[This corrects the article DOI: 10.1016/j.isci.2023.107660.].

Patients with Fanconi anemia (FA) are particularly susceptible to developing squamous cell carcinoma of the head and neck due to impaired DNA repair pathways. However, their hypersensitivity to DNA damaging agents can limit effective treatment with standard radiotherapy due to severe side effects and complications. In pre-clinical models, ultra-rapid FLASH radiotherapy (FLASH) reduces radiation-induced toxicity in normal tissues while maintaining similar tumor control compared to conventional dose rate radiotherapy (CONV). Here, we investigated the safety of FLASH for treatment of the head and neck in a mouse model of FA. 129/Sv wild-type (WT) and Fanca-deficient (Fanca -/-) mice received single-dose oral cavity irradiation with electron beam FLASH or CONV to evaluate radiation-induced toxicity in non-tumor-bearing mice. Fanca WT and Fanca -/- mice were irradiated with 25 and 18 Gy, respectively, of FLASH (190 Gy/sec) or CONV (0.2 Gy/sec), with tongues harvested at 12 hours (hpi) and 10 days (dpi) post-irradiation. At 10 dpi, FLASH-irradiated tongues in both genetic backgrounds demonstrated reduced ulceration at the dorsal tongue surface compared to CONV-irradiated counterparts. Histopathological analysis of the tongue revealed lower mucositis severity scores with decreased epithelial thinning and ulceration in FLASH-irradiated tongues compared to CONV-irradiated ones. Analysis of γ-H2AX foci formation at 12 hpi demonstrated fewer foci in WT mice treated with FLASH compared to CONV, with a similar trend observed in Fanca -/- mice. These findings suggest a potential normal tissue-sparing effect with FLASH and hold important clinical implications for the treatment of patients with Fanconi anemia and head and neck cancers.

Threats of irradiation (IR) exposure increase the need for radiomitigators. An important contributor to radiation injury is ferroptosis, triggered by the disbalanced redox metabolism. We showed that 15-lipoxygenase (15-LOX) catalyzed peroxidation of arachidonoyl-phosphatidyl-ethanolamine is an essential ferroptotic response of ileum to total body IR (TBI). Given that nitric oxide (NO● ) can suppress ferroptosis by inhibiting 15-LOX and by directly scavenging lipid radicals, we tested NO●-donors with optimized half decay times as radiomitigators. Here, we report that diethylenetriamine-NONOate (DETA-NONOate) (with a half decay-time of 20 hr) acted as an effective radiomitigator when administered 24 hr after exposure to TBI (9.25Gy) and markedly prolonged survival of C57BlJ6 mice by - i) decreasing the levels of pro-ferroptotic HOO-PUFA-PE signals, and ii) decreasing the expression of 15-LOX2 – in the ileum on day 4 after TBI. Redox lipidomics LC-MS and two mass spectrometric imaging (MSI) protocols: i) single-cell multi-omics Dual C60/gas cluster ion beam (GCIB) secondary ion mass spectrometry (SIMS), and ii) matrix-assisted laser desorption ionization (MALDI)-MSI, visualized DETA-NONOate's effectiveness in suppressing TBI-induced HOO-PUFA-PE production and preserving intestinal epithelium structural integrity. In vitro, NO● donors were effective in suppressing PUFA-PE peroxidation and ferroptotic death in human intestinal epithelial cells (FHs 74 Int) exposed to radiation (8Gy) plus enzymatic (15-LOX2) pro-ferroptotic stimulation.

