RESUMO
The Ca2+/calmodulin-dependent protein kinase II (CaMKII) is a central regulator of learning and memory, which poses a problem for targeting it therapeutically. Indeed, our study supports prior conclusions that long-term interference with CaMKII signaling can erase pre-formed memories. By contrast, short-term pharmacological CaMKII inhibition with the neuroprotective peptide tatCN19o interfered with learning in mice only mildly and transiently (for less than 1 h) and did not at all reverse pre-formed memories. These results were obtained with ≥500-fold of the dose that protected hippocampal neurons from cell death after a highly clinically relevant pig model of transient global cerebral ischemia: ventricular fibrillation followed by advanced life support and electrical defibrillation to induce the return of spontaneous circulation. Of additional importance for therapy development, our preliminary cardiovascular safety studies in mice and pig did not indicate any concerns with acute tatCN19o injection. Taken together, although prolonged interference with CaMKII signaling can erase memory, acute short-term CaMKII inhibition with tatCN19o did not cause such retrograde amnesia that would pose a contraindication for therapy.
Assuntos
Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina , Memória , Animais , Camundongos , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/antagonistas & inibidores , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Hipocampo/metabolismo , Memória/efeitos dos fármacos , Memória/fisiologia , Neurônios/metabolismo , Fosforilação/fisiologia , Suínos , Peptídeos/farmacologiaRESUMO
BACKGROUND: Methyl mercaptan occurs naturally in the environment and is found in a variety of occupational settings, including the oil, paper, plastics, and pesticides industries. It is a toxic gas and deaths from methyl mercaptan exposure have occurred. The Department of Homeland Security considers it a high threat chemical agent that could be used by terrorists. Unfortunately, no specific treatment exists for methyl mercaptan poisoning. METHODS: We conducted a randomized trial in 12 swine comparing no treatment to intramuscular injection of the vitamin B12 analog cobinamide (2.0 mL, 12.5 mg/kg) following acute inhalation of methyl mercaptan gas. Physiological and laboratory parameters were similar in the control and cobinamide-treated groups at baseline and at the time of treatment. RESULTS: All six cobinamide-treated animals survived, whereas only one of six control animals lived (17% survival) (p = 0.0043). The cobinamide-treated animals returned to a normal breathing pattern by 3.8 ± 1.1 min after treatment (mean ± SD), while all but one animal in the control group had intermittent gasping, never regaining a normal breathing pattern. Blood pressure and arterial oxygen saturation returned to baseline values within 15 minutes of cobinamide-treatment. Plasma lactate concentration increased progressively until death (10.93 ± 6.02 mmol [mean ± SD]) in control animals, and decreased toward baseline (3.79 ± 2.93 mmol [mean ± SD]) by the end of the experiment in cobinamide-treated animals. CONCLUSION: We conclude that intramuscular administration of cobinamide improves survival and clinical outcomes in a large animal model of acute, high dose methyl mercaptan poisoning.
Assuntos
Antídotos/farmacologia , Cobamidas/farmacologia , Compostos de Sulfidrila/toxicidade , Animais , Antídotos/administração & dosagem , Cobamidas/administração & dosagem , Feminino , Exposição por Inalação , Injeções Intramusculares , Masculino , Distribuição Aleatória , SuínosRESUMO
STUDY OBJECTIVE: Cyanide is a deadly poison, particularly with oral exposure, in which larger doses can occur before any symptoms develop. Multiple governmental agencies highlight oral cyanide as an agent that can be used in a terrorist attack because it can be easily weaponized and is readily available. Currently, there are no Food and Drug Administration-approved antidotes specifically for oral cyanide. An oral countermeasure that can neutralize and prevent absorption of cyanide from the gastrointestinal tract after oral exposure is needed. The objective of this study is to determine if the combination of glycine and sodium thiosulfate administered orally is effective in reducing mortality in a large, swine model of oral cyanide toxicity. METHODS: Nine swine (45 to 55 kg) were instrumented, sedated, and stabilized. Potassium cyanide (at 8 mg/kg) in saline solution was delivered as a onetime bolus through an orogastric tube. Three minutes after cyanide administration, animals that were randomized to the treatment group received sodium thiosulfate (508.2 mg/kg, 3.25-M solution) and glycine (30 mg/kg, 3.5-M solution) through an orogastric tube. Survival at 60 minutes was the primary outcome. We compared survival between groups by log-rank Mantel-Cox analysis and trended laboratory results and vital signs. RESULTS: At baseline and treatment, all animals were similar. Survival at 60 minutes was 100% in treated animals compared with 0% in the control group (P=.003). By the study end, defined as death or 60 minutes after cyanide administration, there was a significant difference in the lactate concentration between the treatment and control groups (control 9.43 mmol/L [SD 4.08]; treatment 1.66 mmol/L [SD 0.82]; difference between means 7.69 mmol/L [SD 2.07]; 95% confidence interval difference -14.05 to -1.32). Mean arterial pressure was significantly different between the treatment and control groups at study end (control 26 mm Hg [SD 6.7]; treatment 81 mm Hg [SD 14]; difference between means 55.2 mm Hg [SD 7.1]; 95% confidence interval difference 37.8 to 72.6). pH and oxygen saturation were also significantly different between the treatment and control groups at study end. CONCLUSION: The combination of oral sodium thiosulfate and glycine significantly improved survival and physiologic parameters in a large-animal model of oral cyanide toxicity.
