Mitigation of Nitrogen Mustard-Induced Skin Injury by the ß-Blocker Carvedilol and Its Enantiomers.

The chemical warfare agent sulfur mustard and its structural analog nitrogen mustard (NM) cause severe vesicating skin injuries. The pathologic mechanisms for the skin injury following mustard exposure are poorly understood; therefore, no effective countermeasure is available. Previous reports demonstrated the protective activity of carvedilol, a US Food and Drug Administration (FDA)-approved ß-blocker, against UV radiation-induced skin damage. Thus, the current study evaluated the effects of carvedilol on NM-induced skin injuries in vitro and in vivo. In the murine epidermal cell line JB6 Cl 41-5a, ß-blockers with different receptor subtype selectivity were examined. Carvedilol and both of its enantiomers, R- and S-carvedilol, were the only tested ligands statistically reducing NM-induced cytotoxicity. Carvedilol also reduced NM-induced apoptosis and p53 expression. In SKH-1 mice, NM increased epidermal thickness, damaged skin architecture, and induced nuclear factor κB (NF-κB)-related proinflammatory genes as assessed by RT2 Profiler PCR (polymerase chain reaction) Arrays. To model chemical warfare scenario, 30 minutes after exposure to NM, 10 µM carvedilol was applied topically. Twenty-four hours after NM exposure, carvedilol attenuated NM-induced epidermal thickening, Ki-67 expression, a marker of cellular proliferation, and multiple proinflammatory genes. Supporting the in vitro data, the non-ß-blocking R-enantiomer of carvedilol had similar effects as racemic carvedilol, and there was no difference between carvedilol and R-carvedilol in the PCR array data, suggesting that the skin protective effects are independent of the ß-adrenergic receptors. These data suggest that the ß-blocker carvedilol and its enantiomers can be repurposed as countermeasures against mustard-induced skin injuries. SIGNIFICANCE STATEMENT: The chemical warfare agent sulfur mustard and its structural analog nitrogen mustard cause severe vesicating skin injuries for which no effective countermeasure is available. This study evaluated the effects of US Food and Drug Administration (FDA)-approved ß-blocker carvedilol on nitrogen mustard-induced skin injuries to repurpose this cardiovascular drug as a medical countermeasure.

Genomic Adaption and Mutational Patterns in a HaCaT Subline Resistant to Alkylating Agents and Ionizing Radiation.

Sulfur mustard (SM) is a chemical warfare agent that can damage DNA via alkylation and oxidative stress. Because of its genotoxicity, SM is cancerogenic and the progenitor of many chemotherapeutics. Previously, we developed an SM-resistant cell line via chronic exposure of the popular keratinocyte cell line HaCaT to increasing doses of SM over a period of 40 months. In this study, we compared the genomic landscape of the SM-resistant cell line HaCaT/SM to its sensitive parental line HaCaT in order to gain insights into genetic changes associated with continuous alkylation and oxidative stress. We established chromosome numbers by cytogenetics, analyzed DNA copy number changes by means of array Comparative Genomic Hybridization (array CGH), employed the genome-wide chromosome conformation capture technique Hi-C to detect chromosomal translocations, and derived mutational signatures by whole-genome sequencing. We observed that chronic SM exposure eliminated the initially prevailing hypotetraploid cell population in favor of a hyperdiploid one, which contrasts with previous observations that link polyploidization to increased tolerance and adaptability toward genotoxic stress. Furthermore, we observed an accumulation of chromosomal translocations, frequently flanked by DNA copy number changes, which indicates a high rate of DNA double-strand breaks and their misrepair. HaCaT/SM-specific single-nucleotide variants showed enrichment of C > A and T > A transversions and a lower rate of deaminated cytosines in the CpG dinucleotide context. Given the frequent use of HaCaT in toxicology, this study provides a valuable data source with respect to the original genotype of HaCaT and the mutational signatures associated with chronic alkylation and oxidative stress.

Relevance of Erk1/2-PI3K/Akt signaling pathway in CEES-induced oxidative stress regulates inflammation and apoptosis in keratinocytes.

