RESUMEN
Geographic isolation and other factors result in evolution-driven diversity of the enzymatic composition of venom of pit vipers in the same genus. The present investigation sought to characterize venoms obtained from such genetically diverse Ovophis and Trimeresurus pit vipers utilizing thrombelastographic coagulation kinetic analyses. The coagulation kinetics of human plasma were assessed after exposure to venom obtained from two Ovophis and three Trimeresurus species. The potency of each venom was defined (µg/mL required to equivalently change coagulation); additionally, venoms were exposed to carbon monoxide (CO) or a metheme-inducing agent to modulate any enzyme-associated heme. All venoms had fibrinogenolytic activity, with four being CO-inhibitable. While Ovophis venoms had similar potency, one demonstrated the presence of a thrombin-like activity, whereas the other demonstrated a thrombin-generating activity. There was a 10-fold difference in potency and 10-fold different vulnerability to CO inhibition between the Trimeresurus species. Metheme formation enhanced fibrinogenolytic-like activity in both Ovophis species venoms, whereas the three Trimeresurus species venoms had fibrinogenolytic-like activity enhanced, inhibited, or not changed. This novel "venom kinetomic" approach has potential to identify clinically relevant enzymatic activity and assess efficacy of antivenoms between genetically and geographically diverse species.
Asunto(s)
Venenos de Crotálidos/toxicidad , Crotalinae , Hemo/metabolismo , Animales , Coagulación Sanguínea/efectos de los fármacos , Monóxido de Carbono , Humanos , TromboelastografíaRESUMEN
Envenomation by hemotoxic enzymes continues to be a major cause of morbidity and mortality throughout the world. With regard to treatment, the gold standard to abrogate coagulopathy caused by these venoms is still the administration of antivenom; however, despite antivenom therapy, coagulopathy still occurs and recurs. Of interest, this laboratory has demonstrated in vitro and in vivo that coagulopathy inducing venom derived from snakes of the family Viperidae exposed to carbon monoxide (CO) is inhibited, potentially by an attached heme. The present investigation sought to determine if venoms derived from snakes of the Elapidae family (taipans and cobras) could also be inhibited with CO or with the metheme inducing agent, O-phenylhydroxylamine (PHA). Assessing changes in coagulation kinetics of human plasma with thrombelastography, venoms from Elapidae snakes were exposed in isolation to CO (five species) or PHA (one specie) and placed in human plasma to assess changes in procoagulant or anticoagulant activity. The procoagulant activity of two taipan venoms and anticoagulant activity of three cobra venoms were significantly inhibited by CO. The venom of the inland taipan was also inhibited by PHA. In sum, these data demonstrate indirectly that the biometal heme is likely bound to these disparate venoms as an intermediary modulatory molecule. In conclusion, CO may not just be a potential therapeutic agent to treat envenomation but also may be a potential modulator of heme as a protective mechanism for venomous snakes against injury from their own proteolytic venoms.
Asunto(s)
Antivenenos/farmacología , Coagulación Sanguínea/efectos de los fármacos , Monóxido de Carbono/farmacología , Venenos Elapídicos/antagonistas & inhibidores , Hemo/metabolismo , Compuestos Organometálicos/farmacología , Animales , Antivenenos/química , Monóxido de Carbono/química , Venenos Elapídicos/sangre , Elapidae/fisiología , Hemo/química , Humanos , Hidroxilaminas/farmacología , Cinética , Compuestos Organometálicos/química , Soluciones , TromboelastografíaRESUMEN
In addition to degrading fibrinogen as a source of consumptive coagulopathy, rattlesnake venom has also been demonstrated to enhance fibrinolysis and degrade alpha-2-antiplasmin. The goals of this investigation was to characterize the kinetic fibrinolytic profile of Crotalus atrox venom in the absence and presence of tissue-type plasminogen activator (tPA), and to also ascertain if iron and carbon monoxide (CO, a positive modulator of alpha-2-antiplasmin) could attenuate venom-enhanced fibrinolysis. Utilizing thrombelastographic methods, the coagulation and fibrinolytic kinetic profiles of human plasma exposed to C. atrox venom (0-2âµg/ml) were determined in the absence or presence of tPA (0-100âIU/ml). Then, either separately or in combination, plasma was exposed to iron (ferric chloride, 10âµmol/l) or CO (carbon monoxide-releasing molecule-2, 100âµmol/l) prior to incubation with venom; the plasma sample was subsequently subjected to thrombelastographic analysis with addition of tPA. Venom exposure in the absence of tPA did not result in detectable fibrinolysis. In the presence of tPA, venom markedly enhanced fibrinolysis. Iron and CO, markedly attenuated venom enhancement of fibrinolysis. C. atrox venom enhances tPA-mediated fibrinolysis, and interventions that enhance/protect alpha-2-antiplasmin activity significantly attenuate venom-enhanced fibrinolysis. Future preclinical investigation is required to determine if iron and CO can attenuate venom-mediated degradation of alpha-2-antiplasmin-dependent fibrinolytic resistance.