Assuntos
Antídotos/administração & dosagem , Glicina/administração & dosagem , Cianeto de Potássio/intoxicação , Tiossulfatos/administração & dosagem , Administração Oral , Animais , Antídotos/farmacocinética , Modelos Animais de Doenças , Quimioterapia Combinada , Feminino , Glicina/farmacologia , Humanos , Venenos , Distribuição Aleatória , Suínos , Tiossulfatos/farmacologia , Fatores de TempoRESUMO
Inhalation of powerful chemical agents, such as sulfur mustard (SM), can have debilitating pulmonary consequences, such as bronchiolitis obliterans (BO) and parenchymal fibrosis (PF). The underlying pathogenesis of disorders after SM inhalation is not clearly understood, resulting in a paucity of effective therapies. In this study, we evaluated the role of profibrotic pathways involving transforming growth factor-ß (TGF-ß) and platelet-derived growth factor (PDGF) in the development of BO and PF after SM inhalation injury using a rat model. Adult Sprague-Dawley rats were intubated and exposed to SM (1.0 mg/kg), then monitored daily for respiratory distress, oxygen saturation changes, and weight loss. Rats were killed at 7, 14, 21, or 28 days, and markers of injury were determined by histopathology; pulmonary function testing; and assessment of TGF-ß, PDGF, and PAI-1 concentrations. Respiratory distress developed over time after SM inhalation, with progressive hypoxemia, respiratory distress, and weight loss. Histopathology confirmed the presence of both BO and PF, and both gradually worsened with time. Pulmonary function testing demonstrated a time-dependent increase in lung resistance, as well as a decrease in lung compliance. Concentrations of TGF-ß, PDGF, and PAI-1 were elevated at 28 days in lung, BAL fluid, and/or plasma. Time-dependent development of BO and PF occurs in lungs of rats exposed to SM inhalation, and the elevated concentrations of TGF-ß, PDGF, and PAI-1 suggest involvement of these profibrotic pathways in the aberrant remodeling after injury.
Assuntos
Bronquiolite Obliterante/induzido quimicamente , Gás de Mostarda/administração & dosagem , Gás de Mostarda/toxicidade , Fibrose Pulmonar/induzido quimicamente , Administração por Inalação , Animais , Bronquiolite Obliterante/metabolismo , Bronquiolite Obliterante/mortalidade , Bronquiolite Obliterante/patologia , Líquido da Lavagem Broncoalveolar , Substâncias para a Guerra Química/toxicidade , Relação Dose-Resposta a Droga , Inibidor 1 de Ativador de Plasminogênio/metabolismo , Fator de Crescimento Derivado de Plaquetas/metabolismo , Fibrose Pulmonar/metabolismo , Fibrose Pulmonar/mortalidade , Ratos Sprague-Dawley , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Testes de Função Respiratória , Fator de Crescimento Transformador beta1/metabolismo , Redução de Peso/efeitos dos fármacosRESUMO
Autopsy specimens from human victims or experimental animals that die due to acute chlorine gas exposure present features of cardiovascular pathology. We demonstrate acute chlorine inhalation-induced reduction in heart rate and oxygen saturation in rats. Chlorine inhalation elevated chlorine reactants, such as chlorotyrosine and chloramine, in blood plasma. Using heart tissue and primary cardiomyocytes, we demonstrated that acute high-concentration chlorine exposure in vivo (500 ppm for 30 min) caused decreased total ATP content and loss of sarcoendoplasmic reticulum calcium ATPase (SERCA) activity. Loss of SERCA activity was attributed to chlorination of tyrosine residues and oxidation of an important cysteine residue, cysteine-674, in SERCA, as demonstrated by immunoblots and mass spectrometry. Using cardiomyocytes, we found that chlorine-induced cell death and damage to SERCA could be decreased by thiocyanate, an important biological antioxidant, and by genetic SERCA2 overexpression. We also investigated a U.S. Food and Drug Administration-approved drug, ranolazine, used in treatment of cardiac diseases, and previously shown to stabilize SERCA in animal models of ischemia-reperfusion. Pretreatment with ranolazine or istaroxime, another SERCA activator, prevented chlorine-induced cardiomyocyte death. Further investigation of responsible mechanisms showed that ranolazine- and istaroxime-treated cells preserved mitochondrial membrane potential and ATP after chlorine exposure. Thus, these studies demonstrate a novel critical target for chlorine in the heart and identify potentially useful therapies to mitigate toxicity of acute chlorine exposure.
