PLATELET MICROPARTICLES BIND, ACTIVATE AND AGGREGATE NEUTROPHILS IN-VITRO

The interaction of activated platelets with leukocytes are believed to play an important role in ischemic reperfusion injury and other throm botic conditions. Upon activation, platelets shed platelet micropartic les (PMP) and express activation markers, CD62P expressed on activated platelets mediates adhesion of platelets to leukocytes, chiefly neutr ophils, but little is known of the interaction of PMP with neutrophils . We investigated this interaction as compared to platelet/neutrophil interaction, PMP isolated from stored platelets or thrombin activated platelets was incubated with leukocytes and binding assessed by flow c ytometry. FITC-labeled alpha-CD41 was used to assess platelet material associated with WBC. Like platelets, PMP bound preferentially to neut rophils rather than lymphocytes, and exhibited an absolute dependence on the presence of Ca2+. Binding was time- and concentration-dependent , reaching a plateau at 10 min at a ratio of PMP to neutrophils of 150 :1. Fluorescence microscopy showed that most of the neutrophils were a ggregated into clusters of 5-20 cells, Clustering of neutrophils was n ot observed to result from interaction with platelets. In these cluste rs the adherent PMP appeared to serve as bridges between the neutrophi ls. Addition of EGTA after brief incubation (5-10 min) released most o f the bound PMP but if added after >10 min, only similar to 60% of bou nd PMP were released, In contrast, nearly all bound platelets were rel eased by EGTA at the same time of incubation, Incubation of neutrophil s with PMP gave a significantly higher percentage of CD41a(+) neutroph ils than did platelets incubated at the same numerical ratio, PMP asso ciation with neutrophils was less markedly inhibited by alpha-CD62P (A C1.2) than platelets, but binding of both PMP and activated platelets was inhibited similar to 90% by antisialyl Lewis X. PMP binding to neu trophils induced a significant increase in both CD11b expression and p hagocytic activity in a concentration-dependent manner. These findings suggest a possible role for PMP in addition to providing platelet fac tor 3, specifically, as an activator and mediator of neutrophils in is chemic injury, thrombosis, and inflammation.