The integrin alpha IIb/beta 3 in human platelet signal transduction

Platelets are critical for the maintenance of the integrity of the vascular system and are the first line of defence against haemorrhage. When they encounter a subendothelial matrix exposed by injury to a vessel, platelets adhere, are activated, and become adhesive for other platelets so that they aggregate. alpha IIb/beta 3, a platelet-specific integrin, is largely prominent amongst the adhesion receptors and is essential for platelet aggregation. The ligands for alpha IIb/beta 3 are the multivalent adhesive proteins fibrinogen and von Willebrand factor. In resting platelets, alpha IIb/beta 3 is normally in a low activation state, unable to interact with soluble fibrinogen. Stimulation of platelets with various agonists will induce a conformational change in alpha IIb/beta 3 (inside-out signalling), which is then able to bind soluble fibrinogen resulting in the onset of platelet aggregation. However, fibrinogen binding to its membrane receptor is not simply a passive event allowing the formation of intercellular bridges between platelets. Indeed, a complex signalling pathway triggered by integrin ligation and clustering (outside-in signalling) will regulate the extent of irreversible platelet aggregation and clot retraction. Amongst the signalling enzymes activated downstream of alpha IIb/beta 3 engagement, phosphoinositide 3-kinase plays an important role in the control of the irreversible phase of aggregation.     

YD-3, a novel inhibitor of protease-induced platelet activation

1. In the present study, the antiplatelet effects and mechanisms of a new synthetic compound YD-3 [1-benzyl-3(ethoxycarbonylphenyl)-indazole] were examined. 2. YD-3 inhibited the aggregation of washed rabbit platelets caused by thrombin (IC(50)=28.3 microM), but had no or little inhibitory effect on that induced by arachidonic acid, collagen, platelet-activating factor (PAF) or U46619. YD-3 also suppressed generation of inositol phosphates caused by thrombin. On the other hand, thrombin-induced fibrin formation was not affected by YD-3, indicating YD-3 does not inhibit the proteolytic activity of thrombin. 3. In washed human platelets, however, YD-3 had only mild inhibitory effect on the low concentration (0.05 u ml(-1)) of thrombin-induced human platelet aggregation, and did not affect that induced by higher concentrations (> or =0.1 u ml(-1)) of thrombin or SFLLRN, the protease-activated receptor 1 (PAR1) agonist peptide. By contrast, YD-3 inhibited both human and rabbit platelet aggregation elicited by trypsin with IC(50) values of 38.1 microM and 5.7 microM, respectively. 4. YD-3, at 100 microM, had no effect on ristocetin-induced glycoprotein Ib (GPIb)-dependent aggregation of human platelets. In addition, platelets treated with chymotrypsin, which cleaves GPIb, enhanced rather than attenuated the inhibition of YD-3 on thrombin-induced human platelet aggregation. These data indicate that GPIb plays no role in the antiplatelet effect of YD-3. 5. In SFLLRN-desensitized human platelets, high concentration of thrombin (1 u ml(-1)) could still elicit intracellular Ca(2+) mobilization, and the rise of [Ca(2+)](i) was prevented by either leupeptin or YD-3. 6. Our results suggest that YD-3 inhibits a non-PAR1 thrombin receptor which mediates the major effect of thrombin in rabbit platelets, but in human platelets, this receptor function becomes significant only when the function of PAR1 has been blocked or attenuated.     

