Hirudin interruption of heparin-resistant arterial thrombus formation in baboons

Abstract

To determine the role of thrombin in high blood flow, platelet- dependent thrombotic and hemostatic processes we measured the relative antithrombotic and antihemostatic effects in baboons of hirudin, a highly potent and specific antithrombin, and compared the effects of heparin, an antithrombin III-dependent inhibitor of thrombin. Thrombus formation was determined in vivo using three relevant models (homologous endarterectomized aorta, collagen-coated tubing, and Dacron vascular graft) by measuring: (1) platelet deposition, using gamma camera imaging of 111In-platelets; (2) fibrin deposition, as assessed by the incorporation of circulating 125I-fibrinogen; and (3) occlusion. The continuous intravenous infusion of 1, 5, and 20 nmol/kg per minute of recombinant hirudin (desulfatohirudin) maintained constant plasma levels of 0.16 +/- 0.03, 0.79 +/- 0.44, and 3.3 +/- 0.77 mumol/mL, respectively. Hirudin interrupted platelet and fibrin deposition in a dose-dependent manner that was profound at the highest dose for all three thrombogenic surfaces and significant at the lowest dose for thrombus formation on endarterectomized aorta. Thrombotic occlusion was prevented by all doses studied. In contrast, heparin did not inhibit either platelet or fibrin deposition when administered at a dose that maximally prolonged clotting times (100 U/kg) (P greater than .1), and only intermediate effects were produced at 10-fold that dose (1,000 U/kg). Moreover, heparin did not prevent occlusion of the test segments. Hirudin inhibited platelet hemostatic function in concert with its antithrombotic effects (bleeding times were prolonged by the intermediate and higher doses). By comparison, intravenous heparin failed to affect the bleeding time at the 100 U/kg dose (P greater than .5), and only minimally prolonged the bleeding time at the 1,000 U/kg dose (P less than .05). We conclude that platelet-dependent thrombotic and hemostatic processes are thrombin-mediated and that the biologic antithrombin hirudin produces a potent, dose-dependent inhibition of arterial thrombus formation that greatly exceeds the minimal antithrombotic effects produced by heparin.

Fibrinogen γ′ chain carboxy terminal peptide selectively inhibits the intrinsic coagulation pathway

“…In experiments with infused γ′ 410–427 peptide, thrombosis was initiated using a Dacron graft segment, which accumulates a platelet‐rich, arterial‐type thrombus, that was proximal to a chamber region of low‐shear flow expansion, which accumulates fibrin‐rich, venous‐type thrombus (Cadroy et al , 1989; Hanson et al , 1993). This model was chosen because the thrombus that forms on the Dacron graft segment is sensitively inhibited by antiplatelet agents, but much less by anticoagulants, such as heparin (Kelly et al , 1991), while the thrombus that forms within the disturbed flow region is sensitively inhibited by heparin and other agents that interfere with blood coagulation (Cadroy et al , 1989; Hanson et al , 1993). The data from two animals treated with the γ′ peptide were averaged and compared with eight control animals that received no peptide.…”

A four-hour infusion of recombinant hirudin results in long-term inhibition of arterial-type thrombosis in baboons

“…It also confirms the findings that r-hirudin interrupts platelet-rich, arterial-type thrombus formation induced by Dacron vascular graft material, collagen coated tubing, endarterectomized aorta segments, and following angioplasty. 6,26 When treatment was stopped, platelets were again deposited (Fig 1). However, the rate of deposition was much slower and markedly less platelets were deposited than in ment with r-hirudin reduced the maximum size of the thrombus by approximately 59% and that maximum deposition was attained at a much later stage.…”

Reduction in Vascular Lesion Formation by Hirudin Secreted From Retrovirus-Transduced Confluent Endothelial Cells on Vascular Grafts in Baboons

“…23 Baboon fibrinogen was purified by ␤-alanine precipitation and labeled with 125 I by the Iodine monochloride method as described previously. 10…”

“…Because systemic concentrations of hirudin that prevent TR-dependent platelet recruitment and fibrin generation concurrently impair hemostatic function, 10,31 we adopted the safety strategy of generating inhibitory concentrations of hirudin locally using AVG-attached hirudin-transduced ECs. These safety concerns regarding systemic hirudin therapy have been heightened by the recent controlled clinical trials evaluating the effects of systemic hirudin.…”