2-Chloroethyl ethyl sulfide (CEES) is a well-known chemical warfare agent that induces cellular stress in exposed individuals. However, molecular mechanisms of CEES-induced oxidative stress-mediated metabolic deregulation are not clearly elucidated. Here we investigated CEES-induced free radical production act as key functional mediators of metabolic stress via Erk1/2 mitogen-activated protein kinases (MAPKs) and phosphatidylinositol-3-kinase (PI3K/Akt) signaling cascades in keratinocytes. We observed that CEES exposure disrupts the cellular antioxidant defense capacities leading to increase in free oxygen and nitrogen radical accumulation in keratinocytes. These unusual cellular abnormalities initiate cellular stress via Erk1/2-PI3K/Akt signaling pathways. Biochemical tools were used to analyze the changes in metabolites including sulfur amino acids (SAAs), namely, L-glutathione (GSH) and L-cysteine (Cys), in the presence of selective inhibitors of reactive oxygen/nitrogen species (ROS/RNS), Erk1/2, or PI3K/Akt after CEES exposure. Importantly, these metabolite changes were accompanied by a decrease in the glycolytic flux, consistent with the observed decrease in 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFK-2) concentration and these CEES-induced phenomena were attenuated by pretreatment of Erk1/2 or PI3-K/Akt inhibitors. On the other hand, CEES exposure disrupts the protein carbonylation (PC) and lipid peroxidation (LPO) in keratinocytes leading to inflammation, crash of the cell-cell communication, cell cycle deregulation, and apoptosis via Erk1/2-PI3K/Akt pathways. However, pretreatment of Erk1/2 or PI3K/Akt inhibitors attenuated the CEES action. Collectively, these results illustrated that accumulated free radicals act as key functional mediators for inflammation, and apoptosis via Erk1/2-PI3K/Akt regulatory signaling cascades induced by CEES exposure. Treatment of pharmacological Erk1/2-PI3K/Akt inhibitors attenuated the CEES-induced keratinocyte injury that may provide the basis for the development of therapeutic strategy to work against CEES exposure.

Carvacrol ameliorates haematological parameters, oxidant/antioxidant biomarkers and pulmonary function tests in patients with sulphur mustard-induced lung disorders: A randomized double-blind clinical trial.

WHAT IS KNOWN AND OBJECTIVE: In this study, the effect of carvacrol (CAR) on pulmonary function tests (PFT), haematological indices and oxidant/antioxidant biomarkers in patients with sulphur mustard (SM)-induced lung disorders was examined. METHODS: Twenty patients exposed to SM 27-30 years ago were divided into two groups and treated with either placebo (P) or CAR (1.2 mg/kg per day) (n = 10 for each group). Forced vital capacity (FVC), peak expiratory flow (PEF), total and different white blood cell (WBC), haematological parameters and oxidant/antioxidant biomarkers were measured at the baseline (step 0), one and two months (steps I and II, respectively) after starting the treatment. RESULTS AND DISCUSSION: PEF was significantly increased in the CAR-treated group in step II compared to step 0 (P < .01). Total WBC (P < .01) and neutrophil (P < .05) count in the CAR-treated group were significantly decreased in the group in steps I and II (P < .01 for both cases) compared to step 0. The levels of thiol, superoxide dismutase and catalase in the CAR-treated group were significantly increased (P < .05 to P < .001) in steps I and II, but malondialdehyde significantly decreased in step II compared to step 0 (P < .01). The percentage of total and differential WBC, oxidant/antioxidant biomarkers, FVC and PEF values following a two-month treatment period were significantly improved in the CAR-treated group compared to the placebo group (P < .05 to P < .001). WHAT IS NEW AND CONCLUSION: Two-month treatment with CAR reduced inflammatory cells and oxidant biomarkers, whereas increased antioxidant biomarkers and improved PFT tests in SM-exposed patients.

NMR spectroscopy-based metabolomic study of serum in sulfur mustard exposed patients with lung disease.

Sulfur mustard (SM) is a vesication chemical warfare agent for which there is currently no antidote. Despite years of research, there is no common consensus about the pathophysiological basis of chronic pulmonary disease caused by this chemical warfare agent. In this study, we combined chemometric techniques with nuclear magnetic resonance (NMR) spectroscopy to explore the metabolic profile of sera from SM-exposed patients. A total of 29 serum samples obtained from 17 SM-injured patients, and 12 healthy controls were analyzed by Random Forest. Increased concentrations of seven amino acids, glycerol, dimethylamine, ketone bodies, lactate, acetate, citrulline and creatine together with the decreased very low-density lipoproteins (VLDL) levels were observed in patients compared with control subjects. Our study reveals the metabolic profile of sera from SM-injured patients and indicates that NMR-based methods can distinguish these patients from healthy controls.

