Role and localization of urokinase receptor in the formation of new microvascular structures in fibrin matrices.
Fibrin or a fibrinous exudate can facilitate angiogenesis in many pathological conditions. In vitro, the outgrowth of capillary-like structures in fibrin can be mimicked by exposing human microvascular endothelial cells (hMVECs) to an angiogenic growth factor and tumor necrosis factor (TNF)-alpha. Urokinase-type plasminogen activator (u-PA) and plasmin activities are required for this angiogenic process. This study focuses on the role and localization of the u-PA receptor (u-PAR) in newly formed microvascular structures. The u-PAR-blocking monoclonal antibody (MAb) H-2 completely inhibited the formation of capillary-like tubular structures induced by exposure of hMVECs to basic fibroblast growth factor and TNF-alpha. This was accompanied by a several-fold increase in u-PA accumulation in the conditioned medium. The effect of MAb H-2 was not caused by blocking cellular activation by u-PA/u-PAR interaction, as the amino-terminal fragment (ATF) of u-PA, which also activates u-PAR, prevented tube formation. In addition, the inhibition by MAb H-2 was not due to an effect of the antibody on u-PAR-vitronectin binding. These data show that inhibition of tube formation can be caused not only by inhibition of u-PA or plasmin activities but also by unavailability of the u-PAR for cell-bound proteolysis. Immunohistochemical analysis showed that in in vitro angiogenesis u-PAR and u-PA were localized on the invading, tube-forming hMVECs and not on the endothelial cells that are located on top of the fibrin matrix. u-PAR and u-PA were also prominently expressed on endothelial cells of neovessels present in an atherosclerotic plaque. These data may give more insight into the role of u-PAR in repair-associated angiogenesis.
Kroon
M E
ME
Gaubius Laboratory, Leiden University Hospital, Groningen Leiden The Netherlands.
Koolwijk
P
P
van Goor
H
H
Weidle
U H
UH
Collen
A
A
van der Pluijm
G
G
van Hinsbergh
V W
VW
D000911:Antibodies, Monoclonal / Q000378:metabolism / Q000494:pharmacology; D001161:Arteriosclerosis / Q000378:metabolism; D002352:Carrier Proteins / Q000494:pharmacology; D002478:Cells, Cultured; D016207:Cytokines / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D005337:Fibrin / Q000502:physiology; D005982:Glutathione Transferase / Q000494:pharmacology; D006801:Humans; D007150:Immunohistochemistry; D018919:Neovascularization, Physiologic / Q000187:drug effects / Q000502:physiology*; D010960:Plasminogen Activators / Q000276:immunology / Q000378:metabolism* / Q000502:physiology*; D011956:Receptors, Cell Surface / Q000276:immunology / Q000378:metabolism* / Q000502:physiology*; D055293:Receptors, Urokinase Plasminogen Activator; D011993:Recombinant Fusion Proteins / Q000494:pharmacology; D014568:Urokinase-Type Plasminogen Activator / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000493:pharmacokinetics; D019096:Vitronectin / Q000378:metabolism
D000911:Antibodies, Monoclonal; D002352:Carrier Proteins; D016207:Cytokines; C524898:PLAUR protein, human; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D011993:Recombinant Fusion Proteins; D019096:Vitronectin; D005337:Fibrin; C103332:GST-RAP protein, recombinant; D005982:Glutathione Transferase; D010960:Plasminogen Activators; D014568:Urokinase-Type Plasminogen Activator