Abstract
Human platelets contain platelet-derived growth factor (PDGF) in their alpha-granules which is released during platelet exocytosis. We show by immunoprecipitation and 125I-PDGF binding experiments that human platelets have functionally active PDGF alpha-receptors, but not beta-receptors. The PDGF alpha-receptor (PDGFR-alpha) was identified as a 170-kDa glycosylated protein-tyrosine kinase as found in other cell types. Stimulation of platelets with 0.1 unit/ml thrombin resulted in a significant increase (2-5-fold) of the tyrosine phosphorylation of the PDGFR-alpha, as determined by immunoprecipitation with phosphotyrosine antiserum as well as with PDGFR-alpha antiserum. The observed thrombin-induced autophosphorylation of the PDGFR-alpha was inhibited by the addition of a neutralizing monoclonal PDGF antibody. Thus, our results suggest that the platelet PDGFR-alpha is stimulated in an autocrine manner by PDGF secreted during platelet activation. Preincubation of platelets with PDGF inhibited thrombin-induced platelet aggregation and secretion of ATP + ADP and beta-hexosaminidase. Thrombin-induced platelet aggregation was also reversed when PDGF was added 30 s after thrombin stimulation. Inhibition of the autocrine PDGF pathway during platelet activation by the PDGF antibody led to a potentiation of thrombin-induced beta-hexosaminidase secretion. Thus, the PDGFR-alpha takes part in a negative feedback regulation during platelet activation. Our demonstration of PDGF alpha-receptors on human platelets and its inhibitory function during platelet activation identifies a new possible role of PDGF in the regulation of thrombosis.
Highlights
Human platelets contain platelet-derivedgrowth fac- gesting normal physiological roolefsPDGF during wound healtor (PDGF)in theira-granules which is released during ing, placental growth, early embryogenesis, as well as during platelet exocytosis
ThePDGF a-receptor (PDGFR-a) wasiden- PDGF isoforms,PDGF-AA, - A B, and -BB, mediate their effects tified as a 1'70-kDaglycosylated protein-tyrosinekinase by binding to two protein-tyrosine kinase receptors (PDGFR), as found in other cell types
Thuos,ur re- tosis [1].Interestingly, the PDGFP-receptor and a-receptor sults suggest that theplatelet PDGFR-a is stimulated in haveagonisticandantagonisticeffects,respectively,onthe an autocrine manner by PDGF secreted during platelet chemotactic response of human fibroblasts [12, 13]
Summary
Human platelets contain platelet-derivedgrowth fac- gesting normal physiological roolefsPDGF during wound healtor (PDGF)in theira-granules which is released during ing, placental growth, early embryogenesis, as well as during platelet exocytosis. The addition of exogenous PDGF or PDGF antibodies inhibitedor potentiated thrombin-induced platelet responses, respectively. PDGF, PDGF antibodies, or vehicle for 60 s a t 37 "C before incubation In Vitro Immune Complex Kinase Assay-Human platelets and sub- with thrombin(0.04-0.3 unitlml final concentration)or vehicle for 60 s.
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