Oxidative stress and altered expression of peroxiredoxin genes family (PRDXS) and sulfiredoxin-1 (SRXN1) in human lung tissue following exposure to sulfur mustard.

BACKGROUND: Sulfur mustard (SM) is a potent and mutagenic agent that targets human lung tissue. PURPOSE OF THE STUDY: The purpose of this investigation is to characterize the expression of sulfiredoxin-1 (SRXN1) and peroxiredoxin (PRDXs) genes and oxidative stress (OS) status in human lung after exposure to SM. MATERIALS AND METHODS: Lung biopsy specimens bronchoalveolar lavage (BAL) fluids were provided from SM-exposed patients (n = 6) and controls (n = 5). Changes in gene expression were measured using RT(2) Profiler PCR Array. OS was considered by measuring BAL fluid levels of malondialdehyde (MDA) and protein carbonyls (PC). RESULTS: Mean of MDA and PC values in BAL fluid of patients (0.6467 ± 0.05922 nmol/l and 1.391 ± 0.421 nmol/mg, respectively) was higher than in controls (0.486 ± 0.04615 nmol/l and 0.949 ± 0.149 nmol/mg, respectively). Expression of all examined genes was in the order PRDX1> PRDX3> PRDX6> SRXN1> PRDX2> PRDX4> PRDX5. Among the most upregulated genes was the PRDX1, which was overexpressed by 10.1029-fold (p = 0.000634). SM-exposed individuals demonstrated expression of PRDX3 4.6231 (p = 0.000134), PRDX6 3.4964 (p = 0.001102), SRXN1 3.3719 (p < 0.0001) and PRDX2 2.7725-folds (p = 0.000383) higher than those of controls that reveal. CONCLUSIONS: Upregulation of PRDXs and SRXN1 genes may be because of reactive oxygen species (ROS) production and OS in lung tissue of patients after SM exposure. Expression of SRXN1 and PRDXNs genes, especially I, II, III, and VI is increased in SM-injured lungs, suggesting the induction of cellular responses to increased production of ROS and OS in lung of the patients. Therefore, sulfiredoxin and peroxiredoxins can be targeted as biomarkers of OS in these patients.

Analysis of different fates of DNA adducts in adipocytes post-sulfur mustard exposure in vitro and in vivo using a simultaneous UPLC-MS/MS quantification method.

Sulfur mustard (SM) is a powerful alkylating vesicant that can rapidly penetrate skin, ocular, and lung bronchus mucous membranes and react with numerous nucleophiles in vivo. Although the lesion mechanisms of SM remain unclear, DNA damage is believed to be the most crucial factor in initiating SM-induced toxicity. Four major DNA adducts were identified for retrospective detection and DNA lesion evaluation, namely, N(7)-[2-[(2-hydroxyethyl)thio]-ethyl]guanine (N(7)-HETEG), bis(2-ethyl-N(7)-guanine)thioether (Bis-G), N(3)-(2-hydroxyethylthioethyl)-2'-adenine (N(3)-HETEA), and O(6)-[2-[(2-hydroxyethyl)thio]-ethyl]guanine (O(6)-HETEG). Because of previous observations that the levels of SM-DNA adducts were relatively higher in adipose-rich organs, such as the brain, we focused on the in vitro and in vivo fates of the DNA adducts in exposed adipocytes. A UPLC-MS/MS method developed in our laboratory was used to profile the N(7)-HETEG, Bis-G, and N(3)-HETEA levels in human mature adipocytes (HA-s) that had differentiated from human subcutaneous preadipocytes (HPA-s). This method was also used to profile three other cell lines related to the targeting of major tissues, including human keratinocytes (HaCaT), human hepatocytes (L-02), and human lung fibroblasts (HLF). Long-lasting adduct persistence and a high proportion of Bis-G were found in exposed adipocytes in vitro. The survival properties of exposed adipocytes were also tested. At the same time, the fate of SM-DNA adducts in vivo was characterized using a rat model exposed to 1 and 10 mg/kg doses of SM. The level of DNA adducts in the exposed adipose tissue (AT) was much lower than those in other organs studied in our previous work. The adduct persistence behavior was observed in AT with an extremely high proportion of Bis-G, which was higher than N(7)-HETEG. In light of these results, we suggest that an adipose-rich environment may promote the formation of Bis-G and that adipocyte-specific DNA repair mechanisms may result in adduct persistence and the survival of adipocytes after SM exposure. These conclusions should be further investigated.