Inhibition of platelet adhesion to collagen as a new target for antithrombotic drugs

Platelet adhesion to a damaged blood vessel is the initial trigger for arterial hemostasis and thrombosis. Platelets adhere to the subendothelium through an interaction with von Willebrand factor (VWF), which forms a bridge between collagen within the damaged vessel wall and the platelet receptor glycoprotein Ib/V/IX (GPIb), an interaction especially important under high shear conditions[1]. This reversible adhesion allows platelets to roll over the damaged area, which is then followed by a firm adhesion mediated by the collagen receptors (alpha(2)beta(1), GPVI, ) in addition[2] resulting in platelet activation. This leads to the conformational activation of the platelet alpha(IIb)beta3 receptor, fibrinogen binding and finally to platelet aggregation. Over the past decades, modulation of platelet function has been a strategy for the control of cardiovascular disease. Lately, drugs have been developed that target the fibrinogen receptor alphaIIbbeta3 or the ADP receptor and many of these promising compounds have been tested in clinical trials. However the development of products that interfere with the first step of hemostasis, i.e. the platelet adhesion, has lagged behind. In this review we want to discuss (i) the in vivo studies that were performed with compounds that target proteins involved in different adhesion steps i.e. the VWF-GPIb-axis, the collagen-VWF axis and the collagen-collagen receptor axis and (ii) the possible advantages these putative new drugs could have over the current antiplatelet agents.     

眼镜蛇毒蛋白酶natrahagin水解纤维蛋白原的特性及对人血小板聚集的

AIM: To study the fibrinogenolytic properties of natrahagin and its effect on platelet aggregation. METHOD: SDS-PAGE, fibrinogenolytic activity assay, platelet aggregation. RESULTS: Upon incubation of fibrinogen with natrahagin at the ratio of 50:1 (w/w), A alpha-chains of fibrinogen were almost completely hydrolyzed in 5 min; however, at least 6 h was needed for the complete degradation of gamma-chains. Fibrinogenolytic activity of natrahagin was 0.349 +/- 0.044 g.min-1.g-1 as determined by its ability to reduce the clottable fibrinogen. On the other hand, natrahagin concentration-dependently inhibited platelet aggregation induced by ristocetin in platelet-rich plasma and thrombin (80 U.L-1) in washed platelets with IC50 (95% confidence limit) of 56 (40-79) and 3.3 (1.4-8.0) mg.L-1. No inhibitory effect was found on collagen- and ADP-induced platelet aggregation even when the dose of natrahagin reached 200 mg.L-1. CONCLUSION: Natrahagin is an alpha, gamma-fibrinogenase with an inhibitory effect on platelet membrane glycoprotein Ib (GPIb)-dependent platelet aggregation.     

The Role of the Platelet in the Pathogenesis of Atherothrombosis

Platelet adhesion, activation, and aggregation at sites of vascular endothelial disruption caused by atherosclerosis are key events in arterial thrombus formation. Platelet tethering and adhesion to the arterial wall, particularly under high shear forces, are achieved through multiple high-affinity interactions between platelet membrane receptors (integrins) and ligands within the exposed subendothelium, most notably collagen and von Willebrand factor (vWF). Platelet adhesion to collagen occurs both indirectly, via binding of the platelet glycoprotein (GP) Ib-V-IX receptor to circulating vWF, which binds to exposed collagen, and directly, via interaction with the platelet receptors GP VI and GP Ia/IIb. Platelet activation, initiated by exposed collagen and locally generated soluble platelet agonists (primarily thrombin, ADP, and thromboxane A2), provides the stimulus for the release of platelet-derived growth factors, adhesion molecules and coagulation factors, activation of adjacent platelets, and conformational changes in the platelet alpha(IIb)beta3 integrin (GP IIb/IIIa receptor). Platelet aggregation, mediated primarily by interaction between the activated platelet GP IIb/IIIa receptor and its ligands, fibrinogen and vWF, results in the formation of a platelet-rich thrombus. Currently available antiplatelet drugs (aspirin [acetylsalicylic acid], dipyridamole, clopidogrel, ticlopidine, abciximab, eptifibatide, tirofiban) act on specific targets to inhibit platelet activation and aggregation. Elucidation of the multiple mechanisms involved in platelet thrombus formation provides opportunities for selectively inhibiting the pathways most relevant to the pathophysiology of atherothrombosis.     

