Platelet adhesion to collagen and collagen-related peptide under flow: roles of the [alpha]2[beta]1 integrin, GPVI, and Src tyrosine kinases.
OBJECTIVE
Platelet stimulation by collagen and collagen-related peptides (CRPs) is associated with activation of protein tyrosine kinases. In the present study, we investigated the role of Src family tyrosine kinases in the initial adhesion events of human platelets to collagen and cross-linked CRP.


RESULTS
Under arterial flow conditions, a glycoprotein VI-specific substrate, cross-linked CRP, caused rapid (<2 second) platelet retention and protein tyrosine phosphorylation that were markedly decreased by the Src family kinase inhibitor pyrozolopyrimidine (PP2) or by aggregation inhibitor GRGDSP. CRP-induced platelet retention was transient, and 90% of single platelets or aggregates detached within seconds. PP2, although having no effect on RGD peptide-binding to CRP, completely blocked aggregation and tyrosine phosphorylation of Syk and phospholipase Cgamma2 (PLCgamma2). In contrast, PP2 weakly (<30%) suppressed firm adhesion to collagen mediated primarily by the alpha2beta1 integrin. Although PP2 prevented activation of Syk and PLCgamma2 in collagen-adherent platelets, tyrosine phosphorylation of several unidentified protein bands persisted, as did autophosphorylation of pp125FAK.


CONCLUSIONS
These findings indicate that activation of Src-tyrosine kinases Syk and PLCgamma2 is not required for the initial stable attachment of human platelets to collagen and for FAK autophosphorylation. However, Src-tyrosine kinases are critical for glycoprotein VI-mediated signaling leading to platelet aggregation.
Polanowska-Grabowska
Renata
R
Department of Biochemistry and Molecular Genetics, University of Virginia, Charlottesville, VA 22908, USA. rp4t@virginia.edu
Gibbins
Jonathan M
JM
Gear
Adrian R L
AR
D002352:Carrier Proteins / Q000502:physiology*; D003094:Collagen / Q000502:physiology*; D006801:Humans; D016023:Integrins / Q000502:physiology; D010455:Peptides; D010766:Phosphorylation; D010973:Platelet Adhesiveness / Q000187:drug effects* / Q000502:physiology; D010980:Platelet Membrane Glycoproteins / Q000502:physiology; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011743:Pyrimidines / Q000494:pharmacology; D019061:src-Family Kinases / Q000037:antagonists & inhibitors / Q000502:physiology*
C412373:AG 1879; D002352:Carrier Proteins; D016023:Integrins; D010455:Peptides; D010980:Platelet Membrane Glycoproteins; D011743:Pyrimidines; C435908:collagen-related peptide; C098341:platelet membrane glycoprotein VI; D003094:Collagen; D011505:Protein-Tyrosine Kinases; D019061:src-Family Kinases