The Glycosaminoglycan of Recombinant Human Soluble Thrombomodulin Affects Antithrombotic Activity in a Rat Model of Tissue Factor-Induced Disseminated Intravascular Coagulation

Abstract

SummaryPrevious studies on recombinant human soluble thrombomodulin (rsTM) from Chinese hamster ovary cells revealed that rsTM was expressed as two proteins that differed functionally in vitro due to the presence (rsTMp) or absence (rsTMa) of chondroitin-4-sulfate. The current study evaluates the in vivo behavior of rsTM in rats and in a rat model of tissue factor-induced disseminated intravascular coagulation (DIC). rsTMp was more potent than rsTMa for prolongation of the activated partial thromboplastin time (APTT) and their in vivo half-lives determined by ELISA were 20 min for rsTMp and 5.0 h for rsTMa. Injection of a tissue factor suspension (5 mg/kg) resulted in DIC as judged by decreased platelet counts and fibrinogen concentrations, prolonged APTT, and increased fibrin and fibrinogen degradation products (FDP) levels. A bolus injection of either rsTM (0.2 mg/kg) 1 min before induction of DIC essentially neutralized effects on platelets, fibrinogen, and FDP levels, and had only a moderate effect on APTT prolongation. The dose of anticoagulant to inhibit the drop in platelet counts by 50% (ED50) was 0.2 mg/kg rsTMa, 0.07 mg/kg rsTMp, and 7 U/ kg heparin. The effect of increasing concentrations of rsTM and heparin on bleeding times were compared in experiments involving incision of the rat tail. Doubling of the bleeding times occurred at 5 mg/kg rsTMa, 3 mg/kg rsTMp or 90 U/kg heparin. These values represent a 25-fold increase over the ED50 for rsTMa, 43-fold for rsTMp and 13-fold for heparin. These results suggest that rsTMp is a potent anticoagulant to inhibit the platelet reduction when injected prior to the induction of DIC in rats.

Antiinflammatory and Anticoagulant Effects of Transgenic Expression of Human Thrombomodulin in Mice

“…The impact of thrombomodulin on the tail bleeding assay is in keeping with the results of several groups (40,41). Aoki demonstrated a prolongation of bleeding times using rhs-TBM, although the doses required were in excess of that needed to prevent tissue factor-induced disseminated intravascular coagulation in rats (41).…”

ART‐123: Recombinant Human Soluble Thrombomodulin

“…The half-lives of ART-123 are relatively long compared with those of other thrombomodulin derivatives described previously. Nawa et al (38) demonstrated that recombinant human soluble thrombomodulin with glycosaminoglycan had a shorter half-life than that treated with chondroitinase AC I. The half-lives of thrombomodulin composed of the last three epidermal growth factor-like structures (E456) are very short in monkeys at 9 (t 1/2a ) and 50 min (t 1/2b ) (48).…”

Local Overexpression of Thrombomodulin for In Vivo Prevention of Arterial Thrombosis in a Rabbit Model

“…19 Further, systemic infusion of recombinant soluble THM has shown promise as a useful anticoagulant therapy. 20 Endothelial cells reflect the local pathophysiological milieu and play active roles in determining, among other processes, the procoagulant/anticoagulant balance. 21 A perturbed endothelial cell dramatically decreases expression of THM as a critical mechanism for shifting the local environment toward a procoagulant state.…”