Antiangiogenic antithrombin blocks the heparan sulfate-dependent binding of proangiogenic growth factors to their endothelial cell receptors: evidence for differential binding of antiangiogenic and anticoagulant forms of antithrombin to proangiogenic heparan sulfate domains.
The anticoagulant serpin antithrombin acquires a potent antiangiogenic activity upon undergoing conformational alterations to cleaved or latent forms. Here we show that antithrombin antiangiogenic activity is mediated at least in part through the ability of the conformationally altered serpin to block the proangiogenic growth factors fibroblast growth factor (FGF)-2 and vascular endothelial growth factor (VEGF) from forming signaling competent ternary complexes with their protein receptors and heparan sulfate co-receptors on endothelial cells. Cleaved and latent but not native forms of antithrombin blocked the formation of FGF-2-FGF receptor-1 ectodomain-heparin ternary complexes, and the dimerization of these complexes in solution and similarly inhibited the formation of FGF-2-heparin binary complexes and their dimerization. Only antiangiogenic forms of antithrombin likewise inhibited (125)I-FGF-2 binding to its low affinity heparan sulfate co-receptor and blocked FGF receptor-1 autophosphorylation and p42/44 MAP kinase phosphorylation in cultured human umbilical vein endothelial cells (HUVECs). Moreover, treatment of HUVECs with heparinase III to specifically eliminate the FGF-2 heparan sulfate co-receptor suppressed the ability of antiangiogenic antithrombin to inhibit growth factor-stimulated proliferation. Antiangiogenic antithrombin inhibited full-length VEGF(165) stimulation of HUVEC proliferation but did not affect the stimulation of cells by the heparin-binding domain-deleted VEGF(121). Taken together, these results demonstrate that antiangiogenic forms of antithrombin block the proangiogenic effects of FGF-2 and VEGF on endothelial cells by competing with the growth factors for binding the heparan sulfate co-receptor, which mediates growth factor-receptor interactions. Moreover, the inability of native antithrombin to bind this co-receptor implies that native and conformationally altered forms of antithrombin differentially bind proangiogenic heparan sulfate domains.
Zhang
Weiqing
W
Center for Molecular Biology of Oral Diseases, University of Illinois at Chicago, Chicago, Illinois 60612, USA. zhang98@uic.edu
Swanson
Richard
R
Xiong
Yan
Y
Richard
Benjamin
B
Olson
Steven T
ST
D020533:Angiogenesis Inhibitors / Q000378:metabolism / Q000494:pharmacology*; D000925:Anticoagulants / Q000378:metabolism / Q000494:pharmacology; D000991:Antithrombins / Q000378:metabolism / Q000494:pharmacology*; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D005346:Fibroblast Growth Factors / Q000378:metabolism*; D006493:Heparin / Q000378:metabolism; D019763:Heparin Lyase / Q000494:pharmacology; D006497:Heparan Sulfate / Q000378:metabolism*; D006801:Humans; D011485:Protein Binding; D051496:Receptor, Fibroblast Growth Factor, Type 1 / Q000378:metabolism; D040262:Receptors, Vascular Endothelial Growth Factor / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism*
D020533:Angiogenesis Inhibitors; D000925:Anticoagulants; D000991:Antithrombins; D005346:Fibroblast Growth Factors; D006493:Heparin; D019763:Heparin Lyase; D006497:Heparan Sulfate; D051496:Receptor, Fibroblast Growth Factor, Type 1; D040262:Receptors, Vascular Endothelial Growth Factor; D042461:Vascular Endothelial Growth Factor A; C416003:FGF20 protein, human; C467484:VEGFA protein, human; C496348:FGFR1 protein, human