Antiplatelet Mechanism of 2‐Chloro‐3‐(4‐hexylphenyl)‐amino‐1,4‐naphthoquinone (NQ304), an Antithrombotic Agent

Abstract: The effects of 2‐chloro‐3‐(4‐hexylphenyl)‐amino‐1,4‐naphthoquinone (NQ304), an antithrombotic agent, on aggregation, binding of fibrinogen to glycoprotein IIb/IIIa and intracellular signals were investigated using human platelets. NQ304 inhibited thrombin‐, arachidonic acid‐ and thapsigargin‐induced aggregation of washed human platelets with the IC₅₀ values of 22.2±0.7, 6.5±0.2, and 7.6±0.1 μM, respectively. NQ304 significantly inhibited fluorescein isothiocyanate‐conjugated fibrinogen binding to human platelet surface glycoprotein IIb/IIIa receptor by 75%, but failed to inhibit the fibrinogen binding to purified glycoprotein IIb/IIIa receptor. This result suggests that NQ304 inhibit platelet aggregation by suppression of an intracellular pathway that involves exposure of the glycoprotein IIb/IIIa receptor, rather than by direct inhibition of fibrinogen‐glycoprotein IIb/IIIa binding. NQ304 significantly inhibited thrombin‐induced increase in intracellular Ca²⁺ mobilization at the dose of 30 μM and ATP secretion in a dose‐dependent manner. It also inhibited thrombin‐ and arachidonic acid‐induced thromboxane A₂ formation in human platelet dose‐dependently. In conclusion, the antiplatelet mechanism of NQ304 may be due to the reduction of the thromboxane A₂ formation, inhibition of adenosine triphosphate release and intracellular calcium mobilization.     

The antithrombotic efficacy of lotrafiban (SB 214857) in canine models of acute coronary thrombosis

In patients with acute coronary syndromes, inhibition of platelet aggregation with parenteral alpha(IIb)/beta(III) antagonists has proven effective at preventing nonfatal myocardial infarction and repeat percutaneous coronary interventions. Paradoxically, the efficacy observed for acute indications and parenteral agents has not extended to oral agents and chronic prevention of secondary thrombotic events, despite robust antithrombotic properties in preclinical thrombosis models. This report documents the preclinical data of Lotrafiban, an oral alpha(IIb)/beta(III) antagonist that recently failed in a phase III clinical trial (BRAVO) for the prevention of secondary thrombosis. Lotrafiban was characterized in a dog circumflex artery electrical injury model, and a cyclic flow reduction model ("Folts"). The data demonstrate that both oral (1.0-50.0 mg/kg) and intravenous (0.1-0.8ug/kg/min) administration of lotrafiban produced dose-related inhibition (45%-95%) of ex vivo platelet aggregation. In the electrical injury model, the dose-related inhibition correlated with a significant reduction in the frequency of coronary occlusion, size of the developing thrombus, and the extent of left ventricular ischemic damage. Effects on blood flow and bleeding time were also dose related. The combination of low dose lotrafiban (0.1ug/kg/min) and aspirin (5.0 mg/kg) generated additive antithrombotic effects, approximating the antithrombotic efficacy of a 2-4 fold higher dose of lotrafiban while only modestly prolonging the bleeding time. For purposes of comparison, the ADP receptor antagonist clopidogrel was also assessed in the electrical injury model. Clopidogrel (5.0-10.0 mg/kg, iv.) significantly reduced the resulting left ventricular infarct areas, but lacked the overall efficacy of lotrafiban. In the "Folts" model, lotrafiban inhibited cyclic blood flow reductions (CFR's) by 100% in animals insensitive to the antithrombotic effects of aspirin. Overall, the preclinical data demonstrated that alpha(IIb)/beta(III) antagonism with lotrafiban was a well tolerated and effective strategy for attenuating acute arterial thrombosis. The lack of a correlation between these preclinical data and the outcome of the clinical trial BRAVO are unexplained. However, the combined evidence suggests that these acute canine thrombosis studies may not completely capture the pathology reflected in chronic human atherothrombotic disease.     

