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CC BY 4.0 · TH Open 2019; 03(02): e94-e102
DOI: 10.1055/s-0039-1685495
Original Article
Georg Thieme Verlag KG Stuttgart · New York

The Role of Human Platelet Preparation for Toll-Like Receptors 2 and 4 Related Platelet Responsiveness

Juergen Koessler 
1   Institute of Clinical Transfusion Medicine and Haemotherapy, University of Würzburg, Würzburg, Germany
,
Marius Niklaus
1   Institute of Clinical Transfusion Medicine and Haemotherapy, University of Würzburg, Würzburg, Germany
,
Katja Weber
1   Institute of Clinical Transfusion Medicine and Haemotherapy, University of Würzburg, Würzburg, Germany
,
Angela Koessler
1   Institute of Clinical Transfusion Medicine and Haemotherapy, University of Würzburg, Würzburg, Germany
,
Sabine Kuhn
1   Institute of Clinical Transfusion Medicine and Haemotherapy, University of Würzburg, Würzburg, Germany
,
Markus Boeck
1   Institute of Clinical Transfusion Medicine and Haemotherapy, University of Würzburg, Würzburg, Germany
,
Anna Kobsar
1   Institute of Clinical Transfusion Medicine and Haemotherapy, University of Würzburg, Würzburg, Germany
› Author Affiliations Funding This publication was funded by the German Research Foundation (DFG) and the University of Würzburg in the funding program Open Access Publishing. Juergen Koessler (project number KO 5256/3–1) and Anna Kobsar (project number KO 5294/2–1) have received grants from Deutsche Forschungsgemeinschaft (DFG).
Further Information

Publication History

12 November 2018

25 February 2019

Publication Date:
17 April 2019 (online)

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Abstract

Background Like immune cells, platelets express the repertoire of toll-like receptors (TLR), among them TLR2 and TLR4, which are important for the recognition of bacterial patterns. Receptor-mediated functional effects in platelets have been investigated, but reliable conclusions are tampered due to heterogeneous study designs with variable platelet preparation methods. This study compares TLR2- and TLR4-dependent platelet responsiveness in platelet-rich plasma (PRP) and in washed platelets (WPs).

Material and Methods Fresh peripheral blood samples from healthy donors served for the preparation of PRP and WP. Basal and agonist-stimulated TLR2 and TLR4 expression levels were evaluated by flow cytometry. Light transmission aggregometry was used to investigate functional effects of TLR2 and TLR4 stimulation with Pam3CSK4 or LPS (lipopolysaccharides from Escherichia coli) as ligands. The capacity of chemokine release was determined by immunoassays.

Results Pam3CSK4 and LPS (in combination with thrombin) were able to induce aggregation in WP, but not in PRP, with threshold concentrations of 15 µg/mL. Basal expression levels of TLR2 and TLR4 were higher in WP than in PRP, increasing several-fold rapidly and persistently upon platelet activation with potent agonists. Pam3CSK4 (15 µg/mL) or LPS led to the submaximal release of RANTES, PF4, PDGF, NAP-2, and sCD40L from WP. In PRP, secretory effects are less pronounced for RANTES, PDGF, or PF4, and not detectable for NAP-2 or sCD40L.

Conclusion The effects mediated by TLR2 and TLR4 stimulation are dependent on platelet preparation, an important issue for experimental designs and for manufacturing of platelet concentrates in transfusion medicine.