Asunto(s)
Monóxido de Carbono/farmacología , Cloruros/farmacología , Venenos de Crotálidos/antagonistas & inhibidores , Compuestos Férricos/farmacología , Fibrinólisis/efectos de los fármacos , Fibrinolíticos/farmacología , Hemostáticos/farmacología , Activador de Tejido Plasminógeno/antagonistas & inhibidores , Animales , Monóxido de Carbono/química , Venenos de Crotálidos/farmacología , Crotalus/metabolismo , Tiempo de Lisis del Coágulo de Fibrina , Fibrinógeno/metabolismo , Humanos , Compuestos Organometálicos/química , Compuestos Organometálicos/farmacología , Tromboelastografía , Activador de Tejido Plasminógeno/farmacología , alfa 2-Antiplasmina/metabolismo , alfa 2-Antiplasmina/farmacologíaRESUMEN
Morbid obesity is associated with significant thrombophilia. Of interest, adipocytes obtained from obese patients have increased heme oxygenase (Hmox) activity, the endogenous enzyme responsible for carbon monoxide (CO) production. Given that CO enhances plasmatic coagulation, we determined whether morbidly obese patients undergoing bariatric surgery had an increase in endogenous CO and plasmatic hypercoagulability. CO was determined by noninvasive pulse oximetry measurement of carboxyhemoglobin (COHb). A thrombelastographic method to assess plasma coagulation kinetics and formation of carboxyhemefibrinogen (COHF) was utilized. Nonsmoking bariatric patients (nâ=â20, BMI 47â±â8âkg/m, meanâ±âSD) had abnormally increased COHb concentrations of 2.7â±â1.9%, indicative of Hmox upregulation. When coagulation kinetics of these bariatric patients were compared with values obtained from normal individuals' (nâ=â30) plasma, 70% (95% confidence interval 45.7-88.1%) had abnormally great velocity of clot formation, abnormally large clot strength, and COHF formation. Future investigation of Hmox-derived CO in the pathogenesis of obesity-related thrombophilia is warranted.
