PLATELET POLYMORPHONUCLEAR LEUKOCYTE INTERACTION IN DYNAMIC CONDITIONS - EVIDENCE OF ADHESION CASCADE AND CROSS-TALK BETWEEN P-SELECTIN ANDTHE BETA-2 INTEGRIN CD11B/CD18/

Adhesion between platelets and polymorphonuclear leukocytes (PMN) is a key event in thrombosis and inflammation. Double color fluorescence-a ctivated cell sorter (FACS) analysis was used to determine the extent and kinetics of adhesion of thrombin-activated platelets to resting or activated PMN when mixed cell populations were incubated in dynamic c onditions. Activated platelets bound very rapidly to PMN. Mixed cell c onjugates reached a maximum at 1 minute and were reversible within 10 minutes. Platelet/PMN adhesion required both Ca2+ and Mg2+ and was mar kedly increased by the presence of Mn2+. The latter made mixed cell co njugates stable up to 10 minutes. Adhesion of platelets required metab olic activity of PMN and was abolished by tyrosine kinase inhibitors. Furthermore, adhesion of platelets to PMN resulted in binding of a mon oclonal antibody (MoAb 24) known as beta 2 integrins ''activation repo rter.'' When PMN were activated by exogenous stimuli, the adhesion of platelets was markedly increased: fMLP induced a rapid and transient e ffect, while PMA resulted in a slower, but stable, increase in mixed c onjugates formation. The hypothesis that activated PMN beta 2 integrin s are able to bind a counter-receptor on platelets was directly demons trated by the increase of mixed cell conjugates following PMN treatmen t with KIM127 and KIM185, two anti-CD18 antibodies able to induce the active conformation of beta 2 integrins. Consistently, two other anti- CD18, as well as an anti-CD11b inhibitory antibody abolished platelet/ PMN adhesion. PMN beta 2 integrin activation was not the only mechanis m for activated platelet/PMN adhesion to occur: indeed,this phenomenon could also be inhibited by two anti-P-selectin antibodies. Resting pl atelets did not adhere to resting PMN, but markedly adhered to fMLP- o r PMA-activated PMN. Resting platelet/fMLP-activated PMN adhesion was abolished by anti-CD18 antibodies, but not by anti-P-selectin antibodi es. In conclusion, activated platelet/PMN interaction can be modeled a s an adhesion cascade involving a P-selectin-dependent recognition ste p and a functional signal. The latter proceeds through tyrosine kinase activation and enables a beta 2 integrin-dependent adhesion to a not yet identified counter-receptor constitutively expressed on platelet s urface. (C) 1996 by The American Society of Hematology.