Mechanism of Race-Dependent Platelet Activation Through the Protease-Activated Receptor-4 and Gq Signaling Axis

Abstract

Objective Black individuals are at an increased risk of myocardial infarction and stroke, two vascular diseases with strong thrombotic components. Platelet activation is a key step in platelet clot formation leading to myocardial infarction and stroke and recent work supports a racial difference in platelet aggregation through the thrombin protease-activated receptors (PARs). The underlying mechanism for this racial difference, however has not been established. Determining where in the signaling cascade these racial differences emerge will aid in understanding why individuals of differing racial ancestry may possess an inherent difference in their responsiveness to anti-platelet therapies. Approach and Results Washed human platelets from black volunteers were hyper-aggregable in response to PAR4-mediated platelet stimulation compared to whites. Interestingly, the racial difference in PAR4-mediated platelet aggregation persisted in platelets treated ex vivo with aspirin and 2MeSAMP, suggesting that the racial difference is independent of secondary feedback. Further, stimulation of platelets from black donors with PAR4-activating peptide (PAR4-AP) showed a potentiated level of activation through the Gq pathway compared to platelets from white donors. Differences in signaling included increased Ca2+ mobilization, Rap1 activation, and integrin αIIbβ3 activation with no observed difference in platelet protein expression between the groups tested. Conclusions Our study is the first to demonstrate that the Gq pathway is differentially regulated by race following PAR4 stimulation in human platelets. Furthermore, the racial difference in PAR4-mediated platelet aggregation persisted in the presence of COX and P2Y12 dual inhibition, suggesting that current anti-platelet therapy may provide less protection to blacks than whites.

Effects of genetic variation in protease activated receptor 4 after an acute coronary syndrome: Analysis from the TRACER trial

“…To our knowledge this is the first study testing for associations between a common thrombin receptor polymorphism and clinical outcomes in a large, randomized clinic trial of patients with NSTE ACS [26]. Although thrombin receptors have a broad tissue distribution, our hypotheses were based on abundant in vitro data using human blood that demonstrated enhanced platelet activation and aggregation from healthy individuals expressing the rs773902 A allele encoding Thr in the PAR4 thrombin receptor [23, 24, 27]. The most important finding from our work was that the rs773902 AA genotype conferred a significant reduction in GUSTO moderate or severe bleeding.…”

12-HETrE inhibits platelet reactivity and thrombosis in part through the prostacyclin receptor

“…Platelets were isolated from whole blood via serial centrifugation, as previously reported. 3 Briefly, plateletrich plasma acquired via centrifugation of whole blood at 200 g for 10 minutes was treated with acid citrate dextrose (2.5% sodium citrate tribasic, 1.5% citric acid, 2.0% D-glucose) and apyrase (0.02 U/mL) and then spun for 10 minutes at 2000g. Platelets were resuspended to a final concentration of 3.0 3 10 8 platelets/mL in Tyrode's buffer (N-2-hydroxyethylpiperazine-N9-2-ethanesulfonic acid 10 mM, sodium bicarbonate 12 mM, sodium chloride 127 mM, potassium chloride 5 mM, monosodium phosphate 0.5 mM, magnesium chloride 1 mM, and glucose 5 mM) as determined by a complete blood cell counter (Hemavet 950FS; Drew Scientific, Miami Lakes, FL).…”

Genomics and transcriptomics of megakaryocytes and platelets: Implications for health and disease

“…PAR4 surface expression was not different between white and black individuals, meaning that the difference in aggregation and reactivity is due to PAR4 mediated signaling rather than variable surface expression of PAR4. Further studies demonstrate that the downstream Gq signaling axis is altered in black individuals upon PAR4‐mediated platelet stimulation 58. Interestingly, miR‐376c, which regulates the expression of PCTP mRNA in human megakaryocytes, was differentially expressed by race.…”