Asunto(s)
Cirugía Bariátrica/métodos , Monóxido de Carbono/sangre , Hemo Oxigenasa (Desciclizante)/metabolismo , Obesidad Mórbida/sangre , Trombofilia/etiología , Adulto , Cirugía Bariátrica/efectos adversos , Carboxihemoglobina/metabolismo , Femenino , Humanos , Masculino , Persona de Mediana Edad , Obesidad Mórbida/enzimología , Obesidad Mórbida/cirugía , Trombofilia/sangre , Regulación hacia Arriba , Adulto JovenRESUMEN
Chronic hemodialysis is associated with significant thrombophilia. Of interest, hemodialysis patients have increased carboxyhemoglobin (COHb) and exhaled carbon monoxide (CO), signs of upregulated heme oxygenase (Hmox) activity. Given that CO enhances plasmatic coagulation, we determined whether patients requiring chronic hemodialysis had an increase in endogenous CO, plasmatic hypercoagulability and decreased fibrinolytic vulnerability. Carbon monoxide was determined by noninvasive pulse oximetry measurement of COHb. Blood samples were obtained just before hemodialysis. Thrombelastographic methods to assess plasma coagulation kinetics, fibrinolytic kinetics, and formation of carboxyhemefibrinogen (COHF) were used. Hemodialysis patients (n = 45) had abnormally increased COHb concentrations of 2.2 ± 1.9%, indicative of Hmox upregulation. Coagulation and fibrinolytic parameter normal values were determined with normal individual (n = 30) plasma. Thirty-seven patients of the hemodialysis cohort had COHF formation (82.2%, [67.9%-92.0%]; mean, [95% confidence interval]), and many of this group of patients had abnormally great velocity of clot growth (73.3%, [58.1%-85.4%]) and strength (75.6%, [60.5%-87.1%]). Furthermore, over half of COHF positive patients had a hypofibrinolytic state, evidenced by an abnormally prolonged time to maximum rate of lysis (53.3%, [37.9%-68.6%]) and clot lysis time (64.4%, [48.8%-78.1%]). Carbon monoxide enhanced coagulation and diminished fibrinolytic vulnerability in hemodialysis patients. Future investigation of hemodialysis, CO-related thrombophilia is warranted.
Asunto(s)
Monóxido de Carbono/sangre , Fibrinólisis , Diálisis Renal/efectos adversos , Trombofilia/sangre , Trombofilia/etiología , Adulto , Anciano , Coagulación Sanguínea , Carboxihemoglobina/metabolismo , Femenino , Fibrinógeno/análogos & derivados , Fibrinógeno/metabolismo , Humanos , Fallo Renal Crónico/sangre , Fallo Renal Crónico/complicaciones , Fallo Renal Crónico/terapia , Cinética , Masculino , Persona de Mediana Edad , Tromboelastografía , Adulto JovenRESUMEN
Although cancer-mediated changes in hemostatic proteins unquestionably promote hypercoagulation, the effects of neoplasia on fibrinolysis in the circulation are less well defined. The goals of the present investigation were to determine if plasma obtained from patients with breast, lung, pancreas and colon cancer was less or more susceptible to lysis by tissue-type plasminogen activator (tPA) compared to plasma obtained from normal individuals. Archived plasma obtained from patients with breast (nâ=â18), colon/pancreas (nâ=â27) or lung (nâ=â19) was compared to normal individual plasma (nâ=â30) using a thrombelastographic assay that assessed fibrinolytic vulnerability to exogenously added tPA. Plasma samples were activated with tissue factor/celite, had tPA added, and had data collected until clot lysis occurred. Additional, similar samples had potato carboxypeptidase inhibitor added to assess the role played by thrombin-activatable fibrinolysis inhibitor in cancer-modulated fibrinolysis. Rather than inflicting a hypofibrinolytic state, the three groups of cancers demonstrated increased vulnerability to tPA (e.g. decreased time to lysis, increased speed of lysis, decreased clot lysis time). However, hypercoagulation manifested as increased speed of clot formation and strength compensated for enhanced fibrinolytic vulnerability, resulting in a clot residence time that was not different from normal individual thrombi. In sum, enhanced hypercoagulability associated with cancer was in part diminished by enhanced fibrinolytic vulnerability to tPA.
