Thrombin Inhibition in an Ex Vivo Model of Porcine Heart Xenograft Hyperacute Rejection1

Abstract

Prominent components of vascularized xenograft rejection such as platelet activation and microvascular thrombosis may be dependent upon thrombin generation in vivo. To study potential therapeutic benefits of a synthetic low-molecular-weight thrombin inhibitor, SDZ MTH 958, in hyperacute porcine heart rejection by human blood ex vivo, a working model of hyperacute rejection of porcine by fresh, heparinized (6 microM/ml) human blood with or without 1 microM SDZ MTH 958 was used. Thrombin-antithrombin complexes (TAT) and prothrombin fragment F1.2 levels as markers of thrombin activation were determined, and biopsies from rejected hearts were analyzed by immunohistopathology. Control porcine hearts (n=8) underwent a rapid and consistent decline in cardiac output, ceasing function by 60 min. Experimental cardiac output values of 14 ml/g (SEM 1.2) were significantly higher than seen in controls (5 ml/g SEM 0.6) after 5 min of cardiac work, and prolonged survival times up to 120 min were noted (P<0.05). Activity of SDZ MTH 958 was confirmed by functional assays throughout perfusion. Levels of TAT and F1.2 increased consistently in control samples when compared with plasma samples containing SDZ MTH 958. Immunohistopathological examination confirmed diminished fibrin deposition, reduced leukocyte adherence to endothelium, impaired diapedesis and less tissue necrosis in the hearts perfused with SDZ MTH 958. SDZ MTH 958, in this xenoperfusion model, prolonged survival, enhanced function of the explanted organ, and improved histological features at the time of rejection. Effective and specific antagonism of thrombin may be useful as an adjunct therapy to complement inhibition for xenograft rejection

Hyperacute lung rejection in the pig-to-human model. III. platelet receptor inhibitors synergistically modulate complement activation and lung injury

“…Removal of platelets and neutrophils by Pall filtration dramatically prolongs survival of pig hearts, whereas adding back platelets to pall-filtered blood shortens survival. Although GPIIbIIIa blockade alone is not effective in this model (S. Robson, unpublished data, 1994), thrombin inhibition is moderately protective (26), as it is for the lung (27). We have also previously shown that aspirin-sensitive mechanisms, thrombin activation, and complement activa-FIGURE 2.…”

α1,3-Galactosyltransferase Gene-Knockout Pigs for Xenotransplantation: Where Do We Go From Here?

“…(1) Porcine TFPI (pTFPI) does not completely neutralize human factor Xa (66). (2) Injured porcine EC activate human factor Xa and thrombin (67). (3) Porcine thrombomodulin does not act as an effective cofactor for the activation of human protein C by human thrombin (68).…”

Xenotransplantation as a model of integrated, multidisciplinary research

“…62 Porcine tissue factor pathway inhibitor (TFPI) is not able to neutralize human factor Xa, and is therefore unable to inhibit the direct activation of human prothrombin to thrombin. 63,64 In addition, although porcine thrombomodulin has been shown to bind human thrombin and Protein C, the human thrombin-porcine thrombomodulin complex is a poor activator of Protein C. As a consequence, the insufficient production of activated Protein C contributes to enhanced levels of thrombin, favouring the initiation of clotting. 64,65 In order to combat such incompatibilities, transgenic modulation of the clotting cascade by de novo expression or induction of anticoagulants, or the elimination of pro-coagulant molecules on the xenogenic vascular endothelium, may represent a potential therapeutic strategy.…”