Endogenous nitric oxide acts as a natural antithrombotic agent in vivo by inhibiting platelet aggregation in the pulmonary vasculature.

Nitric oxide (NO) is a powerful vasodilator and an inhibitor of platelet aggregation in vitro. While the ability of NO to modulate vascular tone in vivo has been proven, only a few studies have assessed its platelet inhibitory activity in vivo. We have employed two complementary animal models of pulmonary platelet thromboembolism to assess the antithrombotic activity of endogenous NO in vivo. The inhibition of nitric oxide synthase (NOS) by L-NAME significantly potentiated while the administration of the NOS substrate L-arginine significantly reduced the accumulation of 111In-labelled platelets in the pulmonary vasculature of rabbits induced by intravenous collagen plus epinephrine. L-NAME or L-arginine did not, however, modify 111In-labelled erythrocyte distribution in lungs and phenylephrine had no effect on platelet accumulation following collagen + adrenaline, suggesting that the effects of L-NAME were not due to vasoconstriction but rather to a direct modification of platelet function. In mice, L-NAME significantly reduced the dose of collagen + adrenaline required to induce thromboembolic mortality, increased the fall in circulating platelets and increased the % of pulmonary vessels occluded by platelet thrombi. The effects of L-NAME were reversed by L-arginine but not by a dose of nicardipine exerting maximal vasodilatation. Phenylephrine did not potentiate collagen + adrenaline-induced mortality. In the pulmonary vasculature in vivo, endogenous NO inhibits collagen + adrenaline-induced aggregation and enhances platelet disaggregation. This natural modulator function of NO is exerted via a direct effect on platelets and not as a result of haemodynamic changes.

Activated human protein C prevents thrombin-induced thromboembolism in mice. Evidence that activated protein c reduces intravascular fibrin accumulation through the inhibition of additional thrombin generation.

Activated protein C (APC) is a potent physiologic anticoagulant with profibrinolytic properties, and has been shown to prevent thrombosis in different experimental models. We investigated the effect of human APC on thrombin-induced thromboembolism in mice, a model of acute intravascular fibrin deposition leading to death within minutes. APC given intravenously (i.v.) as a bolus 2 min before thrombin challenge (1,250 U/kg) reduced mortality in a dose-dependent manner despite the lack of thrombin inhibitor activity. Significant inhibition of thrombin-induced death was observed at the dose of 0.05 mg/kg, and maximal protection was obtained with 2 mg/kg (> 85% reduction in mortality rate). Histology of lung tissue revealed that APC treatment (2 mg/kg) reduced significantly vascular occlusion rate (from 89.2 to 46.6%, P < 0.01). The protective effect of APC was due to the inhibition of endogenous thrombin formation as indicated by the fact that (a) the injection of human thrombin caused a marked decrease in the coagulation factors of the intrinsic and common pathways (but not of Factor VII), suggesting the activation of blood clotting via the contact system; (b) APC pretreatment reduced markedly prothrombin consumption; (c) the lethal effect of thrombin was almost abolished when the animals were made deficient in vitamin K-dependent factors by warfarin treatment, and could be restored only by doubling the dose of thrombin, indicating that the generation of endogenous thrombin contributes significantly to death; and (d) APC failed to protect warfarin-treated animals, in which mortality is entirely due to injected thrombin, even after protein S supplementation. Other results suggest that APC protects from thrombin-induced thromboembolism by rendering the formed fibrin more susceptible to plasmin degradation rather than by reducing fibrin formation: in thrombin-treated mice, fibrinogen consumption was not inhibited by APC; and inhibition of endogenous fibrinolysis by epsilon-aminocaproic or tranexamic acid resulted in a significant reduction of the protective effect of APC. Since APC did not enhance plasma fibrinolytic activity, as assessed by the measurement of plasminogen activator (PA) or PA inhibitor (PAI) activities, PAI-1 antigen, or 125I-fibrin degrading activity, we speculate that the inhibition of additional (endogenous) thrombin formation by APC interrupts thrombin-dependent mechanisms that make fibrin clots more resistant to lysis, so that the intravascular deposited fibrin can be removed more rapidly by the endogenous fibrinolytic system.