Metabolomics to investigate the effect of preconditioned mesenchymal stem cells with crocin on pulmonary epithelial cells exposed to 2-chloroethyl ethyl sulfide.

Metabolomics significantly impacts drug discovery and precise disease management. This study meticulously assesses the metabolite profiles of cells treated with Crocin, Dexamethasone, and mesenchymal stem cells (MSCs) under oxidative stress induced by 2-chloroethyl ethyl sulfide (CEES). Gas chromatography/mass spectrometry (GC/MS) analysis unequivocally identified substantial changes in 37 metabolites across the treated groups. Notably, pronounced alterations were observed in pathways associated with aminoacyl-tRNA biosynthesis and the metabolism of aspartate, serine, proline, and glutamate. These findings demonstrate the potent capacity of the analyzed treatments to effectively reduce inflammation, mitigate reactive oxygen species production, and enhance cell survival rates. SIGNIFICANCE.

Tissue plasminogen activator prevents mortality from sulfur mustard analog-induced airway obstruction.

Sulfur mustard (SM) inhalation causes the rare but life-threatening disorder of plastic bronchitis, characterized by bronchial cast formation, resulting in severe airway obstruction that can lead to respiratory failure and death. Mortality in those requiring intubation is greater than 80%. To date, no antidote exists for SM toxicity. In addition, therapies for plastic bronchitis are solely anecdotal, due to lack of systematic research available to assess drug efficacy in improving mortality and/or morbidity. Adult rats exposed to SM analog were treated with intratracheal tissue plasminogen activator (tPA) (0.15-0.7 mg/kg, 5.5 and 6.5 h), compared with controls (no treatment, isoflurane, and placebo). Respiratory distress and pulse oximetry were assessed (for 12 or 48 h), and arterial blood gases were obtained at study termination (12 h). Microdissection of fixed lungs was done to assess airway obstruction by casts. Optimal intratracheal tPA treatment (0.7 mg/kg) completely eliminated mortality (0% at 48 h), and greatly improved morbidity in this nearly uniformly fatal disease model (90-100% mortality at 48 h). tPA normalized plastic bronchitis-associated hypoxemia, hypercarbia, and lactic acidosis, and improved respiratory distress (i.e., clinical scores) while decreasing airway fibrin casts. Intratracheal tPA diminished airway-obstructive fibrin-containing casts while improving clinical respiratory distress, pulmonary gas exchange, tissue oxygenation, and oxygen utilization in our model of severe chemically induced plastic bronchitis. Most importantly, mortality, which was associated with hypoxemia and clinical respiratory distress, was eliminated.

Long-term effects of mustard gas on respiratory system of Iranian veterans after Iraq-Iran war: a review.

To review long-term respiratory effects of mustard gas on Iranian veterans having undergone Iraq-Iran war. Electronic databases of Scopus, Medline, ISI, IranMedex, and Irandoc sites were searched. We accepted articles published in scientific journals as a quality criterion.The main pathogenic factors are free radical mediators. Prevalence of pulmonary involvement is approximately 42.5%. The most common complaints are cough and dyspnea. Major respiratory complications are chronic obstructive pulmonary disease, bronchiectasis, and asthma. Spirometry results can reveal restrictive and obstructive pulmonary disease. Plain chest X-ray does not help in about 50% of lung diseases. High-resolution CT of the lung is the best modality for diagnostic assessment of parenchymal lung and bronchi. There is no definite curative treatment for mustard lung. The effective treatment regimens consist of oxygen administration, use of vaporized moist air, respiratory physiotherapy, administration of mucolytic agents, bronchodilators, corticosteroids, and long-acting beta-2 agonists, antioxidants, surfactant, magnesium ions, therapeutic bronchoscopy, laser therapy, placement of respiratory stents, early tracheostomy in laryngospasm, and ultimately lung transplantation. High-resolution CT of the lung is the most accurate modality for the evaluation of the lung parenchyma and bronchi. The treatment efficacy of patients exposed to mustard gas depends on patient conditions (acute or chronic, upper or lower respiratory tract involvement). There are various treatment protocols, but unfortunately none of them is definitely curable.