Surfactin C inhibits platelet aggregation

This study was designed to investigate the effect of surfactin C, which is derived from Bacillus subtilis, on platelet aggregation and homotypic leucocyte aggregation. Surfactin C strongly and dose-dependently inhibited platelet aggregation, which was stimulated both by thrombin (0.1 U mL(-1)), a potent agonist that activates the G protein-coupled protease receptor, and by collagen (5 microg mL(-1)), a potent ligand that activates alpha(IIb)beta(3) with IC50 values (concentration inhibiting platelet aggregation by 50%) of 10.9 and 17.0 microM, respectively. Moreover, surfactin C significantly suppressed the intracellular Ca(2+) mobilization in thrombin-activated platelets. Surfactin C, however, did not affect various integrin-mediated U937 cell aggregation, implying that the anti-platelet activity of surfactin C was not due to its detergent effect but by its action on the downstream signalling pathway. Therefore, the results suggest that surfactin C may have a beneficial therapeutic effect on aberrant platelet aggregation-mediated cardiovascular diseases.     

Effect of cefaclor on guinea pig platelet aggregation in vitro

Effects of cefaclor (3-chloro-7-D-(2-phenyl-glycinamido)-3-cephem-4-carboxylic acid) on PAF, ADP, collagen, endotoxin, and thrombin-induced platelet aggregation were examined in vitro with the use of guinea pig platelet-rich plasma and washed platelets. PAF, even at concentrations lower than its minimum effective concentration, enhanced ADP- or endotoxin-induced platelet aggregation and prolonged the time to attain the maximum aggregation. PAF also enhanced collagen-induced platelet aggregation and shortened the lag time. Cefaclor (CCL) inhibited the PAF, ADP or thrombin induced platelet aggregation and shortened their maximum aggregation times at higher concentrations such as 300 micrograms/ml or more. CCL also inhibited the collagen-induced platelet aggregation and prolonged the lag time, but showed no effect on endotoxin-induced platelet aggregation. The effect of CCL was almost the same as that of latamoxef (LMOX). CCL and LMOX, however, showed no effect on cellular Ca2+ increase produced by PAF, ADP, or thrombin, suggesting that the inhibitory effect of CCL and LMOX on platelet aggregation is caused by the inhibition of fibrinogen binding to the glycoprotein IIb/IIIa complex.     

TMVA, a snake C-type lectin-like protein from Trimeresurus mucrosquamatus venom, activates platelet via GPIb.

TMVA is a C-type lectin-like protein with potent platelet activating activity from Trimeresurus mucrosquamatus venom. In the absence of von Willebrand factor (vWF), TMVA dose-dependently induced aggregation of washed platelets. Anti-GP Ib monoclonal antibodies (mAbs), HIP1, specifically inhibited TMVA-induced aggregation in a dose-dependent manner. The aggregation was also inhibited by mAb P2 (an anti-GP IIb mAb). Flow cytometric analysis revealed that FITC-TMVA bound to human formalin-fixed platelets in a saturable manner, and its binding was specifically blocked by HIP1 in a dose-dependent manner. Flow cytometric analysis showed that TMVA did not bind to platelet GPIX, GPIIb, GPIIIa, GPIa, GPIIa and GPIV. Moreover, the platelet aggregation induced by TMVA was partially inhibited when platelet was pretreated with mocarhagin, a snake venom protease that specifically cleaves human GPIb. These results suggest that TMVA is a strong platelet agonist via GPIb and it might have multiple functional binding-sites on GPIb molecule or on other unknown receptor.     