Radiation-induced lung injury (RILI) includes early acute phase radiation pneumonitis (RP), and late chronic phase radiation-induced pulmonary fibrosis (RIPF). There is increasing evidence that ionizing radiation-induced cellular senescence is associated with pulmonary fibrosis. We have recently reported that biomarkers of senescence and, specifically, tyrosine kinase Fgr are induced in mouse RIPF, human idiopathic pulmonary fibrosis (IPF), and in human RIPF. We also reported that treatment with an Fgr inhibitor significantly reduced fibrosis of irradiated mouse lungs. Here, we investigated the association of senescence and tyrosine kinase Fgr in non-human primate (NHP) lung fibrosis and determined whether lung fibrosis can be predicted by analyzing the bronchoalveolar lavage (BAL) cells and fluid at early time points after irradiation. We found that markers of senescence (p16, p21) and expression of Fgr are induced in the lungs of NHP with RILI. That fibrosis can be predicted by analyzing BAL cells prior to the appearance of pulmonary fibrosis. We also induced senescence and expression of Fgr in irradiated normal human primary airway epithelial cells in vitro. In a transwell culture system, we established that senescent human airway epithelial cells induced fibrosis biomarkers collagen1, collagen 3, and alpha-smooth-muscle actin in target human primary lung fibroblasts. Whole-thorax lung irradiated (WTLI) NHPs in this study developed moderate to severe pneumonitis and marked variations in the magnitude of RIPF as measured by trichrome staining. In BAL fluid that was collected from WTLI NHP senescence-associated secretory proteins (SASP) were significantly induced, compared to the BAL fluid collected from control non-irradiated NHPs. Moreover, the levels of Fgr and biomarkers of senescence were significantly higher in NHPs with severely injured lungs compared to those with mildly or moderately injured lungs as indicated by fibrosis. Proinflammatory SASP cytokines increased to levels that correlated with the severity of RILI. The results show that senescent cells with induction of Fgr, and SASP cytokines are detectable in NHPs prior to RIPF and suggest that analysis of these proteins can predict the severity of RIPF prior to fully formed fibrosis.

BACKGROUND/AIM:There is concern that people who had COVID-19 will develop pulmonary fibrosis. Using mouse models, we compared pulmonary inflammation following injection of the spike protein of SARS-CoV-2 (COVID-19) to radiation-induced inflammation to demonstrate similarities between the two models. SARS-CoV-2 (COVID-19) induces inflammatory cytokines and stress responses, which are also common to ionizing irradiation-induced acute pulmonary damage. Cellular senescence, which is a late effect following exposure to SARS-CoV-2 as well as radiation, was investigated. MATERIALS AND METHODS:We evaluated the effect of SARS-CoV-2 spike protein compared to ionizing irradiation in K18-hACE2 mouse lung, human lung cell lines, and in freshly explanted human lung. We measured reactive oxygen species, DNA double-strand breaks, stimulation of transforming growth factor-beta pathways, and cellular senescence following exposure to SARS-CoV-2 spike protein, irradiation or SARS-COV-2 and irradiation. We also measured the effects of the antioxidant radiation mitigator MMS350 following irradiation or exposure to SARS-CoV-2. RESULTS:SARS-CoV-2 spike protein induced reactive oxygen species, DNA double-strand breaks, transforming growth factor-β signaling pathways, and senescence, which were exacerbated by prior or subsequent ionizing irradiation. The water-soluble radiation countermeasure, MMS350, reduced spike protein-induced changes. CONCLUSION:In both the SARS-Co-2 and the irradiation mouse models, similar responses were seen indicating that irradiation or exposure to SARS-CoV-2 virus may lead to similar lung diseases such as pulmonary fibrosis. Combination of irradiation and SARS-CoV-2 may result in a more severe case of pulmonary fibrosis. Cellular senescence may explain some of the late effects of exposure to SARS-CoV-2 spike protein and to ionizing irradiation.

Victims of a radiation terrorist event will include pregnant women and unborn fetuses. Mitochondrial dysfunction and oxidative stress are key pathogenic factors of fetal radiation injury. The goal of this preclinical study is to investigate the efficacy of mitigating fetal radiation injury by maternal administration of the mitochondrial-targeted gramicidin S (GS)-nitroxide radiation mitigator JP4-039. Pregnant female C57BL/6NTac mice received 3 Gy total-body irradiation (TBI) at mid-gestation embryonic day 13.5 (E13.5). Using novel time-and-motion-resolved 4D in utero magnetic resonance imaging (4D-uMRI), we found TBI caused extensive injury to the fetal brain that included cerebral hemorrhage, loss of cerebral tissue, and hydrocephalus with excessive accumulation of cerebrospinal fluid (CSF). Histopathology of the fetal mouse brain showed broken cerebral vessels and elevated apoptosis. Further use of novel 4D Oxy-wavelet MRI capable of probing in vivo mitochondrial function in intact brain revealed a significant reduction of mitochondrial function in the fetal brain after 3 Gy TBI. This was validated by ex vivo Oroboros mitochondrial respirometry. One day after TBI (E14.5) maternal administration of JP4-039, which passes through the placenta, significantly reduced fetal brain radiation injury and improved fetal brain mitochondrial respiration. Treatment also preserved cerebral brain tissue integrity and reduced cerebral hemorrhage and cell death. JP4-039 administration following irradiation resulted in increased survival of pups. These findings indicate that JP4-039 can be deployed as a safe and effective mitigator of fetal radiation injury from mid-gestational in utero ionizing radiation exposure.

