The dynamics of interaction between hemostatic cells (platelets) and inflammatory cells (PMN) provides a basis for understanding relevant pathophysiologies. Platelets may aid PMN participation at an inflammatory site and PMN may function in thromboregulation. Data in vitro on platelet-PMN interactions suggest that these cell interactions are of importance for both inflammation and hemostasis. We are investigating platelet-PMN interactions and the functional consequences of platelet-PMN adhesion. Adhesion of activated platelets to PMN is mediated by P-selectin (CD62) on platelets binding to P-selectin glycoprotein ligand-1 on PMN in the presence of calcium ions. Adhesion of unactivated platelets to PMN is mediated through an as yet unknown receptor-ligand interaction independent of divalent cations. PMN adhesion molecules are differentially down-regulated after migration in vitro. Hence, we have investigated the adhesion of unactivated platelets to chemotactically responsive (migrated) and non-responsive (non-migrated) PMN to examine their possible receptor-ligand interactions. Autologous platelets and PMN were isolated from whole blood. Platelets were labeled with a fluorescein isothiocyanate-conjugated CD41 (GPIIb-IIIa complex) mono- clonal antibody. Migrated and non-migrated PMN were separated after PMN chemotaxis to N-formyl peptide on polycarbonate membranes. PMN were labeled with phycoerythrin-conjugated CD45 monoclonal antibody. Platelets (300 million/ml) and PMN (3 million/ml) were coincubated for 30 min. Heterotypic cell adhesion was measured by flow cytometric analysis of platelet marker fluorescence in PMN size-gated events. In platelets coincubated with migrated and non-migrated PMN, 38-78% of PMN bound one or two platelets. In isolated PMN, contaminating platelets were bound to 10-25% of unstimulated PMN, 7-19% of stimulated PMN, 15-24% of non-migrated PMN, and 1-3% of migrated PMN. Corroborative evidence of PMN-platelet adhesion was obtained by fluorescence confocal microscopy. These results indicate that platelets adhered to PMN are detached in the process of PMN migration.