Asunto(s)
Fibrinólisis/efectos de los fármacos , Neoplasias/sangre , Activador de Tejido Plasminógeno/farmacología , Adulto , Neoplasias de la Mama/sangre , Neoplasias de la Mama/patología , Neoplasias del Colon/sangre , Neoplasias del Colon/patología , Femenino , Humanos , Neoplasias Pulmonares/sangre , Neoplasias Pulmonares/patología , Masculino , Persona de Mediana Edad , Neoplasias/patología , Neoplasias Pancreáticas/sangre , Neoplasias Pancreáticas/patología , Adulto JovenRESUMEN
Colon and pancreatic cancer are associated with significant thrombophilia. Colon and pancreas tumor cells have an increase in hemeoxygenase-1 (HO-1) activity, the endogenous enzyme responsible for carbon monoxide production. Given that carbon monoxide enhances plasmatic coagulation, we determined if patients undergoing resection of colon and pancreatic tumors had an increase in endogenous carbon monoxide and plasmatic hypercoagulability. Patients with colon (n = 17) and pancreatic (n = 10) tumors were studied. Carbon monoxide was determined by the measurement of carboxyhemoglobin (COHb). A thrombelastographic method to assess plasma coagulation kinetics and formation of carboxyhemefibrinogen (COHF) was utilized. Nonsmoking patients with colon and pancreatic tumors had abnormally increased COHb concentrations of 1.4 ± 0.9 and 1.9 ± 0.7%, respectively, indicative of HO-1 upregulation. Coagulation analyses comparing both tumor groups demonstrated no significant differences in any parameter; thus the data were combined for the tumor groups for comparison with 95% confidence interval values obtained from normal individuals (n = 30) plasma. Seventy percent of tumor patients had a velocity of clot formation greater than the 95% confidence interval value of normal individuals, with 53% of this hypercoagulable group also having COHF formation. Further, 67% of tumor patients had clot strength that exceeded the normal 95% confidence interval value, and 56% of this subgroup had COHF formation. Finally, 63% of all tumor patients had COHF formation. Future investigation of HO-1-derived carbon monoxide in the pathogenesis of colon and pancreatic tumor-related thrombophilia is warranted.
Asunto(s)
Pruebas de Coagulación Sanguínea/métodos , Carboxihemoglobina/metabolismo , Neoplasias del Colon/sangre , Hemo-Oxigenasa 1/metabolismo , Neoplasias Pancreáticas/sangre , Tromboelastografía/métodos , Adulto , Coagulación Sanguínea , Femenino , Humanos , Masculino , Persona de Mediana Edad , Adulto JovenRESUMEN
BACKGROUND: Patients with brain tumors suffer significant thrombotic morbidity and mortality. In addition to increased thrombin generation via tumor release of tissue factor-bearing microparticles and hyperfibrinogenemia, brain tumors and surrounding normal brain likely generate endogenous carbon monoxide (CO) via the hemeoxygenase-1 (HO-1) system. CO has been shown to enhance plasmatic coagulation via formation of carboxyhemefibrinogen (COHF). Thus, our goals in this study were to determine whether patients with brain tumors had increased HO-1 upregulation/CO production, plasmatic hypercoagulability, and formation of COHF. METHODS: Patients with brain tumors (N = 20) undergoing craniotomy had blood collected for determination of carboxyhemoglobin as a marker of HO-1 activity, plasmatic hypercoagulability (defined as clot strength > 95% confidence interval value of normal subject plasma), and COHF formation (determined with a thrombelastograph-based assay). Plasma obtained from commercially available normal subjects (N = 30) was used for comparison with brain tumor patient samples. RESULTS: Brain tumor patients had carboxyhemoglobin concentrations of 1.5% ± 0.5% (mean ± SD), indicative of HO-1 upregulation. Compared with normal subject plasma, brain tumor patient plasma had significantly (P < 0.0001) greater clot formation velocity (5.2 ± 1.5 vs 9.5 ± 2.3 dynes/cm/s, respectively) and significantly (P = 0.00016) stronger final clot strength (166 ± 28 vs 230 ± 78 dynes/cm, respectively). Ten of the brain tumor patients had plasma clot strength that exceeded the 95% confidence interval value observed in normal subjects, and 12 of the brain tumor patients had COHF formation. Five of the brain tumor patients in the hypercoagulable subgroup had COHF formation. Last, 5 of the hypercoagulable patients had primary brain tumors, whereas the other 5 patients had metastatic tumors or an inflammatory mass lesion. CONCLUSIONS: A subset of patients with brain tumors has increased endogenous CO production, plasmatic hypercoagulability, and COHF formation. Future investigation of the role played by HO-1 derived CO in the pathogenesis of brain tumor-associated thrombophilia is warranted.