Time course pathogenesis of sulphur mustard-induced skin lesions in mouse model.

Sulphur mustard (SM) is a bifunctional alkylating agent that causes cutaneous blistering in humans and animals. In this study, we have presented closer views on pathogenesis of SM-induced skin injury in a mouse model. SM diluted in acetone was applied once dermally at a dose of 5 or 10 mg/kg to Swiss albino mice. Skin was dissected out at 0, 1, 3, 6, 12, 24, 48, 72 and 168 hours, post-SM exposure for studying histopathological changes and immunohistochemistry of inflammatory-reparative biomarkers, namely, transforming growth factor alpha (TGF-α), fibroblast growth factor (FGF), endothelial nitric oxide synthase (eNOS) and interlukin 6 (IL-6). Histopathological changes were similar to other mammalian species and basal cell damage resembled the histopathological signs observed with vesication in human skin. Inflammatory cell recruitment at the site of injury was supported by differential expressions of IL-6 at various stages. Time-dependent expressions of eNOS played pivotal roles in all the events of wound healing of SM-induced skin lesions. TGF-α and FGF were strongly associated with keratinocyte migration, re-epithelialisation, angiogenesis, fibroblast proliferation and cell differentiation. Furthermore, quantification of the tissue leukocytosis and DNA damage along with semiquantitative estimation of re-epithelialisation, fibroplasia and neovascularisation on histomorphologic scale could be efficiently used for screening the efficacy of orphan drugs against SM-induced skin injury.

Evaluation of Decontamination Efficacy of Electrolytically Generated Hypochlorous Acid for the Vesicating Agent: A Multimodel Study.

BACKGROUND: Sulfur Mustard is a strong vesicant and chemical warfare agent that imposes toxicity to the lungs, eyes, and skin after accidental or intended exposure. OBJECTIVES: The current study was intended to explore in vitro and in vivo decontamination properties of electrolytically generated HOCl (hypochlorous acid) against CEES (2-chloroethyle ethyle sulphide), a known sulfur mustard simulant & vesicating agent. METHODS: in vitro studies were carried out using UV spectroscopy and GC-MS methods. In vivo studies were performed in Strain A and immune-compromised mice by subcutaneous as well as prophylactic topical administration of HOCl pretreated CEES. The blister formation and mortality were considered as end-point. Histopathological study was conducted on skin samples by H & E method. DNA damage studies measuring γ-H2AX and ATM have been carried out in human blood using flow cytometry. Anti-bacterial action was tested by employing broth micro dilution methods. A comparative study was also carried out with known oxidizing agents. RESULTS: The topical application of pre-treated CEES at 5, 30 min and 1 h time points showed significant (p<0.001) inhibition of blister formation. DNA damage study showed reduced mean fluorescence intensity of DSBs nearly 17-20 times, suggesting that HOCl plays a protective role against DNA damage. Histopathology showed no sign of necrosis in the epidermis upto 5 min although moderate changes were observed at 30 min. Pretreated samples were analyzed for detection of reaction products with m/z value of 75.04, 69.08, 83.93, 85.95, 123.99, 126.00, and 108.97. HOCl showed a strong bactericidal effect at 40 ppm. The absorbance spectra of HOCl treated CEES showed lowered peaks in comparison to CEES alone and other oxidizing agents. CONCLUSION: In a nutshell, our results signify the decontamination role of HOCl for biological surface application.

Fe-Doped MoS2 Nanozyme for Antibacterial Activity and Detoxification of Mustard Gas Simulant.

