Adhesion of human platelets to albumin is synergistically increased by lysophosphatidic acid and adrenaline in a donor-dependent fashion

Abstract

Lysophosphatidic acid (LPA) and adrenaline are weak platelet activators considered important for thrombus formation, and were previously shown to synergistically increase platelet aggregation. Here we investigate synergistic activation by LPA and adrenaline when measuring platelet adhesion. Platelet-rich plasma from healthy blood donors together with adrenaline and/or LPA were added to protein-coated microplates. Platelets were allowed to adhere and the amount of adhesion detected enzymatically. The LPA and adrenaline combination induced a synergistic increase of platelet adhesion to a normally non-adhesive albumin surface. The degree of synergy varied markedly between individuals; these variations could not be explained by age, gender, blood type or different amounts of platelets, oxidized low-density lipoprotein, insulin or glucose in plasma. There was a trend indicating increased synergistic effect for platelets sensitive to adrenaline stimulation. The synergistic effect was blocked by the alpha2-adrenoceptor antagonist yohimbine and inhibited by the ADP scavenger system creatine phosphate/creatine phosphokinase and antibodies against alphaIIbbeta3. Furthermore, platelets adhering to albumin after adrenaline and LPA treatment expressed P-selectin. In conclusion, LPA and adrenaline act synergistically to increase alphaIIbbeta3-mediated platelet adhesion to albumin, dependent on alpha2-adrenoceptor signalling and platelet secretion. We also confirm that synergistic platelet activation achieved with LPA and adrenaline is highly donor dependent.

Characterization of static adhesion of human platelets in plasma to protein surfaces in microplates

“…2A) or by AK2 directed against GPIb-IX-V (not shown). We have earlier reported adhesion of activated platelets to albumin [15,16], and confirm these findings in this study by using ADP (Fig. 3A), as well as adrenaline alone or in combination with ristocetin ( The effects of 0.1 µmol/L cangrelor or 10 µmol/L BM-531 on platelet adhesion to albumin, collagen and fibrinogen were investigated in the absence (n=6 for cangrelor experiments.…”

“…This assay is simple to use and, since it is performed in 96-well microplates, it is well suited for screening purposes in order to measure several aspects of platelet function simultaneously. The assay has earlier been used for investigation of platelet activity both in basic studies [16,17] and in clinical research [18,19]. The aim of this study was to investigate fundamental adhesion events occurring in this particular assay.…”

Enhanced platelet adhesion in essential thrombocythemia after in vitro activation

“…The appearance of individual variability in the response to adrenaline has been studied in the healthy population, and when including 140 individuals, 16% were classified as non-responders to adrenaline-induced platelet aggregation in plasma [33]. This variability in the adrenaline response in healthy subjects has been confirmed by other investigations also studying platelet function in plasma [20,[34][35][36]. No study has been designed with the purpose of investigating individual variability of adrenaline-response in ET patients.…”

“…In contrast, weaker stimuli represented by adhesion measured on albumin resulted in comparatively few adhesion values that were higher than the control mean + 2SD. In addition, earlier studies show that secretion of ADP is important in this assay [20,21]. Thus, we propose that multiple stimuli, which most likely are of importance in vivo [42], and a long incubation time are both needed to induce secretion and activation of exhausted ET-platelets.…”

“…The inclusion of bacteria-derived ristocetin is motivated since it stimulates the otherwise flow-dependent interaction between von Willebrand factor (vWf) and glycoprotein (GP)-Ib-IX-V on platelets [19]. Albumin was included both as a negative control for basal adhesion and because it allows the detection of LPA-induced adhesion [20]. This experimental setup also allows investigation of both α 2 β 1 -dependent adhesion to collagen and α IIb β 3 -dependent adhesion to fibrinogen and albumin [21].…”