Agonists of toll-like receptor (TLR)2 and TLR4 are unable to modulate platelet activation by adenosine diphosphate and platelet activating factor

Abstract

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.

Human Cytomegalovirus–Platelet Interaction Triggers Toll-Like Receptor 2–Dependent Proinflammatory and Proangiogenic Responses

“…19 Platelets have been reported previously to express TLR2 on the surface of 4% to 16% platelets. [14][15][16] However, in our study, these levels were lower with 1.3% to 3% TLR2-positive platelets, which strongly correlated with the percentage of HCMV-bound platelets. Despite the low percentage of TLR2-positive platelets, ≈20% of platelets became P-selectin positive in response to HCMV, indicating that activation of the TLR2-positive subpopulation triggered further platelet activation in a TLR2-independent fashion.…”

“…[13][14][15] Interestingly, not all platelets express TLRs: 55% to 60% are positive for TLR4, 10% to 40% are positive for TLR9, and only 4% to 16% are positive for TLR2. [14][15][16] It was speculated that platelet expression of TLR might be due to the adsorption of free TLR in the plasma. However, megakaryocytes are also positive for TLRs, 14,17 and platelet TLR1/2, TLR4, and TLR9 have been proven to be functional because they trigger intracellular signaling, resulting in platelet activation.…”

CLEC‐2 signaling via Syk in myeloid cells can regulate inflammatory responses

“…However, rhodocytin, in our assays, activated DCs primarily via a MyD88-dependent mechanism, possibly due to contamination with TLR agonists, which obscured the putative CLEC-2/Syk-dependent response. This has not been seen in studies with platelets, in which the effect of rhodocytin is completely CLEC-2 dependent [20] but it should be noted that TLR agonists do not induce platelet aggregation [39] such that CLEC-2-independent effects of rhodocytin preparations might not be important for platelet studies. To circumvent this problem, we resorted to using our new mAb to selectively crosslink CLEC-2, inducing Syk-dependent calcium signaling and NFAT activation in both reporter and myeloid cells (Fig.…”

Hyaluronan Depolymerization by Megakaryocyte Hyaluronidase-2 Is Required for Thrombopoiesis

“…An increasing body of literature supports the role of HA-mediated signaling through several receptors, including CD44, RHAMM, and TLR-2 and TLR-4, all of which are expressed by MKs. 30,57,62 In particular, both RHAMM and . n Z 8 WT and 12 Hyal-2 KO mice (A); n Z 12 (B).…”