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PubMed1994

Negative feedback regulation of human platelets via autocrine activation of the platelet-derived growth factor alpha-receptor.

Human platelets contain platelet-derived growth factor (PDGF) in their alpha-granules which is released during platelet exocytosis. We show by immunoprecipitation and 125I-PDGF binding experiments that human platelets have functionally active PDGF alpha-receptors, but not beta-receptors. The PDGF alpha-receptor (PDGFR-alpha) was identified as a 170-kDa glycosylated protein-tyrosine kinase as found in other cell types. Stimulation of platelets with 0.1 unit/ml thrombin resulted in a significant increase (2-5-fold) of the tyrosine phosphorylation of the PDGFR-alpha, as determined by immunoprecipitation with phosphotyrosine antiserum as well as with PDGFR-alpha antiserum. The observed thrombin-induced autophosphorylation of the PDGFR-alpha was inhibited by the addition of a neutralizing monoclonal PDGF antibody. Thus, our results suggest that the platelet PDGFR-alpha is stimulated in an autocrine manner by PDGF secreted during platelet activation. Preincubation of platelets with PDGF inhibited thrombin-induced platelet aggregation and secretion of ATP + ADP and beta-hexosaminidase. Thrombin-induced platelet aggregation was also reversed when PDGF was added 30 s after thrombin stimulation. Inhibition of the autocrine PDGF pathway during platelet activation by the PDGF antibody led to a potentiation of thrombin-induced beta-hexosaminidase secretion. Thus, the PDGFR-alpha takes part in a negative feedback regulation during platelet activation. Our demonstration of PDGF alpha-receptors on human platelets and its inhibitory function during platelet activation identifies a new possible role of PDGF in the regulation of thrombosis.

Becaplermin↗
PubMed1985

Botrocetin- and polybrene-induced platelet aggregation in platelet-type von Willebrand disease.

It has been reported that botrocetin, a Bothrops venom factor, induces platelet aggregation dependent on von Willebrand factor (vWF), and that platelet aggregation induced by Polybrene, a synthetic polycation, is enhanced by vWF. This report describes the platelet aggregability on stimulation with botrocetin and Polybrene in four patients with platelet-type von Willebrand disease (vWD) who showed increased platelet aggregation with low concentrations of ristocetin as the result of a platelet abnormality. Enhanced platelet aggregability with botrocetin was observed in platelet-rich plasma (PRP) from the patients. Platelet aggregation induced by botrocetin in a mixture of normal washed platelets and patient plasma was either decreased or normal, being dependent on the amount of plasma vWF. In contrast with ristocetin and botrocetin, Polybrene did not cause increased aggregation of patient PRP. Polybrene aggregated normal washed platelets less extensively in the presence of patient plasma than normal plasma. These studies demonstrated that botrocetin induced heightened interaction between platelets and vWF, but Polybrene did not, in platelet-type vWD, and that the enhanced responsiveness of patient platelets to botrocetin is related to an intrinsic platelet abnormality.

Crotalid Venoms↗
PubMed1991

Effect of calcium concentration and inhibitors on the responses of platelets stimulated with collagen: contrast between human and rabbit platelets.

1. Variations in the concentration of Ca2+ [Ca2+] in the suspending medium have different effects on the responses of human and rabbit platelets to collagen. 2. When rabbit platelets are stimulated with a low concentration of collagen (0.5 micrograms/ml), aggregation, release of granule contents, and formation of thromboxane are maximal when the suspending medium contains [Ca2+] in the physiological range (0.5-2.0 mM), and very slight in a medium with no added Ca2+. 3. In contrast, human platelets respond most strongly when the suspending medium contains no added Ca2+ [( Ca2+] approx. 20 microM); this is attributable to the enhanced formation of thromboxane A2 (TXA2) upon close platelet-to-platelet contact in this medium. 4. When TXA2 formation is blocked by inhibition of cyclo-oxygenase with aspirin or indomethacin, rabbit platelet aggregation and release in response to 1.25-10 micrograms/ml collagen is also maximal at [Ca2+] of 0.5-2.0 mM and least at 20 microM; human platelets do not aggregate and the extent of release is relatively independent of [Ca2+]. 5. In 1 mM [Ca2+], use of apyrase and/or ketanserin with rabbit platelets in which TXA2 formation is blocked shows that released ADP and serotonin make large contributions to aggregation and release in response to high concentrations of collagen; human platelet aggregation is largely dependent on TXA2. 6. Use of fura-2-loaded platelets shows that the collagen-induced rise in cytosolic [Ca2+] is only slightly inhibited by aspirin or indomethacin in rabbit platelets, but almost completely inhibited in human platelets. 7. Responses of rabbit platelets to collagen are less dependent on TXA2 than those of human platelets. Released ADP and serotonin make major contributions to the responses of rabbit platelets to collagen.

Animals↗
PubMed1980

Binding of adenosine diphosphate to human blood platelets and to isolated blood platelet membranes.

