Submitted on December 21, 2007
Revised on July 16, 2008
Accepted on August 12, 2008
Platelets Undergo Phosphorylation of Syk at Y525/526 and Y352 in Response to Pathophysiological Shear Stress
Henry E Speich1, Svetozar Grgurevich2, Teddi J Kueter2, Angela D Earhart2, Steven M Slack3, and Lisa K Jennings2*
1 The University of Memphis/The University of Tennessee Health Science Center
2 University of Tennessee Health Science Center
3 University of Memphis
* To whom correspondence should be addressed. E-mail: ljennings{at}utmem.edu.
Atherosclerotic plaques can lead to partial vascular occlusions that produce abnormally high levels of arterial wall shear stress. Such pathophysiological shear stress can promote shear-induced platelet aggregation (SIPA), which has been linked to acute myocardial infarction, unstable angina and stroke. This study investigated the role of the tyrosine kinase Syk in shear-induced human platelet signaling. The extent of Syk tyrosine phosphorylation induced by pathophysiological levels of shear stress (100 dynes/cm^2) was significantly greater than that resulting from physiological shear stress (10 dynes/cm^2). Using phospho-Syk specific antibodies, these data are the first to show that key regulatory sites of Syk at tyrosines 525/526 (Y525/526) and tyrosine 352 (Y352) were phosphorylated in response to pathophysiological shear stress. Increased phosphorylation at both sites was attenuated by pharmacologic inhibition of Syk using two different Syk inhibitors, piceatannol and 3-(1-Methyl-1H-indol-3-yl-methylene)-2-oxo-2,3-dihydro-1H-indole-5-sulfonamide (OXSI-2), and by inhibition of upstream Src-family kinases (SFKs). Shear-induced response at the Syk 525/526 site was ADP-dependent, but not contingent upon glycoprotein (GP) IIb-IIIa ligation or the generation of thromboxane (Tx) A2. Pretreatment with Syk inhibitors not only reduced SIPA and Syk phosphorylation in isolated platelets, but also diminished, up to 50%, the platelet-mediated thrombus formation when whole blood was perfused over type-III collagen. In summary, this study demonstrated that Syk is a key molecule in both SIPA and thrombus formation under flow. Pharmacological regulation of Syk may prove efficacious in treating occlusive vascular disease.
