Aspirin Reduces the Potentiating Effect of CD40L on Platelet Aggregation via Inhibition of Myosin Light Chain.

Background Antiplatelet therapy with aspirin (acetylsalicylic acid [ASA]) is less efficient in some coronary patients, which increases their risk of developing thrombosis. Elevated blood levels of thromboinflammatory mediators, like soluble CD40L (sCD40L), may explain such variabilities. We hypothesized that in the presence of elevated levels of sCD40L, the efficacy of ASA may vary and aimed to determine the effects of ASA on CD40L signaling and aggregation of platelets. Methods and Results The effects of ASA on CD40L-treated human platelets, in response to suboptimal concentrations of collagen or thrombin, were assessed at levels of aggregation, thromboxane A2 secretion, and phosphorylation of p38 mitogen-activated protein kinase, nuclear factor kappa B, transforming growth factor-ß-activated kinase 1, and myosin light chain. sCD40L significantly elevated thromboxane A2 secretion in platelets in response to suboptimal doses of collagen and thrombin, which was reversed by ASA. ASA did not inhibit the phosphorylation of p38 mitogen-activated protein kinase, nuclear factor kappa B, and transforming growth factor-ß-activated kinase 1, with sCD40L stimulation alone or with platelet agonists. sCD40L potentiated platelet aggregation, an effect completely reversed and partially reduced by ASA in response to a suboptimal dose of collagen and thrombin, respectively. The effects of ASA in sCD40L-treated platelets with collagen were related to inhibition of platelet shape change and myosin light chain phosphorylation. Conclusions ASA does not affect platelet sCD40L signaling but prevents its effect on thromboxane A2 secretion and platelet aggregation in response to collagen, via a mechanism implying inhibition of myosin light chain. Targeting the sCD40L axis in platelets may have a therapeutic potential in patients with elevated levels of sCD40L and who are nonresponsive or less responsive to ASA.

CD40L Priming of Platelets via NF-κB Activation is CD40- and TAK1-Dependent.

Background CD40 ligand (CD40L) is a thromboinflammatory molecule that predicts cardiovascular events. CD40L is a strong activator of nuclear factor kappa B (NF-κB) in platelets that primes and enhances platelet activation in response to thrombotic stimuli. In addition to its classical receptor CD40, CD40L binds αIIbß3, α5ß1, and αMß2 in various cell types. However, the function of the different CD40L receptors on platelets remains unexplored. The present study aims to identify the receptors of CD40L, involved in platelet NF-κB activation, their downstream signaling and their implication in platelet aggregation. Methods and Results We showed that platelets express CD40, αIIbß3, and α5ß1 and release CD40L in response to sCD40L stimulation. sCD40L alone dose-dependently induced platelet NF-κB activation; this effect was absent in CD40-/- mouse platelets and inhibited by the CD40 blockade, but was unaffected by the αIIbß3 or α5ß1 blockade in human platelets. sCD40L/CD40 axis activates transforming growth factor-ß-activated kinase 1 upstream of NF-κB. In functional studies, sCD40L alone did not affect platelet aggregation but potentiated the aggregation response in the presence of suboptimal doses of thrombin; this effect was abolished by CD40, transforming growth factor-ß-activated kinase 1, and NF-κB inhibitors. Conclusions CD40L primes platelets via signaling pathways involving CD40/transforming growth factor-ß-activated kinase 1/NF-κB, which predisposes platelets to enhanced activation and aggregation in response to thrombotic stimuli.