Keywords

toll-like receptor - lipopolysaccharides - chemokine - platelet preparation
 

  • References

  • 1 Cognasse F, Nguyen KA, Damien P. , et al. The inflammatory role of platelets via their TLRs and Siglec receptors. Front Immunol 2015; 6: 83
    CrossrefPubMedGoogle Scholar
  • 2 Morrell CN, Aggrey AA, Chapman LM, Modjeski KL. Emerging roles for platelets as immune and inflammatory cells. Blood 2014; 123 (18) 2759-2767
    CrossrefPubMedGoogle Scholar
  • 3 von Hundelshausen P, Weber C. Platelets as immune cells: bridging inflammation and cardiovascular disease. Circ Res 2007; 100 (01) 27-40
    CrossrefPubMedGoogle Scholar
  • 4 Cognasse F, Hamzeh H, Chavarin P, Acquart S, Genin C, Garraud O. Evidence of Toll-like receptor molecules on human platelets. Immunol Cell Biol 2005; 83 (02) 196-198
    CrossrefPubMedGoogle Scholar
  • 5 Shiraki R, Inoue N, Kawasaki S. , et al. Expression of Toll-like receptors on human platelets. Thromb Res 2004; 113 (06) 379-385
    CrossrefPubMedGoogle Scholar
  • 6 Hamzeh-Cognasse H, Damien P, Chabert A, Pozzetto B, Cognasse F, Garraud O. Platelets and infections - complex interactions with bacteria. Front Immunol 2015; 6: 82
    PubMedGoogle Scholar
  • 7 Blair P, Rex S, Vitseva O. , et al. Stimulation of toll-like receptor 2 in human platelets induces a thromboinflammatory response through activation of phosphoinositide 3-kinase. Circ Res 2009; 104 (03) 346-354
    CrossrefPubMedGoogle Scholar
  • 8 Rex S, Beaulieu LM, Perlman DH. , et al. Immune versus thrombotic stimulation of platelets differentially regulates signalling pathways, intracellular protein-protein interactions, and alpha-granule release. Thromb Haemost 2009; 102 (01) 97-110
    Article in Thieme ConnectPubMedGoogle Scholar
  • 9 Kälvegren H, Skoglund C, Helldahl C, Lerm M, Grenegård M, Bengtsson T. Toll-like receptor 2 stimulation of platelets is mediated by purinergic P2 × 1-dependent Ca2+ mobilisation, cyclooxygenase and purinergic P2Y1 and P2Y12 receptor activation. Thromb Haemost 2010; 103 (02) 398-407
    Article in Thieme ConnectPubMedGoogle Scholar
  • 10 Aslam R, Speck ER, Kim M. , et al. Platelet Toll-like receptor expression modulates lipopolysaccharide-induced thrombocytopenia and tumor necrosis factor-alpha production in vivo. Blood 2006; 107 (02) 637-641
    CrossrefPubMedGoogle Scholar
  • 11 Damien P, Cognasse F, Payrastre B. , et al. NF-κB links TLR2 and PAR1 to soluble immunomodulator factor secretion in human platelets. Front Immunol 2017; 8: 85
    PubMedGoogle Scholar
  • 12 Ståhl AL, Svensson M, Mörgelin M. , et al. Lipopolysaccharide from enterohemorrhagic Escherichia coli binds to platelets through TLR4 and CD62 and is detected on circulating platelets in patients with hemolytic uremic syndrome. Blood 2006; 108 (01) 167-176
    CrossrefPubMedGoogle Scholar
  • 13 Clark SR, Ma AC, Tavener SA. , et al. Platelet TLR4 activates neutrophil extracellular traps to ensnare bacteria in septic blood. Nat Med 2007; 13 (04) 463-469
    CrossrefPubMedGoogle Scholar
  • 14 Claushuis TAM, Van Der Veen AIP, Horn J. , et al. Platelet Toll-like receptor expression and activation induced by lipopolysaccharide and sepsis. Platelets 2018; :1–9 DOI: 10.1080/09537104.2018.1445841. [Epub ahead of print]
    CrossrefPubMedGoogle Scholar
  • 15 Rivadeneyra L, Carestia A, Etulain J. , et al. Regulation of platelet responses triggered by Toll-like receptor 2 and 4 ligands is another non-genomic role of nuclear factor-kappaB. Thromb Res 2014; 133 (02) 235-243
    CrossrefPubMedGoogle Scholar
  • 16 Ward JR, Bingle L, Judge HM. , et al. Agonists of toll-like receptor (TLR)2 and TLR4 are unable to modulate platelet activation by adenosine diphosphate and platelet activating factor. Thromb Haemost 2005; 94 (04) 831-838
    Article in Thieme ConnectPubMedGoogle Scholar
  • 17 Klarström Engström K, Brommesson C, Kälvegren H, Bengtsson T. Toll like receptor 2/1 mediated platelet adhesion and activation on bacterial mimetic surfaces is dependent on src/Syk-signaling and purinergic receptor P2 × 1 and P2Y12 activation. Biointerphases 2014; 9 (04) 041003
    CrossrefPubMedGoogle Scholar
  • 18 Berthet J, Damien P, Hamzeh-Cognasse H. , et al. Human platelets can discriminate between various bacterial LPS isoforms via TLR4 signaling and differential cytokine secretion. Clin Immunol 2012; 145 (03) 189-200
    CrossrefPubMedGoogle Scholar