The equilibrium binding of 14C-labeled ADP to intact washed human blood platelets and to platelet membranes was investigated. With both intact platelets and platelet membranes a similar concentration dependence curve was found. It consisted of a curvilinear part below 20 microM and a rectilinear part above this concentration. At high ADP concentrations, the rectilinear part appeared to be saturable. Because of this, two classes of saturable ADP binding sites were proposed. ADP was partly converted to ATP and AMP with intact platelets while this conversion was virtually absent in isolated platelet membranes. ADP was bound to platelet membranes with the same type of curves found for intact platelets. The ADP binding to the high affinity system, which was stimulated by calcium ions, was nearly independent of temperature and had a pH optimum at 7.8. A number of agents were investigated for inhibiting properties. Of the sulfhydryl reagents only p-chloromercuribenzene sulfonate inhibited both high and low affinity binding systems while iodoacetamide and N-ethylmaleimide were without effect. Compounds acting via cyclic AMP on platelet aggregation, such as adenosine and cyclic AMP itself, had no influence on binding. Some nucleosidediphosphates and nucleotide analogs at a concentration of 100 microM had no, or only a slight, effect on high affinity ADP binding. For some other nucleotides inhibitor constants were determined for both platelet ADP aggregation and ADP binding. The inhibitor constants of ATP, adenyl-5'-yl-(beta,gamma-methylene)diphosphate, IDP, adenosine-5'(2-O-thio)diphosphate, for aggregation and high affinity binding were in good correlation with each other. Exceptions formed fluorosulfonylbenzoyl adenosine and AMP. The ATP formation found with intact platelets could be attributed to a nucleosidediphosphate kinase. It was investigated in some detail. The enzyme was magnesium dependent, had a Q10 value of 1.41, a pH optimum at 8.0, was competitively inhibited by AMP and reacted via a ping pong mechanism. All findings described in this paper indicate that platelets as well as platelet membranes bind ADP with the same characteristics and they suggest that the high affinity binding of ADP is involved in platelet aggregation induced by ADP. The results on nucleosidediphosphate kinase did not permit a firm conclusion about the role of the enzyme in induction of platelet aggregation by ADP.

Adenosine Diphosphate↗
PubMedDate not supplied

The utility of platelet washing using an automated procedure for severe platelet allergic reactions.

Increased use of platelets in patients requiring chronic platelet support has increased platelet transfusion reactions. The authors reviewed more than 300 platelet transfusion reactions, evaluated an automated platelet washing technique, and studied the effectiveness of washing platelets to reduce reactions. Febrile reactions (66%) were most frequently reported, followed by moderate and severe allergic reactions (15%), and urticaria alone (19%). Washed platelets were prepared by an automated technique (IBM/COBE 2991). In vitro studies indicated no apparent adverse effects to the platelets due to the wash procedure, and in vivo studies demonstrated good platelet increments in 10 thrombocytopenic patients. Twenty-two patients with histories of platelet transfusion reactions received a total of 554 washed platelet transfusions. Washed platelets were not effective in reducing febrile transfusion reactions in 16 patients receiving 347 washed products. The efficacy of washed platelets in reducing transfusion reactions was demonstrated in six patients with histories of severe allergic reactions who received 207 washed products. Severe allergic reactions were completely alleviated in this group. In conclusion, automated platelet washing is simple and efficacious in preventing or reducing the severity of allergic reactions to platelet transfusions.

Adult↗
PubMed2001

Platelet-specific antibodies in HLA-immunized patients receiving chronic platelet support.

BACKGROUND: In HLA-alloimmunized patients, the unexpected failure of HLA-matched platelet transfusions usually raises the suspicion about concomitant platelet-specific antibodies. As the reported frequency of platelet-specific antibodies in multitransfused patients varies widely, the aim of this study was to determine the prevalence of such antibodies in a population of chronic thrombocytopenic patients with HLA antibodies. STUDY DESIGN AND METHODS: From 1985 to 1997, 11,777 determinations of HLA antibodies were performed in 1330 hematologic patients receiving chronic platelet support. Fifty-two patients with HLA alloimmunization that lasted more than 1 month were selected. The search for platelet-specific antibodies was performed by using a monoclonal antibody immobilization of platelet antigens assay, thus allowing the identification of platelet-specific antibodies directed against the platelet glycoproteins (GP) Ib/IX, GPIIb/IIIa, and GPIa/IIa. Specificity of the platelet-specific antibodies was further investigated by using a solid-phase assay with chloroquine-treated platelets. RESULTS: Only 2 (3.8%) of the 52 patients had platelet-specific antibodies. One antibody reacted with an epitope of the GPIIb/IIIa that was present in all the panel platelets, and that probably was an autoantibody. The other was an anti-HPA-5b. CONCLUSIONS: The prevalence of platelet-specific antibodies in patients with HLA alloimmunization is very small. The search for concomitant platelet-specific antibodies would be indicated only when other causes of refractoriness to HLA-matched platelets are ruled out.

HLA Antigens↗
PubMed1992

The significance of platelets with increased RNA content (reticulated platelets). A measure of the rate of thrombopoiesis.

