In vivo clearance of ternary complexes of vitronectin-thrombin-antithrombin is mediated by hepatic heparan sulfate proteoglycans.
Thrombin is inhibited by its cognate plasma inhibitor antithrombin, through the formation of covalent thrombin-antithrombin (TAT) complexes that are found as ternary complexes with vitronectin (VN-TAT). To determine whether the metabolism of VN-TAT ternary complexes is different from that previously reported for binary TAT complexes, plasma clearance studies were done in rabbits using human VN-TAT. 125I-VN-TAT was shown to be cleared rapidly from the circulation (t1/2alpha = 3.8 min) in a biphasic manner mainly by the liver. 125I-TAT had a similar initial clearance (t1/2alpha = 5.3 min) but had a significantly faster beta-phase clearance (t1/2beta = 42.8 min versus 85.4 min for VN-TAT; p = 0.005). Protamine sulfate and heparin abolished the rapid initial alpha-phase of 125I-VN-TAT clearance and reduced its liver-specific association and in vivo degradation. Heparin also reduced the alpha-phase clearance of 125I-TAT and was associated with the appearance of high molecular weight complexes, suggesting enhanced complex formation between VN and TAT. 125I-VN-TAT binding to HepG2 cells was reduced by competition with VN-TAT or heparin but to a much lesser extent in the presence of TAT. The binding of VN-TAT to HepG2 cells was not inhibited by competition with the low density lipoprotein receptor-related protein ligand, methylamine-alpha2-macroglobulin. 125I-VN-TAT binding was also inhibited by treating HepG2 cells with heparinase or by growing the cells in the presence of beta-D-xyloside. Finally, both heparin and chloroquine, but not methylamine-alpha2-macroglobulin, reduced the internalization and degradation of VN-TAT by HepG2 cells. Taken together, these data indicate the importance of VN in TAT metabolism and demonstrate that VN-TAT binds to liver-associated heparan sulfate proteoglycans, which mediate its internalization and subsequent intracellular degradation.
Department of Pathology, McMaster University, Hamilton, Ontario L8N 3Z5, Canada.
M J
MJ
Wells
M A
MA
Blajchman
D000818:Animals; D000990:Antithrombin III / Q000493:pharmacokinetics*; D001692:Biological Transport; D006528:Carcinoma, Hepatocellular / Q000378:metabolism; D019812:Heparan Sulfate Proteoglycans / Q000378:metabolism*; D006801:Humans; D008099:Liver / Q000378:metabolism*; D008113:Liver Neoplasms / Q000378:metabolism; D008657:Metabolic Clearance Rate; D011485:Protein Binding; D011817:Rabbits; D011869:Radioligand Assay; D013917:Thrombin / Q000493:pharmacokinetics*; D014018:Tissue Distribution; D019096:Vitronectin / Q000493:pharmacokinetics*
D019812:Heparan Sulfate Proteoglycans; D019096:Vitronectin; D000990:Antithrombin III; D013917:Thrombin