Pharmacological analysis of integrin function: inhibition of vascular stenosis and identification of a novel antiplatelet substance

The long-lasting success of PTCA, a common treatment for patients with coronary artery disease, is hampered by acute reocclusion due to thrombus and by late restenosis due to smooth muscle cell (SMC) migration and proliferation. Recently potent antiplatelet agents such as inhibitors of platelet glycoprotein (GP) IIb/IIIa (alpha IIb/beta 3) have become available and are highly effective in humans. We investigated the inhibitory effects of these compounds on vascular stenosis. Administration of an RGD-containing peptide (non-selective alpha IIb/beta 3 antagonist) results in reduction of neointima, but selective alpha IIb/beta 3 antagonists have no effect. This effect is due both to an early event, which could be due to the inhibition of secretion of PDGF from activated platelets with blocked alpha IIb/beta 3, and to a late event, by interference with SMC alpha v/beta 3. Moreover, platelet adhesion via GPIb/V/IX is a trigger for thrombus formation. Inhibition of platelet adhesion results in the prevention of thrombus formation and the suppression of neointima. However, this effect was supported by the reduction of SMC proliferation. Therefore, these dual inhibitions markedly reduced vascular stenosis. Finally some low molecular mass heat shock proteins (HSPs) appear to act as molecular chaperones, but their physiological roles have not been fully elucidated. We have investigated the physiological role of p20, a kind of HSP, on platelet function. p20 inhibited platelet aggregation. Our findings may provide the basis for a novel defensive system against thrombus formation.     

Effect of anethole dithiolthione on human platelet aggregation.

Anethole dithiolthione (ADT) (10 mumol/l) inhibited platelet aggregation and the formation of thromboxane (Tx)B2 in plasma in response to adenosine diphosphate (ADP), epinephrine and arachidonic acid (AA). ADT partially inhibited platelet aggregation and TxB2 formation in plasma induced by thrombin, phorbol myristate acetate and calcium ionophore A23187 and increased the lag time of collagen-induced aggregation at concentrations in the range 10-40 mumol/l. ADT (100 mumol/l) completely inhibited the aggregation of washed platelets challenged with thrombin. ADT had no additive effect on the inhibition of thrombin-induced platelet aggregation by acetylsalicylic acid. ADT was a more effective inhibitor of AA-induced platelet aggregation than butylated hydroxytoluene. ADT inhibited the release of 3H-AA from platelet phospholipids in response to ADP and collagen. It is suggested that ADT inhibits platelet aggregation by inhibiting thromboxane synthesis and preventing AA release.     

Thrombin-induced platelet endostatin release is blocked by a proteinase activated receptor-4 (PAR4) antagonist

Endostatin is a potent endogenous inhibitor of angiogenesis that was recently shown to be stored in platelets and released in response to thrombin, but not ADP. In the present study, we have tested the hypothesis that thrombin-induced endostatin release from rat platelets is mediated via proteinase-activated receptor-4 (PAR4). Immunoprecipitation and Western blotting confirmed that endostatin is contained within rat platelets. Aggregation and release of endostatin could be elicited by thrombin (0.5 - 1.0 U ml(-1)) or by specific PAR4 agonist (AYPGKF-NH(2); AY-NH(2); 15 - 50 microM). Significant release of endostatin could be induced by a dose of thrombin below that necessary for induction of aggregation. An adenosine diphosphate (ADP) scavenger, apyrase, inhibited the platelet aggregation induced by thrombin, but not the release of endostatin. In contrast, a selective PAR4 antagonist (trans-cinnamoyl-YPGKF-NH(2); tcY-NH(2)) prevented endostatin release and aggregation induced by thrombin or by AY-NH(2). We conclude that thrombin-induced endostatin release from rat platelets is PAR4-mediated via an ADP-independent mechanism that can occur independently of platelet aggregation.     

