- Email: [email protected]
Altered regulation of the coagulation system due to endotoxin tolerance in a porcine shock model
S40
ABSTRACTS OF 12TH INTNAT’L CONGRESS
Vol. 65, Suppi. 1
c75 FPA GENERATION IN A THROMBOPLASTIN - INDUCED RAT TROMBOSJS MODEL R. Zienz, M. Herrmann, M. Fredrich, B. Baldus Research Laboratories of Schering AG, Berlin, D Fibrinogen is one of the thrombin substrates. A specific proteolysis mediated by thrombin leads to the liberation of the amino-terminal fibrinopeptides A and B and initiates fibrin polymerization. An ELISA technique for the quantitation of rat tibrinopeptide A (RtFPA) has been established. Synthetic rat FPA (sRtFPA) representing the 17 amino acids of native RtFPA was conjugated to keyhole limpet hemocyanin and was used for immunization to prepare rabbit IgG anti- RtFPA. The assay was based on the neutralization of the 1gG binding activity towards immobilized sRtFPA by free RtFPA in the samples. Interfering fibrinogen was removed by bentonite adsorption. The assay was used to investigate the kinetics of FPA generation in an experimental thrombosis model. Thrombus formation in wistar rats was induced by a transient infusion (5 minutes) of a thromboplastin reagent. During the period of infusion the vena cava was ligated. Thrombus mass formed in the ligated vessel was determined after 60 minutes of stasis (59 & 17 mg; mean 2 SE, n-6). FPA plasma levels reached maximal values (80 2 19 rig/ml) after IO minutes of ligation and almost returned to initial values (20 + 4 rig/ml) after 60 minutes. When thromboplastin was omitted in the stasis model, FPA plasma levels remained at basal levels and thrombus formation was absent. In heparinixed animals (200 IV/kg i.v.) thrombi did not develop despite thromboplastin administration. In addition, heparin inhibited the endogenous thrombin activity as could be detected by a 40% reduction of FPA plasma levels 60 minutes after heparinixation. The assay described will be useful for the determination of a prethrombotic state and for the quantitation of thromboembolic events in rat models, as the presence of free FPA in plasma is a marker for thrombin activity.
C76 ALTERED REGULATION OF THE COAGULATION SYSTEM DUE TO ENDOTOXIN TOLERANCE IN A PORCINE SHOCK MODEL B. Horst, R Kujath, K.H. Staubach, A. Kooistra, S. Jonas Dept. of Surgery, Medical University of Liibeck, D In an animal model the influence of endotoxin tolerance in septic shock and related alterations of the coagulation system were investigated. Endotoxin from salmonella abortis equi H 1178 was administered i.v. to mixed-breed pigs (IO ng on day 1.20 ng on day 2 and 40 ng on day 3 and 4) inducing an immunological response to endotoxin. After a week, the animals were narcotized and mechanically ventilated after tracheostomy. An endotoxin shock was initiated by continuous infusion of 250 @kg/h et&toxin. Cardiovascular monitoring was done by measuring mean arterial pressure, pulmonary capillary wedge pressure and extravascular lung water by intra-arterial and intravenous catheters. The following laboratory parameters were measured: haemoglobin, leucocytes, thrombocytes, fibrinogen, haemoglobin, &tat, GOT, GPT, aZmacroglobulin, al-antiplasmin, TAT, t-PA, D-DIMER, al-antitrypsin, fibrinmonomers, plasminogen. Endotoxin caused an early-phase tolerance and lead to a significantly increased survival time after induction of an ET-shock. Compared to animals without previous endotoxin application, the progress of the septic shock was prolonged and haemodynamic decompensation occurred about 6 hours later. Parallel to haemodynamic decompensation with low cardiac output and peripheral vasodilatation TAT, D-DIMER and fibrinmonomers increased. whereas leucocytes, thrombocytes and plasminogen decreased. Compared to animals without endotoxin tolerance, the fibrinolysis regulating systems (t-PA and aZantiplasmin) decompensated later in the group of animals with endotoxin tolerance. Prophylaxis of toxic shock in surgical patients at risk may become possible it less toxic immunomodulaters can be developed.
