To determine how these markers are distributed on the surface of platelets/particles, we studied Annexin V and PAC-1 binding to platelets/particles of different sizes by flow cytometry analysis and evaluated influences of calpain and caspase inhibitors on thrombin/convulxin-activated platelets.
Analysed platelets/particles were divided by their sizes, according to the standard size beads, into seven populations from 0.37 to 4.8 microm. PAC-1 binding/microm(2) was almost equal in platelets/particles ranging from 1.2 to 4.8 microm and was significantly lower on smaller-sized particles sizes (0.37-0.7 microm). PS surface exposure/microm(2) was high in the particles of 0.37-1.2 microm and very low in platelets (2.6-4.8 microm). Upon thrombin/convulxin stimulation caspase inhibitors prevented microparticle (MP) formation, while a calpain inhibitor stimulated MP formation.
It was also shown that stimulated platelets are heterogeneous not only in their ability to activate alpha IIb beta 3 integrin complex and expose PS on their surface, but also in the distribution of activation markers, which strongly depends on platelet/particle size and that platelets/particles of different sizes provide different responses to the same stimulus.