Assuntos
Cloro/toxicidade , Cardiopatias/enzimologia , Exposição por Inalação , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Antioxidantes/farmacologia , Apoptose , Sinalização do Cálcio , Cardiotônicos/farmacologia , Células Cultivadas , Etiocolanolona/análogos & derivados , Etiocolanolona/farmacologia , Cardiopatias/induzido quimicamente , Masculino , Mitocôndrias Cardíacas , Miocárdio/enzimologia , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Ranolazina/farmacologia , Ratos Sprague-Dawley , Tiocianatos/farmacologiaRESUMO
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.
Assuntos
Obstrução das Vias Respiratórias/tratamento farmacológico , Substâncias para a Guerra Química/efeitos adversos , Fibrinolíticos/farmacologia , Gás de Mostarda/efeitos adversos , Insuficiência Respiratória/tratamento farmacológico , Ativador de Plasminogênio Tecidual/farmacologia , Obstrução das Vias Respiratórias/induzido quimicamente , Obstrução das Vias Respiratórias/metabolismo , Obstrução das Vias Respiratórias/patologia , Animais , Substâncias para a Guerra Química/farmacologia , Modelos Animais de Doenças , Fibrina/metabolismo , Humanos , Gás de Mostarda/farmacologia , Ratos , Ratos Sprague-Dawley , Insuficiência Respiratória/induzido quimicamente , Insuficiência Respiratória/metabolismo , Insuficiência Respiratória/patologiaRESUMO
UNLABELLED: Sulfur mustard (SM) inhalation causes airway injury, with enhanced vascular permeability, coagulation, and airway obstruction. The objective of this study was to determine whether recombinant tissue factor pathway inhibitor (TFPI) could inhibit this pathogenic sequence. METHODS: Rats were exposed to the SM analog 2-chloroethyl ethyl sulfide (CEES) via nose-only aerosol inhalation. One hour later, TFPI (1.5mg/kg) in vehicle, or vehicle alone, was instilled into the trachea. Arterial O2 saturation was monitored using pulse oximetry. Twelve hours after exposure, animals were euthanized and bronchoalveolar lavage fluid (BALF) and plasma were analyzed for prothrombin, thrombin-antithrombin complex (TAT), active plasminogen activator inhibitor-1 (PAI-1) levels, and fluid fibrinolytic capacity. Lung steady-state PAI-1 mRNA was measured by RT-PCR analysis. Airway-capillary leak was estimated by BALF protein and IgM, and by pleural fluid measurement. In additional animals, airway cast formation was assessed by microdissection and immunohistochemical detection of airway fibrin. RESULTS: Airway obstruction in the form of fibrin-containing casts was evident in central conducting airways of rats receiving CEES. TFPI decreased cast formation, and limited severe hypoxemia. Findings of reduced prothrombin consumption, and lower TAT complexes in BALF, demonstrated that TFPI acted to limit thrombin activation in airways. TFPI, however, did not appreciably affect CEES-induced airway protein leak, PAI-1 mRNA induction, or inhibition of the fibrinolytic activity present in airway surface liquid. CONCLUSIONS: Intratracheal administration of TFPI limits airway obstruction, improves gas exchange, and prevents mortality in rats with sulfur mustard-analog-induced acute lung injury.
Assuntos
Obstrução das Vias Respiratórias/induzido quimicamente , Obstrução das Vias Respiratórias/prevenção & controle , Substâncias para a Guerra Química/toxicidade , Lipoproteínas/farmacologia , Gás de Mostarda/análogos & derivados , Insuficiência Respiratória/induzido quimicamente , Insuficiência Respiratória/prevenção & controle , Administração por Inalação , Obstrução das Vias Respiratórias/patologia , Animais , Western Blotting , Líquido da Lavagem Broncoalveolar/química , Substâncias para a Guerra Química/farmacocinética , Ensaio de Imunoadsorção Enzimática , Fibrina/metabolismo , Fibrinólise/efeitos dos fármacos , Imunoglobulina M/metabolismo , Imuno-Histoquímica , Indicadores e Reagentes , Masculino , Microdissecção , Gás de Mostarda/farmacocinética , Gás de Mostarda/toxicidade , Proteínas/farmacologia , Protrombina/metabolismo , Ratos , Ratos Sprague-Dawley , Proteínas Recombinantes/farmacologia , Insuficiência Respiratória/patologiaRESUMO