Polyamines inhibit both platelet aggregation and glycoprotein IIb/IIIa activation

Platelet aggregation is inhibited by the polyamines putrescine, spermidine, and spermine. To date, the mechanism of action has not been clearly identified. Evidence suggests that polyamines may interact with the fibrinogen receptor (GP IIb/IIIa), interfering with platelet-platelet attachment. The effect of polyamines on human platelet aggregation and GP IIb/IIIa activation was evaluated. For the aggregation experiments, platelets were obtained from heparin- or citrate-collected blood. Our results indicate that the polyamines putrescine, spermidine, and spermine cause a dose-dependent inhibition of ADP- or collagen-mediated platelet aggregation with an order of potency spermine>spermidine>putrescine. In addition, spermine arrests or inhibits thrombin-, epinephrine-, arachidonate-, or ristocetin-induced platelet aggregation. Expression of platelet membrane glycoproteins IIb, IIIa, and IX is not reduced by polyamines. However, spermine inhibits the ADP- or thrombin-induced activation of GP IIb/IIIa. It is concluded that the final step in aggregation, common to all agonists, ie, fibrinogen binding to GP IIb/IIIa, is inhibited by spermine through inhibition of the agonist-induced activation of GP IIb/IIIa that precedes fibrinogen-ligand binding.     

Morphine-potentiated agonist-induced platelet aggregation through alpha2-adrenoceptors in human platelets

Morphine dose-dependently (0.6, 1, and 5 microM) potentiated platelet aggregation and ATP release stimulated by agonists (i.e., collagen and U46619) in washed human platelets. Furthermore, morphine (1 and 5 microM) markedly potentiated collagen (1 microg/ml) evoked an increase of intracellular Ca2+ mobilization in fura 2-AM loading human platelets. Morphine (1 and 5 microM) did not influence the binding of fluorescein isothiocyanate-triflavin to platelet glycoprotein IIb/IIIa complex. Yohimbine (0.1 microM), a specific alpha2-adrenoceptor antagonist, markedly abolished the potentiation of morphine in platelet aggregation stimulated by collagen. Moreover, morphine (0.6-5 microM) markedly inhibited prostaglandin E1 (10 microM)-induced cAMP formation in human platelets, and yohimbine (0.1 microM) significantly reversed the inhibition of cAMP by morphine (0.6 and 1 microM) in this study. Morphine (1 and 5 microM) significantly potentiated thromboxane B2 formation stimulated by collagen in human platelets, and yohimbine also reversed this effect of morphine in this study. In addition, morphine (1 and 5 microM) did not significantly affect nitrate production in human platelets. Morphine may exert its potentiation in platelet aggregation by binding to alpha2-adrenoceptors in human platelets, which leads to reduced cAMP formation and subsequently to increased intracellular Ca2+ mobilization; this, in turn, is followed by increased thromboxane A formation and finally potentiates platelet aggregation and ATP release.     

应用于抗血栓治疗的新凝血酶受体抑制剂SCH-530348

喜巴辛衍生物SCH-530348是蛋白酶激活受体1(PAR-1)的抑制剂,PAR-1是人血小板凝血酶最主要的一种受体。SCH-530348是第一种能抑制凝血酶诱导的血小板聚集而不影响凝血酶对胶原酶原活化能力的化合物。文中综述了SCH-530348的临床前和Ⅰ～Ⅲ期临床试验结果。这些研究表明SCH-530348具有降低缺血性事件发生风险的几率,同时不会明显增加机体出血的风险,临床上可应用于急性冠脉综合征的治疗和缺血性心血管事件的预防。     

木瓜蛋白酶体外对血小板聚集的抑制作用

目的：以洗涤血小板为模型,观察木瓜蛋白酶在体外对血小板聚集的抑制作用,以探讨其抗血栓作用的可能机制。方法：将不同剂量木瓜蛋白酶与洗涤血小板作用,以血小板聚集分析仪检测ADP、花生四烯酸（AA）、胶原和凝血酶诱导的血小板最大聚集率,以流式细胞仪检测活化血小板膜纤维蛋白原受体（FIB-R）和P-选择素表达水平,以SDS-PAGE分析血小板肌动蛋白聚合体的变化。检测ADP诱导的原发性高血压（PH）及急性心肌梗死（AMI）患者血小板最大聚集率和FIB-R表达水平。结果：木瓜蛋白酶剂量依赖性地抑制血小板聚集,降低血小板最大聚集水平,血小板聚集水平与木瓜蛋白酶剂量呈负相关（P〈0.01）。木瓜蛋白酶剂量依赖性地抑制ADP诱导的血小板FIB-R表达,降低FIB-R表达水平（P〈0.01）。木瓜蛋白酶降低ADP诱导的血小板膜P-选择素表达水平和抑制肌动蛋白聚合体增加（P〈0.01）。木瓜蛋白酶抑制PH及AMI患者血小板聚集和FIB-R表达（P〈0.01）。结论：木瓜蛋白酶通过抑制活化血小板膜纤维蛋白原受体的表达,并抑制肌动蛋白聚合以及释放反应从而剂量依赖性地抑制血小板的聚集反应,有抗血栓形成作用。     

