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PAR1-stimulated platelet releasate promotes angiogenic activities of endothelial progenitor cells more potently than PAR4-stimulated platelet releasate.

Journal of thrombosis and haemostasis : JTH (2014-12-17)
Z Huang, X Miao, Y Luan, L Zhu, F Kong, Q Lu, J Pernow, G Nilsson, N Li
ABSTRACT

Endothelial progenitor cells (EPCs) are important for endothelial regeneration and angiogenesis. Thrombin protease-activated receptor 1 (PAR1) PAR1 and PAR4 stimulation induces selective release of platelet proangiogenic and antiangiogenic regulators. To investigate if PAR1-stimulated platelet releasate (PAR1-PR) and PAR4-PR regulate angiogenic properties of EPCs in different manners. EPCs were generated from peripheral mononuclear cell culture. Washed platelets (2 × 10(9) mL(-1)) were stimulated by PAR1-activating peptide (PAR1-AP; 10 μmol L(-1)) or PAR4-AP (100 μmol L(-1)) to prepare PAR1-PR and PAR4-PR, respectively. PAR1-PR or PAR4-PR had little influence on EPC proliferation. EPC migration experiments using a modified Boyden chamber showed that both platelet releasates facilitated EPC migration. As for in vitro tube formation on Matrigel, PAR1-PR and PAR4-PR similarly enhanced capillary-like network formation of EPCs in the complete EPC medium containing 10% FBS and a cocktail of growth factors, while PAR1-PR more profoundly increased EPC tube formation in basal culture medium supplemented with only 0.5% FBS than did PAR4-PR. The latter was confirmed in the murine angiogenesis model of subcutaneous Matrigel implantation. Moreover, blockade of vascular endothelial growth factor, stromal cell-derived factor 1α, or matrix metalloproteinases attenuated EPC migration and tube formation, suggesting a cooperation of these factors in the enhancements. PAR1-PR enhances vasculogenesis more potently than PAR4-PR, and the enhancements require a cooperation of multiple platelet-derived angiogenic regulators.

MATERIALS
Product Number
Brand
Product Description

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Propidium iodide, ≥94.0% (HPLC)
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Propidium iodide, ≥94% (HPLC)
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Ala-Tyr-Pro-Gly-Lys-Phe-NH2 trifluoroacetate salt, ≥98% (HPLC), lyophilized powder
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Ser-Phe-Leu-Leu-Arg-Asn-amide trifluoroacetate salt, ≥98% (HPLC), lyophilized powder
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Propidium iodide solution, solution (1.0 mg/ml in water)