BACKGROUND: Traumatic injury with subsequent hemorrhage is one of the leading causes of mortality among military personnel and civilians alike. Post traumatic hemorrhage accounts for 40-50% of deaths in severe trauma patients occurring secondary to direct vessel injury or the development of trauma induced coagulopathy (TIC). Hyperfibrinolysis plays a major role in TIC and its presence increases a patient's risk of mortality. Early therapeutic intervention with intravenous (IV) tranexamic acid (TXA) prevents development of hyperfibrinolysis and subsequent TIC leading to decreased mortality. However, obtaining IV access in an austere environment can be challenging. In this study, we evaluated the efficacy of intramuscular (IM) versus IV TXA at preventing hyperfibrinolysis in a hemorrhaged swine. METHODS: Yorkshire cross swine were randomized on the day of study to receive IM or IV TXA or no treatment. Swine were sedated, intubated, and determined to be hemodynamically stable prior to experimentation. Controlled hemorrhaged was induced by the removal of 30% total blood volume. After hemorrhage, swine were treated with 1000 mg of IM or IV TXA. Control animals received no treatment. Thirty minutes post TXA treatment, fibrinolysis was induced with a 50 mg bolus of tissue plasminogen activator (tPA). Blood samples were collected to evaluate blood TXA concentrations, blood gases, blood chemistry, and fibrinolysis. RESULTS: Blood TXA concentrations were significantly different between administration routes at the early timepoints, but were equivalent by 20 minutes after injection, remaining consistently elevated for up to three hours post administration. Induction of fibrinolysis resulted in 87.18 ± 4.63% lysis in control animals, compared to swine treated with IM TXA 1.96 ± 2.66 % and 1.5 ± 0.42% lysis in the IV TXA group. CONCLUSION: In the large swine model of hemorrhage with hyperfibrinolysis, IM TXA is bioequivalent and equally efficacious in preventing hyperfibrinolysis as IV TXA administration.
RESUMO
Cyanide-a fast-acting poison-is easy to obtain given its widespread use in manufacturing industries. It is a high-threat chemical agent that poses a risk of occupational exposure in addition to being a terrorist agent. FDA-approved cyanide antidotes must be given intravenously, which is not practical in a mass casualty setting due to the time and skill required to obtain intravenous access. Glyoxylate is an endogenous metabolite that binds cyanide and reverses cyanide-induced redox imbalances independent of chelation. Efficacy and biochemical mechanistic studies in an FDA-approved preclinical animal model have not been reported. Therefore, in a swine model of cyanide poisoning, we evaluated the efficacy of intramuscular glyoxylate on clinical, metabolic, and biochemical endpoints. Animals were instrumented for continuous hemodynamic monitoring and infused with potassium cyanide. Following cyanide-induced apnea, saline control or glyoxylate was administered intramuscularly. Throughout the study, serial blood samples were collected for pharmacokinetic, metabolite, and biochemical studies, in addition, vital signs, hemodynamic parameters, and laboratory values were measured. Survival in glyoxylate-treated animals was 83% compared with 12% in saline-treated control animals (p < .01). Glyoxylate treatment improved physiological parameters including pulse oximetry, arterial oxygenation, respiration, and pH. In addition, levels of citric acid cycle metabolites returned to baseline levels by the end of the study. Moreover, glyoxylate exerted distinct effects on redox balance as compared with a cyanide-chelating countermeasure. In our preclinical swine model of lethal cyanide poisoning, intramuscular administration of the endogenous metabolite glyoxylate improved survival and clinical outcomes, and ameliorated the biochemical effects of cyanide.
Assuntos
Cianetos , Intoxicação , Suínos , Animais , Cianetos/toxicidade , Modelos Animais de Doenças , Antídotos/farmacologia , Antídotos/uso terapêutico , Hemodinâmica , Glioxilatos/uso terapêutico , Intoxicação/tratamento farmacológicoRESUMO
The Ca 2+ /calmodulin-dependent protein kinase II (CaMKII) is a central regulator of learning and memory, which poses a problem for targeting it therapeutically. Indeed, our study supports prior conclusions that long-term interference with CaMKII signaling can erase pre-formed memories. By contrast, short-term pharmacological CaMKII inhibition with tatCN19o interfered with learning in mice only mildly and transiently (for less than 1 h) and did not at all reverse pre-formed memories. This was at â¥500fold of the dose that protected hippocampal neurons from cell death after a highly clinically relevant pig model of transient global cerebral ischemia: ventricular fibrillation followed by advanced life support and electrical defibrillation to induce return of spontaneous circulation. Of additional importance for therapeutic development, cardiovascular safety studies in mice and pig did not indicate any concerns with acute tatCN19o injection. Taken together, even though prolonged interference with CaMKII signaling can erase memory, acute short-term CaMKII inhibition with tatCN19o did not cause such retrograde amnesia that would pose a contraindication for therapy.