眼镜蛇毒因子对大鼠血小板的激活作用(英文)

目的:研究补体激活剂眼镜蛇毒因子(CVF)对大鼠血小板的作用及其细胞机制。方法:比浊法测富血小板血浆内血小板聚集并同步记录ATP释放;生色底物法测血小板表面凝血酶原酶活性;用钙离子荧光指示剂Fura-2 AM负载血小板测细胞内游离钙;放免法测细胞内cAMP含量。结果:CVF在19.5-617nmol·L~(-1)范围内浓度依赖性地诱导血小板聚集和ATP释放,195nmol·L~(-1)诱导的ATP释放不依赖于聚集,且明显弱于凝血酶1U/ml的作用。CVF195nmol·L~(-1)时间依赖性地增加血小板表面凝血酶原酶活性。抗血小板膜糖蛋白Ⅰb/Ⅸ、Ⅲa、Ⅱb的单克隆抗体SZ-1、SZ-21、SZ-22抑制CVF诱导的血小板聚集。CVF 195nmol·L~(-1)使血小板内游离钙从静息态的(141±46)nmol·L~(-1)升高到(240±64)nmol·L~(-1),在61.7-617nmol·L~(-1)范围内轻度降低血小板内的cAMP。结论:补体激活剂CVF能诱导大鼠血小板聚集、ATP释放,增加血小板表面凝血酶原酶活性,其激活血小板的作用与血小板表面纤维蛋白原受体及血小板内游离钙升高、cAMP下降有关。     

Sphingosylphosphorylcholine,a naturally occurring lipid mediator,inhibits human platelet function

1 The lysophospholipids, lysophosphatidic acid and sphingosine 1-phosphate, have been reported to activate platelets. Here we examined effects of the naturally occurring related sphingosylphosphorylcholine (SPC) on human platelet function. 2 Platelet activation was determined as aggregation, elevation of intracellular Ca(2+) concentrations, surface expression of P-selectin, GP 53, and GP IIb/IIIa neoepitope PAC-1, and of fibrinogen binding to the platelet surface. 3 Platelets were activated by ADP (5 and 20 micro M), the thrombin receptor-activating peptide TRAP-6 (5 and 20 micro M), the thromboxane A(2) mimetic U-46619 (1 micro M) and collagen (20 and 50 micro g ml(-1)) but not by SPC (up to 20 micro M). 4 SPC concentration-dependently (IC(50) approximately 1-10 micro M) inhibited activation of washed human platelets in response to all of the above agonists, with almost complete inhibition occurring at 20 micro M SPC. 5 The SPC stereoisomers, D-erythro SPC and L-threo SPC, exhibited similar concentration-response curves in inhibiting 20 micro M ADP-induced platelet aggregation, suggesting that SPC did not act via specific lysophospholipid receptors. 6 Although SPC slightly activated platelet protein kinase A (as assessed by VASP phosphorylation), this effect could not explain the marked platelet inhibition. Possible protein kinase C inhibition also did not explain the inhibition of platelet activation by SPC. On the other hand, SPC suppressed agonist-induced Ca(2+) mobilization and phospholipase C stimulation. 7 These results indicate that the lysophospholipid SPC is an effective inhibitor of human platelet activation, apparently primarily by uncoupling agonist-activated receptors from their effectors.