Stimulation of PAR-1 or PAR-4 promotes similar pattern of VEGF and endostatin release and pro-angiogenic responses mediated by human platelets

• Published in: Platelets (Edinburgh) 2015-11, Vol.26 (8), p.799-804
• Main Authors: Etulain, J., Mena, H. A., Negrotto, S., Schattner, M.
• Format: Article
• Language: English
• Subjects: Angiogenesis; Blood Platelets - drug effects; Blood Platelets - metabolism; endostatin; Endostatins - secretion; Endothelial Cells - metabolism; Humans; Neovascularization, Physiologic; Oligopeptides - pharmacology; Platelet Activation - drug effects; platelet alpha-granules; protease-activated receptors; Receptor, PAR-1 - agonists; Receptors, Thrombin - agonists; Vascular Endothelial Growth Factor A - metabolism; VEGF
• ISSN: 0953-7104; 1369-1635
• DOI: 10.3109/09537104.2015.1051953

Background: Platelets mediate angiogenesis through the secretion of several factors, including the pro-angiogenic vascular endothelial growth factor (VEGF) and the anti-angiogenic endostatin. Although previous findings indicated that these molecules are packed into different alpha-granules and selectively released by specific stimulation of protease-activated receptor (PAR)-1 or PAR-4, recent evidences are against this hypothesis. Objectives: To elucidate the controversies about the VEGF and endostatin release and the overall angiogenic effect of PARs-stimulated platelets. Methods: VEGF and endostatin were quantified by enzyme linked-immunosorbent assay (ELISA). Endothelial proliferation (pNPP assay), wound healing (scratch assay) and tubule formation (matrigel) of human microvascular endothelial cells (HMEC-1) and endothelial progenitor cells (EPC) were determined using supernatants from PAR-1- or PAR-4-stimulated platelets. Results: Activation of washed platelets (WPs) by PAR-1- or PAR-4-activating peptide (AP) promoted the VEGF and endostatin secretion in a concentration-dependent manner, being PAR-1-AP more potent than PAR-4-AP. The release of both molecules was abrogated by pre-incubation of platelets with PAR antagonists. Activation of platelet-rich plasma (PRP) with either PAR-1-AP or PAR-4-AP induced a significant VEGF secretion. Quantification of platelet-endostatin secretion was not possible in PRP due to the high levels of plasmatic endostatin vs. platelet content. Releasates from PAR-1- or PAR-4-activated WPs promoted similar pattern of angiogenic responses of HMEC-1 or EPC. Moreover, proliferation of HMEC-1 mediated by PAR-stimulated PRP releasates was delayed and significantly lower compared with that induced by PAR-stimulated WPs. Conclusions: Our results are in contrast with the previously described differential release of VEGF and endostatin induced by the selective PAR-1 or PAR-4 stimulation, and support the notion that while circulating endostatin accounts for the maintenance of a systemic anti-angiogenic state, locally, the release of platelet alpha-granule content promotes angiogenesis.