Acceleration of thrombin-antithrombin complex formation in rat hindquarters via heparinlike molecules bound to the endothelium.
We have examined the role of heparinlike molecules in the regulation of coagulation by perfusing rat hindquarters with purified human thrombin and with its plasma inhibitor, antithrombin. Our data indicate that contact of the hemostatic components with the endothelium enhances the rate of thrombin-antithrombin complex formation by as much as 19-fold over the uncatalyzed rate of enzyme-inhibitor interaction. Heparinlike molecules are responsible for the antithrombin accelerating activity. The amount of thrombin-antithrombin complex generated within the hindlimb preparation after pretreatment of the vasculature with purified Flavobacterium heparinase or with addition of platelet Factor IV to the hemostatic components, was equal to the uncatalyzed levels. These heparinlike molecules appear to be tightly bound to the luminal surface of the endothelium, since they could not be detected within the physiologic buffer that was perfused through the animal. The above mucopolysaccharides function in a manner similar to commercial heparin, since modification of antithrombin at a site critical for heparin-dependent acceleration of the protease inhibitor resulted in a level of interaction product identical to the uncatalyzed amount. Finally, addition of diisofluorophosphate-thrombin to the enzyme perfusion stream reduced the amount of thrombin-antithrombin complex formed in the animal by 30-40%, which suggested that thrombin bound to the endothelium as well as enzyme free in solution are accessible to antithrombin that has interacted with heparinlike molecules present on the endothelium.
D000818:Animals; D000991:Antithrombins / Q000502:physiology*; D002196:Capillaries / Q000502:physiology; D004727:Endothelium / Q000502:physiology; D005417:Flavobacterium / Q000201:enzymology; D006493:Heparin / Q000502:physiology*; D019763:Heparin Lyase; D006801:Humans; D007700:Kinetics; D008297:Male; D009132:Muscles / Q000098:blood supply*; D011133:Polysaccharide-Lyases; D011485:Protein Binding; D051381:Rats; D011919:Rats, Inbred Strains; D013917:Thrombin / Q000502:physiology*
D000991:Antithrombins; D006493:Heparin; D013917:Thrombin; D011133:Polysaccharide-Lyases; D019763:Heparin Lyase