The peroxidase-like catalytic activity of various nanozymes was extensively applied in various fields. In this study, we have demonstrated the preparation of Fe-doped MoS2 (Fe@MoS2) nanomaterials with enhanced peroxidase-like activity of MoS2 in a co-catalytic pathway. In view of Fenton reaction, the peroxidase-like Fe@MoS2 nanozyme prompted the decomposition of hydrogen peroxide (H2O2) to a reactive hydroxyl radical (·OH). The efficient decomposition of H2O2 in the presence of Fe@MoS2 has been employed toward the antibacterial activity and detoxification of mustard gas simulant. The combined effect of Fe@MoS2 and H2O2 showed remarkable antibacterial activity against the drug-resistant bacterial strain methicillin-resistant Staphylococcus aureus and Escherichia coli with the use of minimal concentration of H2O2. Fe@MoS2 was further applied for the detoxification of the chemical warfare agent sulfur mustard simulant, 2-chloroethyl ethyl sulfide, by selective conversion to the nontoxic sulfoxide. This work demonstrates the development of a hybrid nanozyme and its environmental remediation from harmful chemicals to microbes.

Management of mustard gas-induced limbal stem cell deficiency and keratitis.

PURPOSE: To report the clinical findings and compare outcomes of different surgical techniques evolved over time in a large series of patients with delayed-onset mustard gas keratitis (MGK). DESIGN: Retrospective, comparative, interventional case series. PARTICIPANTS: Ninety Iranian male survivors (175 eyes) of Iraqi chemical warfare with chronic or delayed-onset MGK. METHODS: The symptoms and clinical findings of patients are presented, and medical and surgical interventions to address dry eye, limbal ischemia and limbal stem cell deficiency (LSCD), and corneal involvements are explained. The results of limbal stem cell transplantation techniques (living-related conjunctival-limbal allograft [lrCLAL] versus keratolimbal allograft [KLAL]) as well as corneal transplantation techniques (penetrating keratoplasty [PK] versus lamellar keratoplasty [LK]) are compared in terms of clinical outcomes and graft survival rates. MAIN OUTCOME MEASURES: Ocular findings and appropriate surgical approach for LSCD and corneal involvements. RESULTS: A total of 175 eyes of 90 cases (all male) between 34 and 68 years of age were followed up for 101±30.3 months (range, 36-198 months). The most common ocular involvements were chronic blepharitis and dry eye. Conjunctival vascular abnormalities and limbal ischemia were observed in 27.4% and 29.7% of eyes, respectively. Limbal stem cell deficiency necessitating stem cell transplantation developed in 41.1% of eyes. The most common corneal sign was central and peripheral anterior stromal opacity (58.9%), followed by corneal stromal thinning (36.0%) and neovascularization (27.4%). Living-related conjunctival-limbal allograft was performed in 32 eyes, and KLAL was performed in 40 eyes. The rejection-free graft survival rate was 39.1% in the lrCLAL group and 80.7% in the KLAL group at month 40, with a mean length of 24.9 and 68.8 months, respectively (P = 0.02). Thirty eyes underwent PK and 51 underwent LK. Corneal graft failure was observed in 9 PK eyes and in 6 LK eyes. The rejection-free graft survival rate was 39.0% in the PK group and 90.3% in the LK group at month 28, with a mean length of 29.6 and 85.0 months, respectively (P<0.001). CONCLUSIONS: Chemical warfare victims who initially have mild symptoms ultimately may experience significant ocular involvements requiring surgical intervention. Limbal and corneal abnormalities can be managed best by KLAL and LK, respectively.

Nuclear factor κB1/RelA mediates the inflammation and/or survival of human airway exposed to sulfur mustard.

CONTEXT: Sulfur mustard (SM) is known as an effective chemical agent and was used in the 1980s during the Iran-Iraq war against Iranians. At the present time, there are more than 40,000 people suffering from pulmonary lesions due to mustard gas in Iran. Though much is known about the gross pathology of SM damage, the molecular and cellular basis for this pathology is not well understood. OBJECTIVE: One of the most important protein groups involved in inflammatory responses is nuclear factor κB protein (NF-κB1) family. They belong to the category of DNA-binding protein factors necessary for transcription of many proinflammatory molecules. In our research, we examined the role of NF-κB1/RelA in the pathophysiology of the lung. MATERIALS AND METHODS: We investigated 10 normal individuals and 20 SM induced patients. Expression of NF-κB1/RelA in controls and the SM exposed samples was measured by real-time polymerase chain reaction and localization of NF-κB1 protein was detected by immunohistochemistry staining. RESULTS: Our results revealed that expression levels of NF-κB1 and RelA were upregulated 0.64-6.50 fold and 0.83-8.34 fold, respectively, in the SM exposed patients in comparison with control samples. DISCUSSION AND CONCLUSION: As far as we know, this is the first finding of induction of NF-κB in patients exposed to SM. NF-κB1/RelA may play a major role in inflammation induced by mustard gas or even in cell survival in the bronchial wall of affected patients.

