and PAD4 mRNA expression levels in BMDNs were assessed using reverse transcription‑quantitative PCR. In vitro irradiation of neutrophils resulted in a dose‑dependent increase in NET formation, as assessed using flow cytometry and validated using fluorescence microscopy, which demonstrated the characteristic long extracellular DNA structures of NETs in irradiated neutrophils. The in vivo mouse model of TBI exhibited similar results. Furthermore, TREM‑1 expression in BMDNs was significantly increased after irradiation. Protein and mRNA levels of PAD4 were significantly upregulated after irradiation. The addition of eCIRP to BMDNs further increased NET formation post‑irradiation in vitro. Conversely, knockout of CIRP and TREM‑1 in vivo significantly attenuated radiation‑induced NET formation compared with that of WT mice. High‑dose ionizing radiation induced NET formation through the eCIRP/TREM‑1 pathway and may contribute to early neutropenia post‑irradiation.

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.