CONDITIONS INFLUENCING RELEASE OF GRANULE CONTENTS FROM HUMAN PLATELETS IN CITRATED PLASMA-INDUCED BY ADP OR THE THROMBIN RECEPTOR ACTIVATING PEPTIDE SFLLRN - DIRECT MEASUREMENT OF PERCENT RELEASE OF BETA-THROMBOGLOBULIN AND ASSESSMENT BY FLOW-CYTOMETRY OF P-SELECTIN EXPRESSION

Contrary to a recent report [Rinder et al.: Blood 82:505, 1993], aspir in does inhibit the release of cu-granule contents as well as inhibiti ng the release of dense granule contents by human platelets during ADP -induced aggregation in citrated platelet-rich plasma (PRP). Measureme nts were: percent release of C-14-serotonin from prelabeled platelets, radioimmunoassay of beta-thromboglobulin (beta TG), and expression on the platelet surface of the alpha-granule constituent, P-selectin, by flow cytometry. During the second phase of ADP-induced aggregation, 6 9.0+/-8.3% of beta TG and 54.1+/-4.6% of C-14-serotonin were released (means+/-SEM, n=13); aspirin treatment reduced these values to 6.0+/-1 .2 and 1.0+/-0.3%, respectively. In contrast, incubation of platelets with ADP without stirring caused only 6.7+/-1.7% release of beta TG an d 2.1+/-0.4% release of C-14-serotonin; these low values were not appr eciably affected by aspirin. During ADP-induced primary aggregation in PRP anticoagulated with FPRCH(2)Cl (PPACK), only 4.7+/-0.9% release o f beta TG and no detectable release of C-14-serotonin occurred; aspiri n had no effect. In both stirred and unstirred PRP, the thrombin recep tor activating peptide, SFLLRN (50 mu M), caused at least 75% release of the contents of both granules, which was partially inhibited by asp irin. Upon incubation of platelets with ADP (2-10 mu M), the mean fluo rescence intensity due to P-selectin was <14% of that induced by SFLLR N. In this unstirred system used for flow cytometry, aspirin treatment caused no significant inhibition of P-selectin expression, Thus, unde r conditions in which ADP does not cause secondary aggregation (physio logical Ca2+ concentration or unstirred citrated PRP) release of the c ontents of both types of granules is less than 7% and aspirin is not i nhibitory; the P-selectin expression associated with this low percent release is also unaffected by aspirin. However, aspirin does strongly inhibit the extensive release of both alpha-granule and dense granule contents during ADP-induced secondary aggregation in citrated PRP. (C) 1996 Wiley-Liss, Inc.