Ap4A acts as an antagonist of platelet P2Y1 and P2Y12 receptors (inhibiting ADP-induced aggregation with an IC50 of 9.8 ± 2.8 μM), an agonist of P2X1 receptors, and a partial agonist of P2Y12.
Abstract Introduction: Diadenosine 5′,5‴-P1,P4- tetraphosphate (Ap4A) is stored in platelet dense granules, but its effects on platelet function are not well understood. In the current study, we examined the effects of Ap4A on signaling through the human platelet purinergic receptors P2Y1, P2Y12 and P2X1. Methods: Flow cytometry was used to measure the effects of Ap4A in the presence or absence of ADP on: P2Y12-mediated decrease in intraplatelet phosphorylated vasodilator stimulated phosphoprotein (VASP), P2Y1- mediated increase in platelet cytosolic Ca2+ (measured with FLUO-4), and c) P2X1- mediated intraplatelet entry of extracellular Ca2+ (measured with FLUO-4) (Figure). ADP-stimulated platelet shape change (P2Y1-mediated) and aggregation (P2Y1- and P2Y12-mediated) were measured optically. Results: Ap4A inhibited 3 μM ADP-induced: platelet aggregation (IC50 9.8 ± 2.8 μM, n = 3), P2Y1-mediated shape change, P2Y1-mediated increase in platelet cytosolic Ca2+ (IC50 40.8 ± 12.3 μM, n = 3), and P2Y12-mediated decrease in VASP phosphorylation (IC50 250 μM, n = 3). In addition to inhibiting these ADP-induced effects on P2Y1 and P2Y12, Ap4A in the absence of added ADP decreased VASP phosphorylation, albeit with significantly reduced potency compared to ADP. This Ap4A-induced reduction of VASP phosphorylation was reversed by addition of AR-C69931, a selective P2Y12 antagonist. Ap4A, therefore, has partial P2Y12 agonist properties. Ap4A also had agonist effects on platelet P2X1 receptors, as evidenced by intraplatelet entry of extracellular Ca2+. All these effects of Ap4A were significant at pathophysiologic concentrations. Ap4A had no agonist effects on platelet P2Y1 receptors. Conclusions: Ap4A, a known constituent of platelet dense granules, is an antagonist of platelet P2Y1 and P2Y12 receptors, where it inhibits the effects of ADP, an agonist of P2X1 receptors, and a partial agonist of P2Y12 receptors (Figure). Figure Figure
Chang et al. (Sun,) conducted a other in Platelet function. Diadenosine 5′,5‴-P1,P4- tetraphosphate (Ap4A) vs. ADP was evaluated on Platelet aggregation, shape change, cytosolic Ca2+, and VASP phosphorylation. Ap4A acts as an antagonist of platelet P2Y1 and P2Y12 receptors (inhibiting ADP-induced aggregation with an IC50 of 9.8 ± 2.8 μM), an agonist of P2X1 receptors, and a partial agonist of P2Y12.