RESUMO
Acute lung injury is a principal cause of morbidity and mortality in response to mustard gas (SM) inhalation. Obstructive, fibrin-containing airway casts have recently been reported in a rat inhalation model employing the SM analog 2-chloroethyl ethyl sulfide (CEES). The present study was designed to identify the mechanism(s) causing activation of the coagulation cascade after CEES-induced airway injury. Here we report that CEES inhalation elevates tissue factor (TF) activity and numbers of detached epithelial cells present in lavage fluid (BALF) from rats after exposure (18 h). In vitro studies using 16HBE cells, or with rat BALF, indicated that detached epithelial cells could convert factor X (FX) to the active form FXa when incubated with factor VII and could elicit rapid clotting of plasma. In addition, immunocytochemical analysis demonstrated elevated cell surface (TF) expression on CEES-exposed 16HBE cells as a function of time. However, total cell TF expression did not increase. Since membrane surfaces bearing TF are important determinants of clot initiation, anticoagulants directed against these entities were tested for ability to limit plasma clotting or FX activation capacity of BALF or culture media. Addition of tifacogin, a TF pathway inhibitor, effectively blocked either activity, demonstrating that the procoagulant actions of CEES were TF pathway dependent. Lactadherin, a protein capable of competing with clotting factors for phospholipid-binding sites, was partially effective in limiting these procoagulant actions. These findings indicate that TF pathway inhibition could be an effective strategy to prevent airway obstruction after SM or CEES inhalation.
Assuntos
Obstrução das Vias Respiratórias/induzido quimicamente , Brônquios/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Gás de Mostarda/análogos & derivados , Tromboplastina/metabolismo , Animais , Antígenos de Superfície/metabolismo , Brônquios/citologia , Brônquios/enzimologia , Líquido da Lavagem Broncoalveolar/citologia , Linhagem Celular Transformada , Substâncias para a Guerra Química/toxicidade , Modelos Animais de Doenças , Células Epiteliais/citologia , Células Epiteliais/enzimologia , Fator VII/metabolismo , Fator Xa/metabolismo , Humanos , Exposição por Inalação , Masculino , Proteínas do Leite/metabolismo , Gás de Mostarda/toxicidade , Proteínas/farmacologia , Ratos , Ratos Sprague-Dawley , Tromboplastina/antagonistas & inibidores , Fatores de TempoRESUMO
Cyanide, hydrogen sulfide, and methanethiol are common toxic inhalation agents that inhibit mitochondrial cytochrome c oxidase and result in cellular hypoxia, cytotoxic anoxia, apnea, respiratory failure, cardiovascular collapse, seizure and potentially death. While all are occupational gas exposure hazards that have the potential to cause mass casualties from industrial accidents or acts of terrorism, only cyanide has approved antidotes, and each of these has major limitations, including difficult administration in mass-casualty settings. While bisaminotetrazole cobinamide (Cbi(AT)2) has recently gained attention because of its efficacy in treating these metabolic poisons, there is no method available for the analysis of Cbi(AT)2 in any biological matrix. Hence, in this study, a simple and rapid liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for the analysis of Cbi(AT)2 in swine plasma. The method is extremely simple, consisting of protein precipitation, separation and drying of the supernatant, reconstitution in an aqueous solvent, and LC-MS/MS analysis. The method produced an LOD of 0.3 µM with a wide dynamic range (2 - 500 µM). Inter- and intraassay accuracies (100 ± 12 % and 100 ± 19 %, respectively) were acceptable and the precision (<12 % and < 9 % relative standard deviation, respectively) was good. The developed method was used to analyze Cbi(AT)2 from treated swine and the preliminary pharmacokinetic parameters showed impressive antidotal behavior, most notably a long estimated elimination half-life (t1/2 = 37.5 h). This simple and rapid method can be used to facilitate the development of Cbi(AT)2 as a therapeutic against toxic cyanide, hydrogen sulfide and methanethiol exposure.
Assuntos
Antídotos , Sulfeto de Hidrogênio , Animais , Antídotos/uso terapêutico , Cromatografia Líquida , Cobamidas , Cianetos , Cianeto de Hidrogênio , Compostos de Sulfidrila , Sulfetos , Suínos , Espectrometria de Massas em Tandem/métodosRESUMO
CONTEXT: Hydrogen cyanide and methanethiol are two toxic gases that inhibit mitochondrial cytochrome c oxidase. Cyanide is generated in structural fires and methanethiol is released by decaying organic matter. Current treatments for cyanide exposure do not lend themselves to treatment in the field and no treatment exists for methanethiol poisoning. Sodium tetrathionate (tetrathionate), a product of thiosulfate oxidation, could potentially serve as a cyanide antidote, and, based on its chemical structure, we hypothesized it could react with methanethiol. RESULTS: We show that tetrathionate, unlike thiosulfate, reacts directly with cyanide in vitro under physiological conditions, and based on rabbit studies where we monitor cyanide poisoning in real-time, tetrathionate likely reacts directly with cyanide in vivo. We found that tetrathionate administered by intramuscular injection rescues >80% of juvenile, young adult, and old adult mice from exposure to inhaled hydrogen cyanide gas that is >80% lethal. Tetrathionate also rescued young adult rabbits from intravenously administered sodium cyanide. Tetrathionate was reasonably well-tolerated by mice and rats, yielding a therapeutic index of â¼5 in juvenile and young adult mice, and â¼3.3 in old adult mice; it was non-mutagenic in Chinese Hamster ovary cells and by the Ames bacterial test. We found by gas chromatography-mass spectrometry that both tetrathionate and thiosulfate react with methanethiol to generate dimethyldisulfide, but that tetrathionate was much more effective than thiosulfate at recovering intracellular ATP in COS-7 cells and rescuing mice from a lethal exposure to methanethiol gas. CONCLUSION: We conclude that tetrathionate has the potential to be an effective antidote against cyanide and methanethiol poisoning.
