A New Method to Measure P2Y12 Receptor Expression on the Surface of Human Platelets: Effect of Clopidogrel

Abstract number: P1568

Udaya T, Kevin PB, Jason Y, Kevin H, Rolf K, Paul AG

Background:  The measurement of P2Y12 receptor expression by flow cytometry has not been previously described and the effect of clopidogrel on the expression of this receptor is unknown.

Methods:  Healthy volunteers (n= 6) were treated with clopidogrel (300 mg) and aspirin (325 mg). Blood was analyzed at baseline, 6, 24 and 120 hours post-treatment. 5 and 20 uM ADP and 1 mM AA-induced platelet aggregation (LTA); unsrtimulated (UST) and stimulated (ST) GP IIb/IIIa and P-selectin expression were measured. Intracellular VASP phosphorylation was measured by flow cytometry. PP2Y12 receptor density was measured by a novel method developed in the our lab using labeled antibodies and flow cytometry.

Results:  Clopidogrel markedly decreased platelet aggregation (83 ± 10% pre vs. 40 ± 11% post, P<= 0.001), stimulated GP IIb/IIIa expression (153 ± 46 pre vs. 62 ± 27 MFI post, P < 0.001), and P-selectin expression (38 ± 13 pre. Vs. 9 ± 5, P<= 0.001) and preserved the VASP index (I) of P2Y12 reactivity. Clopidogrel loading also has an immediate effect on lowering P2Y12 expression (10 + 3 pre vs. 6.4 + 2 MFI post, P < 0.01) at 6 hours.

Conclusion:  Clopidogrel effectively attenuates platelet reactivity by inhibiting GP IIb/IIIa expression via the preservation of VASP phosphorylation levels. It is inhibits the granular secretion of P-selectin. Early attenuation of the expression of P2Y12 is a newly described effect of clopidogrel and may contribute to its immediate antithrombotic effects.