NF-κB Links TLR2 and PAR1 to Soluble Immunomodulator Factor Secretion in Human Platelets (original) (raw)
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Thrombosis Research, 2014
Introduction: Platelets express Toll-like receptors (TLRs) that recognise molecular components of pathogens and, in nucleated cells, elicit immune responses through nuclear factor-kappaB (NF-κB) activation. We have shown that NF-κB mediates platelet activation in response to classical agonists, suggesting that this transcription factor exerts non-genomic functions in platelets. The aim of this study was to determine whether NF-κB activation is a downstream signal involved in TLR2 and 4-mediated platelet responses. Material and methods: Aggregation and ATP release were measured with a Lumi-aggregometer. Fibrinogen binding, P-selectin and CD40 ligand (CD40L) levels and platelet-neutrophil aggregates were measured by cytometry. I kappa B alpha (IκBα) degradation and p65 phosphorylation were determined by Western blot and von Willebrand factor (vWF) by ELISA. Results: Platelet stimulation with Pam3CSK4 or LPS resulted in IκBα degradation and p65 phosphorylation. These responses were suppressed by TLR2 and 4 blocking and synergised by thrombin. Aggregation, fibrinogen binding and ATP and vWF release were triggered by Pam3CSK4. LPS did not induce platelet responses per se, except for vWF release, but it did potentiate thrombin-induced aggregation, fibrinogen binding and ATP secretion. Pam3CSK4, but not LPS, induced P-selectin and CD40L expression and mixed aggregate formation. All of these responses, except for CD40L expression, were inhibited in platelets treated with the NF-κB inhibitors BAY 11-7082 or Ro 106-9920. Conclusion: TLR2 and 4 agonists trigger platelet activation responses through NF-κB. These data show another non-genomic function of NF-κB in platelets and highlight this molecule as a potential target to prevent platelet activation in inflammatory or infectious diseases.
Platelet Innate Immune Receptors and TLRs: A Double-Edged Sword
International Journal of Molecular Sciences, 2021
Platelets are hematopoietic cells whose main function has for a long time been considered to be the maintenance of vascular integrity. They have an essential role in the hemostatic response, but they also have functional capabilities that go far beyond it. This review will provide an overview of platelet functions. Indeed, stress signals may induce platelet apoptosis through proapoptotis or hemostasis receptors, necrosis, and even autophagy. Platelets also interact with immune cells and modulate immune responses in terms of activation, maturation, recruitment and cytokine secretion. This review will also show that platelets, thanks to their wide range of innate immune receptors, and in particular toll-like receptors, and can be considered sentinels actively participating in the immuno-surveillance of the body. We will discuss the diversity of platelet responses following the engagement of these receptors as well as the signaling pathways involved. Finally, we will show that while pl...
Evidence of Toll-like receptor molecules on human platelets
Immunology and Cell Biology, 2005
Platelets are primarily involved in thrombosis and haemostasis, and they have recently been shown to have a role in innate immunity and in inflammation. We have determined the markers of innate immunity that are expressed by platelets, specifically the Toll-like receptors (TLR), originating from mixes of platelet concentrates (MPC, n = 5) between day zero and day five after blood collection. The surface membrane and intracellular expression of TLR were measured, both after and without permeabilization, using flow cytometry. We observed weak expression of TLR2, TLR4 and TLR9 on the surface of CD41 + platelets. The expression levels of TLR4 were high (59 ± 2.2%). Moreover, there was a significant expression of TLR2 (47.5 ± 4.8%), TLR4 (78.8 ± 1.3%) and TLR9 (34.2 ± 7.5%) in the cytoplasm of CD41 + platelets. The expression of the three receptors did not change significantly during the course of the 5 day observation period. The percentage of TLR expression is significantly modulated between activated versus non-activated platelets, both after and without permeabilization ( P < 0.01). Study of the expression of TLR could increase our knowledge of the level of platelet participation during an immune reaction and inflammation. In the same way as the platelet ligand/receptor pair CD40L/CD40 is, the TLR are expressed by platelets, and could serve as a link between innate and adaptive immunity.
Innate immune receptors in platelets and platelet‐leukocyte interactions
Journal of Leukocyte Biology, 2020
Platelets are chief cells in hemostasis. Apart from their hemostatic roles, platelets are major inflammatory effector cells that can influence both innate and adaptive immune responses. Activated platelets have thromboinflammatory functions linking hemostatic and immune responses in several physiological and pathological conditions. Among many ways in which platelets exert these functions, platelet expression of pattern recognition receptors (PRRs), including TLR, Nod-like receptor, and C-type lectin receptor families, plays major roles in sensing and responding to pathogen-associated or damage-associated molecular patterns (PAMPs and DAMPs, respectively). In this review, an increasing body of evidence is compiled showing the participation of platelet innate immune receptors, including PRRs, in infectious diseases, sterile inflammation, and cancer. How platelet recognition of endogenous DAMPs participates in sterile inflammatory diseases and thrombosis is discussed. In addition, platelet recognition of both PAMPs and DAMPs initiates platelet-mediated inflammation and vascular thrombosis in infectious diseases, including viral, bacterial, and parasite infections. The study also focuses on the involvement of innate immune receptors in platelet activation during cancer, and their contribution to tumor microenvironment development and metastasis. Finally, how innate immune receptors participate in platelet communication with leukocytes, modulating leukocyte-mediated inflammation and immune functions, is highlighted. These cell communication processes, including platelet-induced release of neutrophil extracellular traps, platelet Ag presentation to T-cells and platelet modulation of monocyte cytokine secretion are discussed in the context of infectious and sterile diseases of major concern in human health, including cardiovascular diseases, dengue, HIV infection, sepsis, and cancer.
Inhibition of constitutive NF-κB activity induces platelet apoptosis via ER stress
Biochemical and biophysical research communications, 2017
Platelets are anucleate cells, known for their pivotal roles in hemostasis, inflammation, immunity, and disease progression. Being anuclear, platelets are known to express several transcriptional factors which exert nongenomic functions, including the positive and negative regulation of platelet activation. NF-κB is one such transcriptional factor involved in the regulation of genes for survival, proliferation, inflammation and immunity. Although, the role NF-κB in platelet activation and aggregation is partially known, its function in management of platelet survival and apoptosis remain unexplored. Therefore, two unrelated inhibitors of NF-κB activation, BAY 11-7082 and MLN4924 were used to determine the role of NF-κB in platelets. Inhibition of NF-κB caused decreased SERCA activity and increased cytosolic Ca level causing ER stress which was determined by the phosphorylation of eIF2-α. Further, there was increased BAX and decreased BCl-2 levels, incidence of mitochondrial membrane...
2005
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.
Biochemical Journal, 2011
PARs (protease-activated receptors) 1 and 4 belong to the family of G-protein-coupled receptors which induce both Gα12/13 and Gαq signalling. By applying the specific PAR1- and PAR4-activating hexapeptides, SFLLRN and AYPGKF respectively, we found that aggregation of isolated human platelets mediated via PAR1, but not via PAR4, is abolished upon homologous receptor activation in a concentration- and time-dependent fashion. This effect was not due to receptor internalization, but to a decrease in Ca2+ mobilization, PKC (protein kinase C) signalling and α-granule secretion, as well as to a complete lack of dense granule secretion. Interestingly, subthreshold PAR4 activation rapidly abrogated PAR1 signalling desensitization by differentially reconstituting these affected signalling events and functional responses, which was sufficient to re-establish aggregation. The lack of ADP release and P2Y12 receptor-induced Gαi signalling accounted for the loss of the aggregation response, as mim...