Loss of expression of TGF-ßs and their receptors in chronic skin lesions induced by sulfur mustard as compared with chronic contact dermatitis patients.

BACKGROUND: Sulfur mustard (SM) is a blister-forming agent that has been used as a chemical weapon. Sulfur mustard can cause damage in various organs, especially the skin, respiratory system, and eyes. Generally, the multiple complications of mustard gas result from its alkalizing potency; it reacts with cellular components like DNA, RNA, proteins, and lipid membranes.TGF-ß is a multi-functional cytokine with multiple biological effects ranging from cell differentiation and growth inhibition to extracellular matrix stimulation, immunosuppression, and immunomodulation. TGF-ß has 3 isoforms (TGF-ß 1, 2, 3) and its signaling is mediated by its receptors: R1, R2 and intracellular Smads molecules.TGF-ß has been shown to have anti-inflammatory effects. TGF-ßs and their receptors also have an important role in modulation of skin inflammation, proliferation of epidermal cells, and wound healing, and they have been implicated in different types of skin inflammatory disorders. METHODS: Seventeen exposed SM individuals (48.47 ± 9.3 years), 17 chronic dermatitis patients (46.52 ± 14.6 years), and 5 normal controls (44.00 ± 14.6 years) were enrolled in this study.Evaluation of TGF-ßs and their receptors expressions was performed by semiquantitative RT-PCR. Only TGF1 was analyzed immunohistochemically. RESULTS: Our results showed significant decreases in the expression percentages of TGF-ß 1, 2 and R1, R2 in chemical victims in comparison with chronic dermatitis and normal subjects and significant decreases in the intensity of R1 and R2 expressions in chemical victims in comparison with chronic dermatitis and normal controls. (P value < 0.05) CONCLUSIONS: TGF-ßs and their receptors appear to have a noticeable role in chronic inflammatory skin lesions caused by sulfur mustard.

Skin decontamination efficacy of sulfur mustard and VX in the pig model: A comparison between Fuller's earth and RSDL.

Skin decontamination following exposure to chemical agents is a most important component of the individual defense doctrine, removing the agent, ceasing its penetration and preventing secondary contamination of the first responders. The goal of the current study was to compare the efficacy of Reactive Skin Decontaminant Lotion (RSDL) and Fuller's Earth (FE) following exposure to sulfur mustard (SM) and VX, aiming to find the optimal procedure for mass casualty decontamination protocol. Decontamination efficacy was evaluated in pigs by measurement of lesion area and erythema (SM) and cholinesterase inhibition and clinical symptoms (VX). FE and RSDL were highly effective against both agents. Following SM exposure, the two decontaminants demonstrated a significant decrease in lesions' size together with the decrease in exposure duration. Likewise, skin decontamination following exposure to VX with either FE or RSDL resulted in reduction in clinical symptoms and prevention of death. Decontamination was worthwhile even if postponed, up to 30 min (SM) and 2 h (VX). In conclusion, both decontamination products were efficient in ameliorating the toxic effects even though in a different mechanism. Finally, for mass casualty scenario, FE is preferred as a universal decontaminant, considering its safety, ease of use and longer shelf life.

Pathophysiological Effects of Sulfur Mustard on Skin and its Current Treatments: Possible Application of Phytochemicals.

