Dwivedi, Suman; Pandey, Dharmendra; Khandoga, Anna L.; Brandl, Richard; Siess, Wolfgang:
Rac1-mediated signaling plays a central role in secretion-dependent platelet aggregation in human blood stimulated by atherosclerotic plaque.
In: Journal of Translational Medicine
Background: Platelet activation requires rapid remodeling of the actin cytoskeleton which is regulated by small GTP-binding proteins. By using the Rac1-specific inhibitor NSC23766, we have recently found that Rac1 is a central component of a signaling pathway that regulates dephosphorylation and activation of the actin-dynamising protein cofilin, dense and alpha granule secretion, and subsequent aggregation of thrombin-stimulated washed platelets. Objectives: To study whether NSC23766 inhibits stimulus-induced platelet secretion and aggregation in blood. Methods: Human platelet aggregation and ATP-secretion were measured in hirudin-anticoagulated blood and platelet-rich plasma (PRP) by using multiple electrode aggregometry and the Lumi-aggregometer. Platelet P-selectin expression was quantified by flow cytometry. Results: NSC23766 (300 mu M) inhibited TRAP-, collagen-, atherosclerotic plaque-, and ADP-induced platelet aggregation in blood by 95.1%, 93.4%, 92.6%, and 70%, respectively. The IC50 values for inhibition of TRAP-, collagen-, and atherosclerotic plaque-, were 50 +/- 18 mu M, 64 +/- 35 mu M, and 50 +/- 30 mu M NSC23766 (mean +/- SD, n = 3-7), respectively. In blood containing RGDS to block integrin alpha(IIb)beta(3)-mediated platelet aggregation, NSC23766 (300 mu M) completely inhibited P-selectin expression and reduced ATP-secretion after TRAP and collagen stimulation by 73% and 85%, respectively. In ADP-stimulated PRP, NSC23766 almost completely inhibited P-selectin expression, in contrast to aspirin, which was ineffective. Moreover, NSC23766 (300 mu M) decreased plaque-stimulated platelet adhesion/aggregate formation under arterial flow conditions (1500s(-1)) by 72%. Conclusions: Rac1-mediated signaling plays a central role in secretion-dependent platelet aggregation in blood stimulated by a wide array of platelet agonists including atherosclerotic plaque. By specifically inhibiting platelet secretion, the pharmacological targeting of Rac1 could be an interesting approach in the development of future antiplatelet drugs.