Exp Mol Med.  2003 Dec;35(6):578-585.

Differential inhibition of endothelial cell proliferation and migration by urokinase subdomains: amino-terminal fragment and kringle domain

• Kim KS
• Hong YK
• Lee Y
• Shin JY
• Chang SI
• Chung SI
• Joe YA

Abstract

The serine protease urokinase-type plasminogen activator (uPA) is implicated in pericellular proteolysis in a variety of physiological and pathological processes including angiogenesis and tumor metastasis. The kringle domain of uPA (UK1) has proven to be an anti-angiogenic molecule with unknown mechanism and amino terminal fragment of uPA (u-ATF) with additional growth factor-like domain can be used for blocking interaction of uPA and uPA receptor. Here, we compared anti-angiogenic activities of these two molecules in vitro and in vivo. The recombinant u-ATF from E. coli and refolded in vitro was found to bind to uPAR with high affinity, whereas E. coli-derived UK1 showed no binding by Biacore analysis. In contrast to UK1 having potent inhibitory effect, u-ATF exhibited low inhibitory effect on bovine capillary endothelial cell growth (ED(50)>320 nM). Furthermore, u-ATF inhibition of VEGF-induced migration of human umbilical vein endothelial cell was far less sensitive (IC(50)= 600 nM) than those observed with UK1, and angiogenesis inhibition was marginal in chorioallantoic membrane. These results suggest that kringle domain alone is sufficient for potent anti- angiogenic activity and additional growth factor-like domain diverts this molecule in undergoing different mechanism such as inhibition of uPA/uPAR interaction rather than undergoing distinct anti- angiogenic mechanism driven by kringle domain.

Keyword

angiogenesis inhibitors; angiostatic proteins; cell migration inhibition; endothelial cells; urokinase-type plasminogen activator

MeSH Terms

Animals
Biosensing Techniques
Cattle
Cell Division/drug effects
Cell Movement/*drug effects
Cells, Cultured
Chickens
Cricetinae
Endothelial Cells/*cytology/*drug effects
Humans
Kinetics
*Kringles
Ligands
Peptide Fragments/*chemistry/genetics/metabolism/*pharmacology
Protein Binding
Receptors, Cell Surface/metabolism
Receptors, Urokinase Plasminogen Activator
Urokinase-Type Plasminogen Activator/*chemistry/genetics/pharmacology
Vascular Endothelial Growth Factor A/pharmacology