Asunto(s)
Coagulación Sanguínea/fisiología , Neoplasias Encefálicas/sangre , Neoplasias Encefálicas/enzimología , Hemo-Oxigenasa 1/fisiología , Plasma/fisiología , Adulto , Anciano de 80 o más Años , Biopsia , Monóxido de Carbono/metabolismo , Carboxihemoglobina/análisis , Craneotomía , Femenino , Fibrinógeno/análisis , Fibrinolíticos/farmacología , Hemo-Oxigenasa 1/biosíntesis , Humanos , Cinética , Masculino , Persona de Mediana Edad , Tromboelastografía , Regulación hacia ArribaRESUMEN
OBJECTIVES: Lung cancer is an important health threat worldwide, and is associated with a 3.8-13.9% incidence of thrombophilia. Of interest, patients with lung tumors have been noted to have an increase in endogenous carbon monoxide production via upregulation of hemeoxygenase-1 activity. Given that it has been demonstrated that carbon monoxide enhances plasmatic coagulation in vitro and in vivo via formation of carboxyhemefibrinogen, we sought to determine if patients with thoracic tumors undergoing lung resection/pneumonectomy had an increase in endogenous carbon monoxide and concurrent plasmatic hypercoagulability. MATERIALS AND METHODS: Nonsmoking patients with thoracic tumors (n=19) had preoperative carboxyhemoglobin (a measure of carbon monoxide production) determined, and a thromboelastometric method to assess citrated plasma coagulation kinetics and the formation of carboxyhemefibrinogen was utilized. Thoracic tumor patient coagulation kinetics was compared with normal subject (n=30) plasma samples. RESULTS AND CONCLUSION: Patients with thoracic tumors were determined to have an abnormally increased carboxyhemoglobin concentration of 2.1±0.6%, indicative of hemeoxygenase-1 upregulation. It was found that 84% of thoracic tumor patients had plasma clot strength that exceeded the 95% confidence interval value observed in normal subjects, and 44% of this hypercoagulable subgroup had carboxyhemefibrinogen formation. Future investigation of the role played by plasmatic hypercoagulability and hemeoxygenase-1 derived carboxyhemefibrinogen in the pathogenesis of thoracic tumor related thrombophilia is warranted.
Asunto(s)
Hemo-Oxigenasa 1/metabolismo , Neoplasias Torácicas/enzimología , Neoplasias Torácicas/epidemiología , Trombofilia/enzimología , Trombofilia/epidemiología , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Coagulación Sanguínea , Monóxido de Carbono/metabolismo , Carboxihemoglobina/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Persona de Mediana Edad , Neoplasias Torácicas/sangre , Tromboelastografía , Trombofilia/sangre , Regulación hacia Arriba , Adulto JovenRESUMEN
Breast cancer is an important health threat to women worldwide, and is associated with a 9-14% incidence of thrombophilia. Of interest, patients with breast cancer have been noted to have an increase in endogenous carbon monoxide production via upregulation of heme oxygenase-1 activity. Given that it has been demonstrated that carbon monoxide enhances plasmatic coagulation in vitro and in vivo, we sought to determine whether patients with breast cancer had an increase in endogenous carbon monoxide and concurrent plasmatic hypercoagulability. Breast cancer patients who were not smokers scheduled to undergo partial or complete mastectomy (nâ=â18) had 15âml of whole blood collected via an indwelling intravenous catheter and anticoagulated with sodium citrate. Whole blood was centrifuged and citrated plasma assessed with a thromboelastometric method to measure coagulation kinetics and the formation of carboxyhemefibrinogen. Breast cancer patients were determined to have an abnormally increased carboxyhemoglobin concentration of 2.5â±â1.3%, indicative of heme oxygenase-1 upregulation. Breast cancer patient plasma on average clotted 73% more quickly and had 32% stronger thrombus strength than normal individual (nâ=â30) plasma. Further, 44% of breast cancer patients had plasma clot strength that exceeded the 95% confidence interval value observed in normal individuals, and 75% of this hypercoagulable subgroup had carboxyhemefibrinogen formation. Future investigation of the role played by heme oxygenase-1-derived carbon monoxide in the pathogenesis of breast cancer-related thrombophilia is warranted.