ABSTRACTS OF 12TH INTNAT’L CONGRESS
Vol. 65, Suppi. 1
c75 FPA GENERATION IN A THROMBOPLASTIN - INDUCED RAT TROMBOSJS MODEL R. Zienz, M. Herrmann, M. Fredrich, B. Baldus Research Laboratories of Schering AG, Berlin, D Fibrinogen is one of the thrombin substrates. A specific proteolysis mediated by thrombin leads to the liberation of the amino-terminal fibrinopeptides A and B and initiates fibrin polymerization. An ELISA technique for the quantitation of rat tibrinopeptide A (RtFPA) has been established. Synthetic rat FPA (sRtFPA) representing the 17 amino acids of native RtFPA was conjugated to keyhole limpet hemocyanin and was used for immunization to prepare rabbit IgG anti- RtFPA. The assay was based on the neutralization of the 1gG binding activity towards immobilized sRtFPA by free RtFPA in the samples. Interfering fibrinogen was removed by bentonite adsorption. The assay was used to investigate the kinetics of FPA generation in an experimental thrombosis model. Thrombus formation in wistar rats was induced by a transient infusion (5 minutes) of a thromboplastin reagent. During the period of infusion the vena cava was ligated. Thrombus mass formed in the ligated vessel was determined after 60 minutes of stasis (59 & 17 mg; mean 2 SE, n-6). FPA plasma levels reached maximal values (80 2 19 rig/ml) after IO minutes of ligation and almost returned to initial values (20 + 4 rig/ml) after 60 minutes. When thromboplastin was omitted in the stasis model, FPA plasma levels remained at basal levels and thrombus formation was absent. In heparinixed animals (200 IV/kg i.v.) thrombi did not develop despite thromboplastin administration. In addition, heparin inhibited the endogenous thrombin activity as could be detected by a 40% reduction of FPA plasma levels 60 minutes after heparinixation. The assay described will be useful for the determination of a prethrombotic state and for the quantitation of thromboembolic events in rat models, as the presence of free FPA in plasma is a marker for thrombin activity.
C76 ALTERED REGULATION OF THE COAGULATION SYSTEM DUE TO ENDOTOXIN TOLERANCE IN A PORCINE SHOCK MODEL B. Horst, R Kujath, K.H. Staubach, A. Kooistra, S. Jonas Dept. of Surgery, Medical University of Liibeck, D In an animal model the influence of endotoxin tolerance in septic shock and related alterations of the coagulation system were investigated. Endotoxin from salmonella abortis equi H 1178 was administered i.v. to mixed-breed pigs (IO ng on day 1.20 ng on day 2 and 40 ng on day 3 and 4) inducing an immunological response to endotoxin. After a week, the animals were narcotized and mechanically ventilated after tracheostomy. An endotoxin shock was initiated by continuous infusion of 250 @kg/h et&toxin. Cardiovascular monitoring was done by measuring mean arterial pressure, pulmonary capillary wedge pressure and extravascular lung water by intra-arterial and intravenous catheters. The following laboratory parameters were measured: haemoglobin, leucocytes, thrombocytes, fibrinogen, haemoglobin, &tat, GOT, GPT, aZmacroglobulin, al-antiplasmin, TAT, t-PA, D-DIMER, al-antitrypsin, fibrinmonomers, plasminogen. Endotoxin caused an early-phase tolerance and lead to a significantly increased survival time after induction of an ET-shock. Compared to animals without previous endotoxin application, the progress of the septic shock was prolonged and haemodynamic decompensation occurred about 6 hours later. Parallel to haemodynamic decompensation with low cardiac output and peripheral vasodilatation TAT, D-DIMER and fibrinmonomers increased. whereas leucocytes, thrombocytes and plasminogen decreased. Compared to animals without endotoxin tolerance, the fibrinolysis regulating systems (t-PA and aZantiplasmin) decompensated later in the group of animals with endotoxin tolerance. Prophylaxis of toxic shock in surgical patients at risk may become possible it less toxic immunomodulaters can be developed.