The antiangiogenic activity of cleaved high molecular weight kininogen is mediated through binding to endothelial cell tropomyosin.
Conformationally altered proteins and protein fragments derived from the extracellular matrix and hemostatic system may function as naturally occurring angiogenesis inhibitors. One example of such a protein is cleaved high molecular weight kininogen (HKa). HKa inhibits angiogenesis by inducing apoptosis of proliferating endothelial cells, effects mediated largely by HKa domain 5. However, the mechanisms underlying the antiangiogenic activity of HKa have not been characterized, and its binding site on proliferating endothelial cells has not been defined. Here, we report that the induction of endothelial cell apoptosis by HKa, as well as the antiangiogenic activity of HKa in the chick chorioallantoic membrane, was inhibited completely by antitropomyosin monoclonal antibody TM-311. TM-311 also blocked the high-affinity Zn2+-dependent binding of HKa to both purified tropomyosin and proliferating endothelial cells. Confocal microscopic analysis of endothelial cells stained with monoclonal antibody TM-311, as well as biotin labeling of cell surface proteins on intact endothelial cells, revealed that tropomyosin exposure was enhanced on the surface of proliferating cells. These studies demonstrate that the antiangiogenic effects of HKa depend on high-affinity binding to endothelial cell tropomyosin.
Division of Hematology-Oncology, Case Western Reserve University School of Medicine and University Hospitals of Cleveland, Cleveland, OH 44106, USA.
Jing-Chuan
JC
Zhang
Fernando
F
Donate
Xiaoping
X
Qi
Nicholas P
NP
Ziats
Jose C
JC
Juarez
Andrew P
AP
Mazar
Yuan-Ping
YP
Pang
Keith R
KR
McCrae
D000482:Allantois / Q000098:blood supply / Q000187:drug effects; D020533:Angiogenesis Inhibitors / Q000378:metabolism* / Q000494:pharmacology*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D001483:Base Sequence; D002478:Cells, Cultured; D002642:Chick Embryo; D002823:Chorion / Q000098:blood supply / Q000187:drug effects; D018076:DNA, Complementary / Q000235:genetics; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D006801:Humans; D019679:Kininogen, High-Molecular-Weight / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology*; D018919:Neovascularization, Physiologic / Q000187:drug effects; D011485:Protein Binding; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D014335:Tropomyosin / Q000037:antagonists & inhibitors / Q000378:metabolism*
D020533:Angiogenesis Inhibitors; D000911:Antibodies, Monoclonal; D018076:DNA, Complementary; D019679:Kininogen, High-Molecular-Weight; D011994:Recombinant Proteins; D014335:Tropomyosin