Assuntos
Antídotos , Ácido Tetratiônico , Animais , Antídotos/uso terapêutico , Células CHO , Cricetinae , Cricetulus , Cianetos , Humanos , Camundongos , Coelhos , Ratos , Compostos de Sulfidrila , TiossulfatosRESUMO
BACKGROUND: Cyanide is a rapid acting, lethal, metabolic poison and remains a significant threat. Current FDA-approved antidotes are not amenable or efficient enough for a mass casualty incident. OBJECTIVE: The objective of this study is to evaluate short and long-term efficacy of intramuscular aqueous dimethyl trisulfide (DMTS) on survival and clinical outcomes in a swine model of cyanide exposure. METHODS: Anesthetized swine were instrumented and acclimated until breathing spontaneously. Potassium cyanide infusion was initiated and continued until 5 min after the onset of apnea. Subsequently, animals were treated with intramuscular DMTS (n = 11) or saline control (n = 10). Laboratory values and DMTS blood concentrations were assessed at various time points and physiological parameters were monitored continuously until the end of the experiment unless death occurred. A subset of animals treated with DMTS (n = 5) were survived for 7 days to evaluate muscle integrity by repeat biopsy and neurobehavioral outcomes. RESULTS: Physiological parameters and time to apnea were similar in both groups at baseline and at time of treatment. Survival in the DMTS-treated group was 90% and 30% in saline controls (p = 0.0034). DMTS-treated animals returned to breathing at 12.0 ± 10.4 min (mean ± SD) compared to 22.9 ± 7.0 min (mean ± SD) in the 3 surviving controls. Blood collected prior to euthanasia showed improved blood lactate concentrations in the DMTS treatment group; 5.47 ± 2.65 mmol/L vs. 9.39 ± 4.51 mmol/L (mean ± SD) in controls (p = 0.0310). Low concentrations of DMTS were detected in the blood, gradually increasing over time with no elimination phase observed. There was no mortality, histological evidence of muscle trauma, or observed adverse neurobehavioral outcomes, in DMTS-treated animals survived to 7 days. CONCLUSION: Intramuscular administration of aqueous DMTS improves survival following cyanide poisoning with no observed long-term effects on muscle integrity at the injection site or adverse neurobehavioral outcomes.
Assuntos
Antídotos , Sulfetos , Animais , Antídotos/farmacologia , Antídotos/uso terapêutico , Cianetos , Humanos , Cianeto de Potássio , SuínosRESUMO
Although cyanide's biological effects are pleiotropic, its most obvious effects are as a metabolic poison. Cyanide potently inhibits cytochrome c oxidase and potentially other metabolic enzymes, thereby unleashing a cascade of metabolic perturbations that are believed to cause lethality. From systematic screens of human metabolites using a zebrafish model of cyanide toxicity, we have identified the TCA-derived small molecule glyoxylate as a potential cyanide countermeasure. Following cyanide exposure, treatment with glyoxylate in both mammalian and non-mammalian animal models confers resistance to cyanide toxicity with greater efficacy and faster kinetics than known cyanide scavengers. Glyoxylate-mediated cyanide resistance is accompanied by rapid pyruvate consumption without an accompanying increase in lactate concentration. Lactate dehydrogenase is required for this effect which distinguishes the mechanism of glyoxylate rescue as distinct from countermeasures based solely on chemical cyanide scavenging. Our metabolic data together support the hypothesis that glyoxylate confers survival at least in part by reversing the cyanide-induced redox imbalances in the cytosol and mitochondria. The data presented herein represent the identification of a potential cyanide countermeasure operating through a novel mechanism of metabolic modulation.
