To Örebro University

Platelets are considered to have important functions in inflammatory processes as key players in innate immunity. Toll like receptors (TLRs), expressed on platelets, recognize pathogen associated molecular patterns and trigger immune responses. Pathogens are able to adhere to human tissues and form biofilms which cause a continuous activation of the immune system. The authors aimed to investigate how immobilized Pam(3)CSK(4) (a synthetic TLR2/1 agonist) and IgG, respectively, resembling a bacterial focus, affects adhesion and activation of platelets including release of two cytokines, regulated on activation normal T-cell expressed and secreted (RANTES) and macrophage migration inhibitory factor (MIF). The authors also aim to clarify the signaling downstream of TLR2/1 and Fc gamma RII (IgG receptor) and the role of adenine nucleotides in this process. Biolayers of Pam(3)CSK(4) and IgG, respectively, were confirmed by null-ellipsometry and contact angle measurements. Platelets were preincubated with signaling inhibitors for scr and Syk and antagonists for P2X1 or P2Y1 [adenosine triphosphate (ATP), adenosine diphosphate (ADP) receptors] prior to addition to the surfaces. The authors show that platelets adhere and spread on both Pam(3)CSK(4)- and IgG-coated surfaces and that this process is antagonized by scr and Syc inhibitors as well as P2X1 and P2Y antagonists. This suggests that Pam(3)CSK(4) activated platelets utilize the same pathway as Fc gamma RII. Moreover, the authors show that ATP-ligation of P2X1 is of importance for further platelet activation after TLR2/1-activation, and that P2Y12 is the prominent ADP-receptor involved in adhesion and spreading. RANTES and MIF were secreted over time from platelets adhering to the coated surfaces, but no MIF was released upon stimulation with soluble Pam(3)CSK(4). These results clarify the importance of TLR2/1 and Fc gamma RII in platelet adhesion and activation, and strengthen the role of platelets as an active player in sensing bacterial infections.