center dot Pyroptosis is implicated in the pathogenesis of ionizing radiation so we investigated cell type involvement in a mouse model. center dot Pyroptotic markers were expressed highly at day 3 post total body irradiation (TBI) predominantly in immune cells from ileum. center dot We investigated the effectiveness of MCC950, a pyroptosis inhibitor, in our model showing survival benefit at 50 mg/kg. center dot Treatment with MCC950 showed elevation in full-length mediator IL-1(3 in ileum and decreased level of cleaved IL-1(3 in spleen.

BACKGROUND/AIM:Radiation oncologists are reluctant to treat cancer in Fanconi Anemia (FA) patients due to their lack of homologous recombination repair of DNA strand breaks in normal tissues. To determine the therapeutic effects of irradiation and combination chemotherapy on cancer in syngeneic, radiosensitive FA mice, we derived transplantable cancers of the same genotype in three FA mouse strains. MATERIALS AND METHODS:Fancd2-/- mice on a C57BL/6 or Sv/129 background and Fancg-/- mice (C57BL/6 background) that received 3-methylcholanthrene (3-MCA), were monitored for the development of subcutaneous tumors. RESULTS:Tumors were induced at the site of 3-MCA injection, and tumor cell lines were established and found to be transplantable. Explanted tumors were identified as pleomorphic/rhabdomyosarcomas using immunohistochemical biomarkers. CONCLUSION:These transplantable FA mouse tumor cell lines should be valuable for testing effects of new radiation therapy protocols including FLASH high dose rate radiation delivery, immunotherapies, and combined radiation and chemotherapy treatments for radiosensitive FA patients.

We hypothesize that the injection of JP4-039, a mitochondria-targeted nitroxide, prior to irradiation of the mouse retina may decrease apoptosis and reduce neutrophil and macrophage migration into the retina. In our study, we aimed to examine the effects of JP4-039 in the mouse retina using fluorescent microscopy, a terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, and flow cytometry. Forty-five mice and one eye per mouse were used. In Group 1, fluorescent microscopy was used to determine retinal uptake of 10 µL (0.004 mg/µL) of intravitreally injected BODIPY-labeled JP4-039 at 0, 15, and 60 min after injection. In Group 2, the TUNEL assay was performed to investigate the rate of apoptosis after irradiation in addition to JP4-039 injection, compared to controls. In Group 3, flow cytometry was used to determine the extent of inflammatory cell migration into the retina after irradiation in addition to JP4-039 injection, compared to controls. Maximal retinal uptake of JP4-039 was 15 min after intravitreal injection (p < 0.0001). JP4-039-treated eyes had lower levels of retinal apoptosis (35.8 ± 2.5%) than irradiated controls (49.0 ± 2.7%; p = 0.0066) and demonstrated reduced migration of N1 cells (30.7 ± 11.7% vs. 77.7 ± 5.3% controls; p = 0.004) and M1 cells (76.6 ± 4.2 vs. 88.1 ± 3.7% controls, p = 0.04). Pretreatment with intravitreally injected JP4-039 reduced apoptosis and inflammatory cell migration in the irradiated mouse retina, marking the first confirmed effect of this molecule in retinal tissue. Further studies may allow for safety profiling and potential use for patients with radiation retinopathy.