BACKGROUND: Sulfur-(SM) and nitrogen (NM)-based mustards are the mutagenic incapacitating compounds which are widely used in vesicating the chemical warfare and cause toxicity in many organs, especially skin. SM, as a potent vesicating agent, contributes to the destruction of skin in dermis and epidermis layers. The progression of the lesion depends on the concentration of SM and the duration of exposure. Body responses start with pruritus, erythema, edema and xerosis, which lead to the accumulation of immune cells in the target sites and recruitment of mast cells and paracrine-mediated activity. Pro-inflammatory effectors are accumulated in the epidermis, hair follicles, and sebaceous glands resulting in the destruction of the basement membrane beneath the epidermis. There is still no satisfactory countermeasure against SM-induced lesions in clinical therapy, and the symptomatic or supportive treatments are routine management approaches. OBJECTIVE: The current review highlights the recent progression of herbal medicines application in SM-induced injuries through the illustrative examples and also demonstrates their efficacies, properties and mechanism of actions as therapeutic agents. CONCLUSION: Phytochemicals and herbal extracts with anti-bacterial, anti-inflammatory and antioxidant properties have been recently shown to hold therapeutic promise against the SM-induced cutaneous complications. The present review discusses the possible application of herbal medicines in the healing of SM-induced injuries.

Alterations in Mitochondrial and Inflammasome Homeostasis by 2-Chloroethyl Ethyl Sulfide and Their Mitigation by Curcumin: An in Vitro Study.

The mitochondrion has a substantial role in innate immunity and inflammasome signaling pathways. Sulfur mustard (SM) induces toxicity in cytoplasmic organelles. We aimed to evaluate the potential therapeutic effect of curcumin on the toxicity of SM analog through measuring gene expression levels of mitochondrial dynamics followed by induction of the inflammasome signaling pathway. After the treatment of pulmonary epithelial cell line (A549) by 2-chloroethyl ethyl sulfide (CEES) (2500 mM) for 48h, the transcriptional activity of mitochondrial fission and fusion genes such as dynamin-related protein 1 (Drp1), mitochondrial fission 1 protein (Fis1), mitofusin-1 (Mfn1), mitofusin-2 (Mfn2), and Dominant optic atrophy (Opa1) and inflammasome pathway genes including absent in melanoma 2 (AIM2), NLR family containing protein 3 (NLRP3), and Apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) was measured. Furthermore, the inhibitory effect of curcumin (160 mM) concurrent with SM analog on the expression level of mitochondria and inflammasome genes was investigated. CEES was able to over-express the fission, fusion (Drp1 ~ 8, Fis1 4.5, Mfn2 15, and Opa1 16-fold) and inflammasome genes (AIM2, NLRP3,  8 and 6-fold, respectively), whereas Mfn1 was significantly decreased (0.5-fold) and a not statistically significant decrease was observed in the ASC gene. Curcumin could modulate the effect of CEES, mitigate the expression of fission, fusion, and inflammasome genes exceedingly. However, a major increase in the repairer fusion gene (Mfn1, 6-fold) and complete suppression of the ASC gene were the outcomes of using the curcumin. In conclusion, we suggest curcumin alleviates the disturbance of mitochondrial dynamics and downregulates the inflammasome genes exposed to the CEES.

Selective targeting of selenocysteine in thioredoxin reductase by the half mustard 2-chloroethyl ethyl sulfide in lung epithelial cells.

Thioredoxin reductase (TrxR) is a selenocysteine-containing flavoprotein that catalyzes the NADPH-dependent reduction of oxidized thioredoxin and plays a key role in regulating cellular redox homeostasis. In the present studies, we examined the effects of 2-chloroethyl ethyl sulfide (CEES), a model sulfur mustard vesicant, on TrxR in lung epithelial cells. We speculated that vesicant-induced alterations in TrxR contribute to oxidative stress and toxicity. The treatment of human lung A549 epithelial cells with CEES resulted in a time- and concentration-dependent inhibition of TrxR. Using purified rat liver TrxR, we demonstrated that only the reduced enzyme was inhibited and that this inhibition was irreversible. The reaction of TrxR with iodoacetamide, which selectively modifies free thiol or selenol on proteins, was also markedly reduced by CEES, suggesting that CEES induces covalent modification of the reduced selenocysteine-containing active site in the enzyme. This was supported by our findings that recombinant mutant TrxR, in which selenocysteine was replaced by cysteine, was markedly less sensitive to inhibition by CEES and that the vesicant preferentially alkylated selenocysteine in the C-terminal redox motif of TrxR. TrxR also catalyzes quinone redox cycling, a process that generates reactive oxygen species. In contrast to its inhibitory effects on TrxR activity, CEES was found to stimulate redox cycling. Taken together, these data suggest that sulfur mustard vesicants target TrxR and that this may be an important mechanism mediating oxidative stress and tissue injury.