Assuntos
Glioxilatos , Peixe-Zebra , Animais , Cianetos/toxicidade , Mamíferos , Ácido PirúvicoRESUMO
The inhalation of sulfur mustard (SM) causes substantial deposition in the nasal region. However, specific injury has not been characterized. 2-chloroethyl ethyl sulfide (CEES) is an SM analogue used to model injury and screen potential therapeutics. After the inhalation of CEES, damage to the olfactory epithelium (OE) was extensive. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling-positive cells were present by 4 hours, and maximal at 18-72 hours. Cleaved caspase 3 immunohistochemistry (IHC) was maximal at 18 hours after the inhalation of 5% CEES. Olfactory marker protein (OMP)-positive olfactory neurons were markedly decreased at 18 hours. IHC-positive cells for 3-nitrotyrosine (3-NT) within epithelium were elevated by 8 hours, waning by 18 hours, and absent by 72 hours. AEOL 10150, a catalytic manganoporphyrin antioxidant, administered both subcutaneously (5 mg/kg) and intranasally (50 µM, "combined treatment"), decreased OE injury. CEES-induced increases in markers of cell death were decreased by combined treatment involving AEOL 10150. CEES-induced changes in OMP and 3-NT immunostaining were markedly improved by combined treatment involving AEOL 10150. The selective inducible nitric oxide synthase inhibitor 1400W (5 mg/kg, subcutaneous), administered 1 hour after inhalation and thereafter every 4 hours (five doses), also reduced OE damage with improved OMP and 3-NT staining. Taken together, these data indicate that reactive oxygen and nitrogen species are important mediators in CEES-induced nasal injury.
Assuntos
Células Epiteliais/efeitos dos fármacos , Gás de Mostarda/análogos & derivados , Cavidade Nasal/efeitos dos fármacos , Mucosa Olfatória/efeitos dos fármacos , Mucosa Olfatória/lesões , Espécies Reativas de Nitrogênio/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Administração por Inalação , Animais , Antioxidantes/farmacologia , Apoptose , Western Blotting , Proliferação de Células , Células Epiteliais/metabolismo , Técnicas Imunoenzimáticas , Masculino , Metaloporfirinas/farmacologia , Gás de Mostarda/administração & dosagem , Gás de Mostarda/toxicidade , Cavidade Nasal/lesões , Cavidade Nasal/metabolismo , Óxido Nítrico Sintase Tipo II/antagonistas & inibidores , Óxido Nítrico Sintase Tipo II/metabolismo , Mucosa Olfatória/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Sprague-DawleyRESUMO
RATIONALE: Sulfur mustard (SM) is a frequently used chemical warfare agent, even in modern history. SM inhalation causes significant respiratory tract injury, with early complications due to airway obstructive bronchial casts, akin to those seen after smoke inhalation and in single-ventricle physiology. This process with SM is poorly understood because animal models are unavailable. OBJECTIVES: To develop a rat inhalation model for airway obstruction with the SM analog 2-chloroethyl ethyl sulfide (CEES), and to investigate the pathogenesis of bronchial cast formation. METHODS: Adult rats were exposed to 0, 5, or 7.5% CEES in ethanol via nose-only aerosol inhalation (15 min). Airway microdissection and confocal microscopy were used to assess cast formation (4 and 18 h after exposure). Bronchoalveolar lavage fluid (BALF) retrieval and intravascular dye injection were done to evaluate vascular permeability. MEASUREMENTS AND MAIN RESULTS: Bronchial casts, composed of abundant fibrin and lacking mucus, occluded dependent lobar bronchi within 18 hours of CEES exposure. BALF contained elevated concentrations of IgM, protein, and fibrin. Accumulation of fibrin-rich fluid in peribronchovascular regions (4 h) preceded cast formation. Monastral blue dye leakage identified bronchial vessels as the site of leakage. CONCLUSIONS: After CEES inhalation, increased permeability from damaged bronchial vessels underlying damaged airway epithelium leads to the appearance of plasma proteins in both peribronchovascular regions and BALF. The subsequent formation of fibrin-rich casts within the airways then leads to airways obstruction, causing significant morbidity and mortality acutely after exposure.
Assuntos
Obstrução das Vias Respiratórias/induzido quimicamente , Brônquios/irrigação sanguínea , Brônquios/efeitos dos fármacos , Substâncias para a Guerra Química/toxicidade , Gás de Mostarda/toxicidade , Animais , Western Blotting , Líquido da Lavagem Broncoalveolar , Permeabilidade Capilar/efeitos dos fármacos , Modelos Animais de Doenças , Fibrina/efeitos dos fármacos , Imunoglobulina M/efeitos dos fármacos , Exposição por Inalação , Masculino , Microdissecção , Microscopia Confocal , Ratos , Ratos Sprague-DawleyRESUMO
The superoxide dismutase (SOD)-mimetic effectiveness of [meso-tetrakis (R)porphyrinato]manganese with R = 1,3-di-N-ethylimidazolium-2-yl (Mn-TDEIP), 1,3-di-N-methylimidazolium-2-yl (Mn-TDMIP), 1,3-di-N-propylimidazolium-2-yl (Mn-TDPIP), N-ethyl-2-pyridyl (Mn-T2EPyP), 4-sulphonatophenyl (Mn-TSP), 1-methyl-4-pyridyl (Mn-T4PyP), 4-carboxyphenyl (Mn-TBAP), and ß-octabromomeso-tetrakis(4-carboxyphenyl porphyrinato)manganese (MnBr8TBAP) was compared with Cu, Zn SOD. Superoxide generated by reaction of xanthine oxidase with hypoxanthine was trapped with 5-tert-butoxycarbonyl-5-methyl-1-pyrroline N-oxide (BMPO), forming BMPO-OOH, which was monitored by electron paramagnetic resonance. Manganoporphyrins with redox potentials ranging from -0.190 to 0.346 V relative to the standard hydrogen electrode were selected for this study. With 0.1 µM manganoporphyrins and 20 mM BMPO, the effectiveness of the manganoporphyrins in inhibiting formation of BMPO-OOH increases in the order Mn-TSP < Mn-TBAP < MnBr8TBAP < Mn-T4PyP < Mn-T2EPyP < Mn-TDEIP ~ Mn-TDMIP ~ Mn-TDPIP ~ Cu, Zn SOD. However, at higher concentrations of manganoporphyrin and BMPO, a BMPO-OH signal was observed. The formation of BMPO-OH was not inhibited by catalase or dimethylsulfoxide, which demonstrated that it was not produced from hydroxyl radical. The artifactual formation of BMPO-OH is attributed to oxidation of the water adduct of BMPO by the manganoporphyrins or decomposition of BMPO-OOH. Although spin trapping is an effective method for evaluating SOD-mimetic efficacy, caution must be exercised to ensure that artifact signals are not interpreted improperly.