Bacterial biotherapeutic delivery vehicles have the potential to treat a variety of diseases. This approach obviates the need to purify the recombinant effector molecule, allows delivery of therapeutics in situ via oral or intranasal administration, and protects the effector molecule during gastrointestinal transit. Lactic acid bacteria have been broadly developed as therapeutic delivery vehicles though risks associated with the colonization of a genetically modified microorganism have so-far not been addressed. Here, we present an engineered Limosilactobacillus reuteri strain with reduced colonization potential. We applied a dual-recombineering scheme for efficient barcoding and generated mutants in genes encoding five previously characterized and four uncharacterized putative adhesins. Compared with the wild type, none of the mutants were reduced in their ability to survive gastrointestinal transit in mice. CmbA was identified as a key protein in L. reuteri adhesion to HT-29 and enteroid cells. The nonuple mutant, a single strain with all nine genes encoding adhesins inactivated, had reduced capacity to adhere to enteroid monolayers. The nonuple mutant producing murine IFN-β was equally effective as its wild-type counterpart in mitigating radiation toxicity in mice. Thus, this work established a novel therapeutic delivery platform that lays a foundation for its application in other microbial therapeutic delivery candidates and furthers the progress of the L. reuteri delivery system towards human use.IMPORTANCEOne major advantage to leverage gut microbes that have co-evolved with the vertebrate host is that evolution already has taken care of the difficult task to optimize survival within a complex ecosystem. The availability of the ecological niche will support colonization. However, long-term colonization of a recombinant microbe may not be desirable. Therefore, strategies need to be developed to overcome this potential safety concern. In this work, we developed a single strain in which we inactivated the encoding sortase, and eight genes encoding characterized/putative adhesins. Each individual mutant was characterized for growth and adhesion to epithelial cells. On enteroid cells, the nonuple mutant has a reduced adhesion potential compared with the wild-type strain. In a model of total-body irradiation, the nonuple strain engineered to release murine interferon-β performed comparable to a derivative of the wild-type strain that releases interferon-β. This work is an important step toward the application of recombinant L. reuteri in humans.

Background Radiation therapy often leads to late radiation-induced skin fibrosis (RISF), causing movement impairment and discomfort. We conducted a comprehensive study to assess the effectiveness of metformin and adipose-derived stem cells (ASCs), whether autologous or allogeneic, individually or in combination therapy, in mitigating RISF. Methods Using a female C57BL/6J mouse model subjected to hind limb irradiation as a representative RISF model, we evaluated metformin, ASCs, or their combination in two contexts: prophylactic (started on day 1 post-irradiation) and therapeutic (initiated on day 14 post-irradiation, coinciding with fibrosis symptoms). We measured limb movement, examined skin histology, and analyzed gene expression to assess treatment efficacy. Results Prophylactic metformin and ASCs, whether autologous or allogeneic, effectively prevented late fibrosis, with metformin showing promising results. However, combination therapy did not provide additional benefits when used prophylactically. Autologous ASCs, alone or with metformin, proved most effective against late-stage RISF. Prophylactic intervention outperformed late therapy for mitigating radiation skin damage. Co-culture studies revealed that ASCs and metformin downregulated inflammation and fibrotic gene expression in both mouse and human fibroblasts. Conclusions Our study suggests metformin's potential as a prophylactic measure to prevent RISF, and the combination of ASCs and metformin holds promise for late-stage RISF treatment. These findings have clinical implications for improving the quality of life for those affected by radiation-induced skin fibrosis.

Despite recent advances in cancer therapy, ovarian cancer remains the most lethal gynecological cancer worldwide, making it crucial and of the utmost importance to establish novel therapeutic strategies. Adjuvant radiotherapy has been assessed historically, but its use was limited by intestinal toxicity. We recently established the role of Limosilactobacillus reuteri in releasing IL-22 (LR-IL-22) as an effective radiation mitigator, and we have now assessed its effect in an ovarian cancer mouse model. We hypothesized that an LR-IL-22 gavage would enable intestinal radioprotection by modifying the tumor microenvironment and, subsequently, improving overall survival in female C57BL/6MUC-1 mice with widespread abdominal syngeneic 2F8cis ovarian cancer. Herein, we report that the LR-IL-22 gavage not only improved overall survival in mice when combined with a PD-L1 inhibitor by inducing differential gene expression in irradiated stem cells but also induced PD-L1 protein expression in ovarian cancer cells and mobilized CD8+ T cells in whole abdomen irradiated mice. The addition of LR-IL-22 to a combined treatment modality with fractionated whole abdomen radiation (WAI) and systemic chemotherapy and immunotherapy regimens can facilitate a safe and effective protocol to reduce tumor burden, increase survival, and improve the quality of life of a locally advanced ovarian cancer patient.

