Inappropriate plateletactivation is afeatureofacute and chronic diseasess uch as disseminated intravascular coagulation (DIC) and atherosclerosis. Since proinflammatorym icrobial-derived agonists can be involved in the pathogenesiso ft hese diseases, we examined the potential role ofTLR4 (mediating responsesto LPS) andTLR2(which responds to bacteriallipopeptides)inplateleta ctivation. Our datas uggestedl ow-level expression of TLR2 andTLR4 on platelets, determinedbyflowcytometry,and we also observed expression of TLR4 on amegakaryocytic cell line by bothflowcytometry and immunohistochemistry. Stimulation of the platelets with the TLR4 agonist LPS, and thes ynthetic TLR2 agonist Pam 3 CSK 4 ,r esulted in no plateleta ggregation,noincrease in CD62P surfaceexpression andnoincrease Keywords Platelets, TLR, LPS, atherosclerosis in thec ytosolic concentration of Ca 2+ .The TLRa gonists were also unablet od irectlya ctivate platelets primedw ith epinephrine,orpretreatedwith alow concentration ofADP or PAF. Pretreatment of platelets with LPSo rP am 3 CSK 4 also failedt o modulate thep lateletr esponset os ubmaximal concentrations of the classical plateletagonists ADP and PAF. We conclude that theTLRagonists LPS andPam 3 CSK 4 have no direct effect on plateleta ctivation and that plateletTLRsm ay be ar emnant from megakaryocytes.TLR2 and TLR4 agonists arethought to have a significant role in diseasessuch as atherosclerosis and DIC, but our research suggests that this is through am echanism other than directplateletactivation or by modification of plateletresponsestootheragonists.

Objective-Clinical studies suggest that platelet P2Y 12 inhibitors reduce mortality from sepsis, although the underlying mechanisms have not been clearly defined in vivo. We hypothesized that P2Y 12 inhibitors may improve survival from sepsis by suppressing systemic inflammation and its prothrombotic effects. We therefore determined whether clopidogrel and the novel, more potent P2Y 12 inhibitor, ticagrelor, modify these responses in an experimental human model. Approach and Results-We randomized 30 healthy volunteers to ticagrelor (n=10), clopidogrel (n=10), or no antiplatelet medication (controls; n=10). We examined the effect of P2Y 12 inhibition on systemic inflammation, which was induced by intravenous injection of Escherichia coli endotoxin. Both P2Y 12 inhibitors significantly reduced platelet-monocyte aggregate formation and peak levels of major proinflammatory cytokines, including tumor necrosis factor α, interleukin-6, and chemokine (C-C motif) ligand 2. In contrast to clopidogrel, ticagrelor also significantly reduced peak levels of IL-8 and growth colony-stimulating factor and increased peak levels of the anti-inflammatory cytokine IL-10. In addition, ticagrelor altered leukocyte trafficking. Both P2Y 12 inhibitors suppressed D-dimer generation and scanning electron microscopy revealed that ticagrelor also suppressed prothrombotic changes in fibrin clot ultrastructure. Ticagrelor is a novel P2Y 12 inhibitor, which causes more potent and consistent P2Y 12 inhibition than clopidogrel 13 and also weakly inhibits cellular uptake of adenosine. Conclusions-Potent14 In the PLATelet inhibition and patient Outcomes (PLATO) study of >18 000 patients with acute coronary syndromes, ticagrelor reduced all-cause mortality compared with clopidogrel (hazard ratio, 0.78; P<0.001), which was out of proportion to its incremental cardiovascular benefit.15 Intriguingly, ticagrelor was associated with lower mortality related to infection (hazard ratio, 0.67; P<0.05) 16,17 and fewer deaths after sepsis and pulmonary infections than clopidogrel. 18 We therefore sought to determine the mechanistic impact of P2Y 12 inhibitors on pathophysiological processes that are central to sepsis responses in humans. We hypothesized that P2Y 12 inhibitors may reduce mortality from sepsis by suppressing systemic inflammation and its prothrombotic effects, mediated by inhibition of platelet-leukocyte interactions. We hypothesized that the more potent P2Y 12 inhibitor, ticagrelor, suppresses these responses more potently than clopidogrel. To test these hypotheses in humans, we used a well-established model of systemic inflammation, which involves intravenous injection of Escherichia coli endotoxin (LPS) into healthy volunteers. 19 The particular strength of this unique model is that it allows direct assessment of dynamic cellular and molecular pathways that are also major mediators of the pathophysiology of sepsis in humans. Nonstandard Abbreviations and Acronyms CCL2chemokine (C-C motif) ligand 2 G-CSF growth colony-stimulating factor hs...

Ticagrelor 60 mg bid achieved high levels of peak and trough platelet inhibition in nearly all patients, similar to that with 90 mg bid, helping to explain the efficacy of the lower ticagrelor dose in PEGASUS-TIMI 54.

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