INCREASED ANTICOAGULANT ACTIVITY OF RECOMBINANT THROMBOMODULIN MODIFIED WITH GLYCOSAMINOGLYCAN

Thrombomodulin (TRI) is a thrombin receptor on the endothelial cell su rface, effective as an anticoagulant by changing procoagulant thrombin to an anticoagulant one. As rabbit TM with glycosaminoglycan (GAG) ha s a more potent anticoagulant activity than that without GAG, we expre ssed recombinant GAG-modified urinary thrombomodulin (GAG-UTM) in C-12 7 cells. The effect of an additional GAG chain on anticoagulant activi ty was investigated in comparison with unmodified recombinant UTM (r-U TM). In vitro, the activity of cleavage of fibrinogen by thrombin or p rothrombinase activity was more potently depressed by GAG-UTM than by r-UTM, and the generation of activated protein C by TM-thrombin comple x was accelerated by GAG modification. The acceleration of antithrombi n III-dependent anticoagulant activity was shown only by GAG-UTM. Para meters like thrombin time, prothrombin time and activated partial thro mboplastin time in human plasma were prolonged by GAG-UTM more than by r-UTM. In vivo, the effect of GAG-UTM and r-UTM in endotoxin-induced disseminated intravascular coagulation (DIC) rats was investigated usi ng hematological parameters. GAG-UTM and r-UTM significantly reduced t he decrease in fibrinogen and platelet number induced by endotoxin at the dosage of 0.1 and 1.0 mg/kg/h, respectively, suggesting that the a ntithrombotic effect of GAG-UTM in endotoxin-induced DIC rats was 10-f old as potent as that of r-UTM, GAG-UTM reduced the prolongation of th e bleeding time induced by endotoxin, while r-UTM accelerated it. Thes e results suggest that the addition of a GAG chain may increase availa bility as an anticoagulant.