Background/Aim: Patients with radiation sensitive Fanconi anemia (FA) are presenting with cancers of the oral cavity, oropharynx, and other anatomic locations. Materials and Methods: Animal models for cancer in FA mice used orthotopic tumors from wild type mice. We derived a cancer cell line from Fanca-/- mice by topical application of the chemical carcinogen dimethyl benzanthracene (DMBA). Results: A Fanca-/- mouse rhabdomyosarcoma was derived from a Fanca-/- (129/Sv) mouse. The in vitro clonogenic survival of the Fanca-/- clone 6 cancer cell line was consistent with the FA genotype. Transplanted tumors demonstrated hypoxic centers surrounded by senescent cells. Conclusion: This Fanca-/- mouse syngeneic cancer should provide a valuable resource for discovery and development of new normal tissue radioprotectors for patients with FA and cancer.

Cellular senescence is involved in the development of pulmonary fibrosis as well as in lung tissue repair and regeneration. Therefore, a strategy of removal of senescent cells by senolytic drugs may not produce the desired therapeutic result. Previously we reported that tyrosine kinase Fgr is upregulated in ionizing irradiation-induced senescent cells. Inhibition of Fgr reduces the production of profibrotic proteins by radiation-induced senescent cells in vitro; however, a mechanistic relationship between senescent cells and radiation-induced pulmonary fibrosis (RIPF) has not been established. We now report that senescent cells from the lungs of mice with RIPF, release profibrotic proteins for target cells and secrete chemotactic proteins for marrow cells. The Fgr inhibitor TL02-59, reduces this release of profibrotic chemokines from the lungs of RIPF mice, without reducing numbers of senescent cells. In vitro studies demonstrated that TL02-59 abrogates the upregulation of profibrotic genes in target cells in transwell cultures. Also, protein arrays using lung fibroblasts demonstrated that TL02-59 inhibits the production of chemokines involved in the migration of macrophages to the lung. In thoracic-irradiated mice, TL02-59 prevents RIPF, significantly reduces levels of expression of fibrotic gene products, and significantly reduces the recruitment of CD11b+ macrophages to the lungs. Bronchoalveolar lavage (BAL) cells from RIPF mice show increased Fgr and other senescent cell markers including p16. In human idiopathic pulmonary fibrosis (IPF) and in RIPF, Fgr, and other senescent cell biomarkers are increased. In both mouse and human RIPF, there is an accumulation of Fgr-positive proinflammatory CD11b+ macrophages in the lungs. Thus, elevated levels of Fgr in lung senescent cells upregulate profibrotic gene products, and chemokines that might be responsible for macrophage infiltration into lungs. The detection of Fgr in senescent cells that are obtained from BAL during the development of RIPF may help predict the onset and facilitate the delivery of medical countermeasures.

PURPOSE: Radiation-induced fibrosis (RIF) is a pathology characterized by large extra-cellular matrix deposition and excessive proliferation of fibroblasts leading to reduced tissue function and quantity of life. Currently, very few safe and effective treatment options are available. Our goal was to test early and late regimens of a safe and well-tolerated FDA-approved agent (metformin) for RIF inhibition. METHODS: We used 40 Gy hind limb irradiated C57BL/6 mice as a skin fibrosis model and injected 100 mg/kg of metformin intraperitoneally twice a week. The early cohort started the treatment immediately; the late cohort was delayed to 14th post-irradiation day to investigate the inhibition of fibrosis on day 42. PBS-injected mice were used as a control group. Regimens were combined with either autologous or allogeneic adipose stem cells. The degree of limb excursion, epithelial thickness, and a histological fibrosis scale were employed. RESULTS: Allogenic and autologous cells successfully mitigated fibrosis; superior histological scoring and significantly improved limb excursion and epithelial thickness were achieved compared to the control. Earlier treatment increased the mitigation of fibrosis in all cohorts. The acute metformin treatment gave outcomes comparable to cell injections; when delayed, metformin group failed to inhibit significant fibrosis. In contrast, the effect of cell therapies did not diminish to the level of insignificance. CONCLUSION: Our findings suggest that allogeneic stem cells and early onset metformin treatment are possible promising therapeutic options to inhibit fibrosis. Further investigating regulatory markers in depth will be a step toward clinical adaptation as a RIF prophylaxis.