  • 19 McCabe White M, Jennings LK. Platelet Protocols: Research and Clinical Laboratory Procedures. San Diego, CA: Academic Press; 1999
    Google Scholar
  • 20 Flo TH, Halaas O, Lien E. , et al. Human toll-like receptor 2 mediates monocyte activation by Listeria monocytogenes, but not by group B streptococci or lipopolysaccharide. J Immunol 2000; 164 (04) 2064-2069
    CrossrefPubMedGoogle Scholar
  • 21 Tsukamoto H, Ukai I, Yamagata Y. , et al. Leucine-rich repeat 2 of human Toll-like receptor 4 contains the binding site for inhibitory monoclonal antibodies. FEBS Lett 2015; 589 (24, Pt B): 3893-3898
    CrossrefPubMedGoogle Scholar
  • 22 Lievens D, Zernecke A, Seijkens T. , et al. Platelet CD40L mediates thrombotic and inflammatory processes in atherosclerosis. Blood 2010; 116 (20) 4317-4327
    CrossrefPubMedGoogle Scholar
  • 23 Kato H, Adachi S, Doi T. , et al. Mechanism of collagen-induced release of 5-HT, PDGF-AB and sCD40L from human platelets: role of HSP27 phosphorylation via p44/p42 MAPK. Thromb Res 2010; 126 (01) 39-43
    CrossrefPubMedGoogle Scholar
  • 24 Brandt E, Petersen F, Ludwig A, Ehlert JE, Bock L, Flad HD. The beta-thromboglobulins and platelet factor 4: blood platelet-derived CXC chemokines with divergent roles in early neutrophil regulation. J Leukoc Biol 2000; 67 (04) 471-478
    CrossrefPubMedGoogle Scholar
  • 25 von Hundelshausen P, Koenen RR, Sack M. , et al. Heterophilic interactions of platelet factor 4 and RANTES promote monocyte arrest on endothelium. Blood 2005; 105 (03) 924-930
    CrossrefPubMedGoogle Scholar
  • 26 Fleischer J, Grage-Griebenow E, Kasper B. , et al. Platelet factor 4 inhibits proliferation and cytokine release of activated human T cells. J Immunol 2002; 169 (02) 770-777
    CrossrefPubMedGoogle Scholar
  • 27 Brown AJ, Sepuru KM, Sawant KV, Rajarathnam K. Platelet-derived chemokine CXCL7 dimer preferentially exists in the glycosaminoglycan-bound form: implications for neutrophil-platelet crosstalk. Front Immunol 2017; 8: 1248
    CrossrefPubMedGoogle Scholar
  • 28 Antczak AJ, Singh N, Gay SR, Worth RG. IgG-complex stimulated platelets: a source of sCD40L and RANTES in initiation of inflammatory cascade. Cell Immunol 2010; 263 (01) 129-133
    CrossrefPubMedGoogle Scholar
  • 29 Anselmo A, Riva F, Gentile S. , et al. Expression and function of IL-1R8 (TIR8/SIGIRR): a regulatory member of the IL-1 receptor family in platelets. Cardiovasc Res 2016; 111 (04) 373-384
    CrossrefPubMedGoogle Scholar
  • 30 Tsai JC, Lin YW, Huang CY. , et al. The role of calpain-myosin 9-Rab7b pathway in mediating the expression of Toll-like receptor 4 in platelets: a novel mechanism involved in α-granules trafficking. PLoS One 2014; 9 (01) e85833
    CrossrefPubMedGoogle Scholar
  • 31 Duerschmied D, Bode C, Ahrens I. Immune functions of platelets. Thromb Haemost 2014; 112 (04) 678-691
    Article in Thieme ConnectPubMedGoogle Scholar
  • 32 Semple JW, Italiano Jr JE, Freedman J. Platelets and the immune continuum. Nat Rev Immunol 2011; 11 (04) 264-274
    CrossrefPubMedGoogle Scholar
  • 33 Aloui C, Prigent A, Sut C. , et al. The signaling role of CD40 ligand in platelet biology and in platelet component transfusion. Int J Mol Sci 2014; 15 (12) 22342-22364
    CrossrefPubMedGoogle Scholar
  • 34 Gerskowitch VP, Hodge J, Hull RA. , et al. Unexpected relationship between plasma protein binding and the pharmacodynamics of 2-NAP, a CCK1-receptor antagonist. Br J Clin Pharmacol 2007; 63 (05) 618-622
    CrossrefPubMedGoogle Scholar
  • 35 Koessler J, Hermann S, Weber K. , et al. Role of purinergic receptor expression and function for reduced responsiveness to adenosine diphosphate in washed human platelets. PLoS One 2016; 11 (01) e0147370
    CrossrefPubMedGoogle Scholar
  • 36 Koessler J, Weber K, Koessler A, Yilmaz P, Boeck M, Kobsar A. Expression and function of purinergic receptors in platelets from apheresis-derived platelet concentrates. Blood Transfus 2016; 14 (06) 545-551
    PubMedGoogle Scholar
  • 37 Jonnalagadda D, Izu LT, Whiteheart SW. Platelet secretion is kinetically heterogeneous in an agonist-responsive manner. Blood 2012; 120 (26) 5209-5216
    CrossrefPubMedGoogle Scholar
  • 38 Kaufman J, Spinelli SL, Schultz E, Blumberg N, Phipps RP. Release of biologically active CD154 during collection and storage of platelet concentrates prepared for transfusion. J Thromb Haemost 2007; 5 (04) 788-796
    CrossrefPubMedGoogle Scholar