RESUMO
INTRODUCTION: Cyanide is a deadly poison, particularly with oral exposure where larger doses can occur before symptoms develop. Prior studies and multiple governmentagencies highlight oral cyanide as an agent with the potential for use in a terrorist attack. Currently, there are no FDA approved antidotes specific to oralcyanide. An oral countermeasure that can neutralize and prevent absorption of cyanide from the GI tract after oral exposure is needed. Our objective was toevaluate the efficacy of oral sodium thiosulfate on survival and clinical outcomes in a large, swine model of severe cyanide toxicity. METHODS: Swine (45-55kg) were instrumented, sedated, and stabilized. Potassium cyanide (8 mg/kg KCN) in saline was delivered as a one-time bolus via an orogastric tube. Three minutes after cyanide, animals randomized to the treatment group received sodium thiosulfate (510 mg/kg, 3.25 M solution) via orogastric tube. Our primary outcome was survival at 60 minutes after exposure. We compared survival between groups by log-rank, Mantel-Cox analysis and trended labs and vital signs. RESULTS: At baseline and time of treatment all animals had similar weights, vital signs, and laboratory values. Survival at 60 min was 100% in treated animals compared to 0% in the control group (p=0.0027). Animals in the control group became apneic and subsequently died by 35.0 min (20.2,48.5) after cyanide exposure. Mean arterial pressure was significantly higher in the treatment group compared to controls (p=0.008). Blood lactate (p=0.02) and oxygen saturation (p=0.02) were also significantly different between treatment and control groups at study end. CONCLUSION: Oral administration of sodium thiosulfate improved survival, blood pressure, respirations, and blood lactate concentrations in a large animal model of acute oral cyanide toxicity.
Assuntos
Antídotos/uso terapêutico , Cianetos/toxicidade , Tiossulfatos/uso terapêutico , Administração Oral , Animais , Humanos , Modelos Animais , Suínos , Tiossulfatos/administração & dosagem , Resultado do TratamentoRESUMO
Plant parasites and soilborne pathogens directly reduce the overall yield of crops, vegetables, and fruits, negatively impacting the market demand for these products and their net profitability. While preplant soil fumigation helps maintain the consistent production quality of high-value cash crops, most soil fumigants are toxic to off-target species, including humans. Dimethyl disulfide (DMDS) has recently been introduced as a relatively low toxicity soil fumigant. Although DMDS exhibits low toxicity compared to other soil fumigants, it is volatile and exposure can cause eye, nasal, and upper respiratory tract irritation, skin irritation, nausea, dizziness, headache, and fatigue. While there is one analysis method available for DMDS from biological matrices, it has significant disadvantages. Hence, in this study, a dynamic headspace gas chromatography-mass spectroscopy (DHS-GC-MS) method was developed for the analysis of DMDS in swine whole blood. This method is highly sensitive and requires only three steps: 1) acid denaturation, 2) addition of internal standard, and 3) DHS-GC-MS analysis. The method produced a wide linear range from 0.1 - 200 µM with an excellent limit of detection of 30 nM. Intra- and interassay accuracy (100±14% and 100±11%, respectively) and precision (<5% and <6% relative standard deviation, respectively) were also excellent. The method worked well to quantify the DMDS levels in the blood of dimethyl trisulfide (DMTS)-treated swine (i.e., DMDS is a byproduct of DMTS treatment) with no interfering substances at or around the retention time of DMDS (i.e., 2.7 min). This simple, rapid, and extremely sensitive method can be used for the quantification of DMDS levels in blood to verify exposure to DMDS or to monitor levels of DMDS following DMTS treatment (e.g., for cyanide poisoning).