Generation of platelet angiostatin mediated by urokinase plasminogen activator: effects on angiogenesis

Abstract

To cite this article: Jurasz P, Santos-Martinez MJ, Radomska A, Radomski MW. Generation of platelet angiostatin mediated by urokinase plasminogen activator: effects on angiogenesis. J Thromb Haemost 2006; 4: 1095-106.Summary. Background: Angiogenesis, the growth of new capillaries from pre-existing blood vessels, is regulated by a balance between its promoters and inhibitors. Platelets are an important circulating store of angiogenesis regulators. We have previously identified the angiogenesis inhibitor angiostatin in human platelets. Aim: To identify the mechanism of platelet angiostatin generation and its pharmacological regulation. Methods: Platelet aggregometry, flow cytometry, Western blot, zymography, immunofluorescence microscopy, matrigelinduced angiogenesis of human umbilical vein endothelial cells (HUVECs), and a panel of selective proteinase inhibitors were used to study the mechanism of angiostatin generation by platelets, its pharmacological regulation, and effects on angiogenesis. Release of pro-MMP-2 by HUVECs was also used to quantify angiogenesis. Results: Platelet membranes were identified as the site of angiostatin generation from plasminogen. Generation of angiostatin by platelet membranes was not affected by a matrix metalloproteinase (MMP) inhibitor, phenanthroline, but was inhibited by serine proteinase inhibitors aprotinin, leupeptin, plasminogen activator inhibitor-1, and selective inhibitor of urokinase plasminogen activator (uPA), uPA-STOP TM . Angiostatin generation by intact platelets was inhibited by aprotinin, and the resulting incubate promoted angiogenesis to a greater extent than incubate where angiostatin generation occurred. Furthermore, HUVECs incubated with reaction mixture, where angiostatin generation was inhibited, released more pro-MMP-2 than HUVECs incubated with supernatants, where angiostatin generation occurred.Conclusions: We conclude that; (i) platelets constitutively generate angiostatin on their membranes; (ii) this mechanism is dependent on uPA, but not, MMPs; and (iii) inhibition of platelet angiostatin generation can further promote angiogenesis.

Activated Platelets Provide a Functional Microenvironment for the Antiangiogenic Fragment of Histidine-Rich Glycoprotein

“…Platelets contain plasminogen and enzymes such as matrix metalloproteinases and urokinase-type plasminogen activator, capable of generating angiostatin. Platelets release functional angiostatin, but the inhibitor is also generated from plasminogen during platelet activation and aggregation (36,37). PF4 and angiostatin exemplify two different mechanisms used by platelets to increase the local concentration of an angiogenesis inhibitor: release from α-granules or generation of the inhibitor during platelet activation.…”

RNA interference-mediated targeting of urokinase plasminogen activator receptor and matrix metalloproteinase-9 gene expression in the IOMM-lee malignant meningioma cell line inhibits tumor growth, tumor cell invasion and angiogenesis

“…Angiostatin is an angiogenesis inhibitor generated by the membranes of platelets. Inhibition of generation of angiostatin and a subsequent favorable effect on angiogenesis was reported by Jurasz et al (60) in HUVEC cells.…”

Pharmacologic Protein Kinase CαInhibition Uncouples Human Platelet-Stimulated Angiogenesis from Collagen-Induced Aggregation

“…In vitro angiogenesis assays in which endothelial cells align and form hollow tube capillary-like structures on Matrigel were performed as described previously elsewhere (Jurasz et al, 2003(Jurasz et al, , 2006 with the following modifications. Briefly, Matrigel was mixed with platelet releasates (1:3) on ice and allowed to solidify at 37°C for 30 minutes in 96-well plates.…”