Radiation therapy can lead to late radiation-induced skin fibrosis (RISF), causing movement restriction, pain, and organ dysfunction. This study evaluated adipose-derived extracellular matrix (Ad-ECM) as a mitigator of RISF. Female C57BL/6J mice that were irradiated developed fibrosis, which was mitigated by a single local Ad-ECM injection, improving limb movement and reducing epithelium thickness and collagen deposition. Ad-ECM treatment resulted in decreased expression of pro-inflammatory and fibrotic genes, and upregulation of anti-inflammatory cytokines, promoting M2 macrophage polarization. Co-culture of irradiated human fibroblasts with Ad-ECM down-modulated fibrotic gene expression and enhanced bone marrow cell migration. Ad-ECM treatment also increased interleukin (IL)-4, IL-5, and IL-15 expression in endothelial cells, stimulating M2 macrophage polarization and alleviating RISF. Prophylactic use of Ad-ECM showed effectiveness in mitigation. This study suggests Ad-ECM's potential in treating chronic-stage fibrosis.

"Vitamin C is Mandatory for the TCA Cycle Production of Anti-Inflammatory Itaconate." American Journal of Respiratory and Critical Care Medicine, 0(ja), pp.

BACKGROUND/AIM:The earliest cellular and molecular biologic changes in the esophagus that lead to esophageal cancer were evaluated in a mouse model. We correlated numbers of senescent cells with the levels of expression of potentially carcinogenic genes in sorted side population (SP) cells containing esophageal stem cells and non-stem cells in the non-side population cells in the 4-nitroquinolone oxide (NQO)-treated esophagus. MATERIALS AND METHODS:We compared stem cells with non-stem cells from the esophagus of mice treated with the chemical carcinogen 4-NQO (100 μg/ml) in drinking water. We also compared gene expression in human esophagus samples treated with 4-NQO (100 μg/ml media) to non-treated samples. We separated and quantitated the relative levels of expression of RNA using RNAseq analysis. We identified senescent cells by luciferase imaging of p16+/LUC mice and senescent cells in excised esophagus from tdTOMp16+ mice. RESULTS:A significant increase in the levels of RNA for oncostatin-M was found in senescent cells of the esophagus from 4-NQO-treated mice and human esophagus in vitro. CONCLUSION:Induction of OSM in chemically-induced esophageal cancer in mice correlates with the appearance of senescent cells.

Introduction: Cardiac irradiation causes heart failure and arrhythmias. We reported that cardiac-targeted irradiation (CTI) led to cardiac dilation with increased end diastolic volume (EDV) by echocardiogram (echo) after 9 months in wild-type C57Bl6/J (WT) male (M) but not female (F) mice. Both sexes developed cardiac fibrosis and evidence of lung congestion, and the radiation mitigator MMS350 did not protect either sex. Here, we studied the effect of CTI on Nitric Oxide Synthase 1 haploinsufficient (Nos +/- ) mice treated with/without MMS350. Methods: 10 week-old Nos +/- and WT mice were exposed to 20 Gy CTI (R) with or without MMS350 (D) and followed for 9 months (Nos +/- : +R+D, n = 9M, 6F; +R-D, n = 6M, 7F; WT: +R+D, n = 12M, 14F; +R-D, n = 13M, 14F), with parallel unirradiated controls (Nos +/- : -R+D, n = 8M, 6F; -R-D, n = 10M, 7F; WT: -R+D, n = 8M, 12F; -R-D, n = 7M, 8F)). Echos and EKGs were performed at baseline and every 3 months. Heart weight (HW), lung weight (LW), and body weight (BW) were measured. Results: Unlike irradiated male WT mice where EDV was increased (p=0.001), EDV was smaller in male (p=0.04) and EF was decreased in both male (p=0.01) and female (p=0.05) Nos +/- irradiated mice compared to unirradiated controls, independent of MMS350 treatment (Figure). In addition, the PR interval was shorter in irradiated Nos +/- male (p=0.008) and female (p=0.003) mice compared to unirradiated controls. HW/BW was not significantly changed in any group, while LW/BW was significantly increased only in WT male (p=0.04) and female (p=0.001) irradiated mice. Conclusions: In long term follow-up of CTI-treated mice, Nos1 deficiency prevents sex-specific cardiac dilation in males, promotes left ventricular systolic dysfunction and alters cardiac conduction system properties. Further investigation into the role of nitric oxide in radiation-induced cardiac damage and the mechanisms underlying sex differences in the response to radiation are warranted.