Direct flow cytometric measurement of nucleic acid content in individual platelets is possible using the fluorescent dye Thiazole Orange (Becton-Dickinson, San Jose, CA). When applied to studies of thrombocytopenic patients, platelets with elevated nucleic acid content ("reticulated platelets") can be identified and quantitated. Labeling of these platelets is saturable and is abolished by treatment with RNAse. It has been suggested that, similar to the erythrocyte reticulocyte response to anemia, the number of these platelets appearing in the circulation may provide an estimate of the rate of thrombopoiesis. The authors studied 229 thrombocytopenic patients, measuring both reticulated platelets and platelet-associated immunoglobulin. The results show that for the subset of patients with normal levels of platelet-associated immunoglobulin, the average absolute number of reticulated platelets is independent of platelet count and remains in the normal range. For those with elevated levels of platelet-associated immunoglobulin, the absolute number of reticulated platelets increases in patients who are moderately thrombocytopenic (60 to 100 x 10(9)/L) but decreases to normal or subnormal levels as thrombocytopenia worsens. The latter finding has been duplicated in studies of mice made thrombocytopenic by injection of anti-platelet antiserum. These results are consistent with the hypothesis that reticulated platelets are subject to peripheral destruction at the same rate as mature platelets, and that in the severely thrombocytopenic patient their level may decrease despite an appropriate marrow thrombopoietic response.

Animals↗
PubMed2001

Activated platelets enhance microparticle formation and platelet-leukocyte interaction in severe trauma and sepsis.

BACKGROUND: Activated platelets have been recently reported to produce platelet microparticles and to enhance platelet-leukocyte interaction. The precise role of platelets in systemic inflammatory response syndrome (SIRS) has not been clarified. The objective of this study was to evaluate microparticle formation and platelet-leukocyte interaction in severe trauma and sepsis. METHODS: Twenty-six patients with severe SIRS (SIRS criteria and serum C-reactive protein > 10 mg/dL) and 12 healthy volunteers were studied. The severe SIRS was caused by trauma in 12 patients and sepsis in 14. Microparticle formation, P-selectin expression on platelets, platelet-monocyte binding, and platelet-polymorphonuclear leukocyte (PMNL) binding were measured by flow cytometry in the presence or absence of ionomycin, N-formyl-methionyl-leucyl-phenylalanine, or anti-CD62p monoclonal antibody. Soluble P-selectin, thrombomodulin, neopterin, and PMNL elastase in blood were also measured. RESULTS: Microparticle formation, P-selectin expression on platelets, platelet-monocyte binding with or without ionomycin, and platelet-PMNL binding with ionomycin significantly increased in patients with severe SIRS in comparison with values in normal volunteers. The increased platelet-leukocyte binding in severe SIRS patients was markedly inhibited by P-selectin blockade and was not enhanced by N-formyl-methionyl-leucyl-phenylalanine. Soluble P-selectin, thrombomodulin, neopterin, and PMNL elastase in blood also increased in these patients. CONCLUSION: Activated platelets enhance microparticle formation and platelet-leukocyte interaction in severe trauma and sepsis. Enhanced platelet-leukocyte interaction is dependent on P-selectin expression and may be involved in the systemic inflammatory response after severe inflammatory insult.

Adolescent↗
PubMed1996

Oxidized LDL decreases L-arginine uptake and nitric oxide synthase protein expression in human platelets: relevance of the effect of oxidized LDL on platelet function.

BACKGROUND: Oxidized LDL (ox-LDL) promotes vasoconstriction and platelet activation. The present study was undertaken to determine the involvement of the L-arginine-nitric oxide (NO) pathway in ox-LDL-mediated platelet activation. METHODS AND RESULTS: Washed human platelets were incubated with native LDL or ox-LDL for 1 hour at 37 degrees C followed by measurement of platelet function and indexes of the L-arginine-NO pathway. Ox-LDL but not native LDL caused a concentration-dependent increase in thrombin-induced platelet aggregation and 14C-serotonin release. These effects of ox-LDL were inhibited by pretreatment of platelets with L-arginine, the precursor of NO. Ox-LDL also caused a concentration-dependent reduction in the uptake of 3H-L-arginine by platelets. In addition, NO synthase activity, measured as conversion of 3H-L-arginine to 3H-L-citrulline, decreased on incubation of platelet cytosol with ox-LDL. Nitrite production was also reduced by treatment of platelets with ox-LDL. These effects of ox-LDL on NO synthase activity and nitrite production were reversed by pretreatment of platelets with L-arginine. Concurrent with the decrease in NO production, cytosolic cGMP was inhibited in ox-LDL-treated platelets. The inhibitory effects of ox-LDL were dependent in part on the increase of cholesterol in the platelets. Western blot analysis demonstrated approximately 50% reduction in the expression of NO synthase protein in platelets treated with ox-LDL. CONCLUSIONS: These observations indicate that the L-arginine-NO pathway is involved in the effects of ox-LDL on platelet function and that ox-LDL stimulates platelet function primarily by diminishing NO synthase expression as well as decreasing the uptake of L-arginine.

Arginine↗
PubMed1968

The effect of platelet age on platelet adherence to collagen.

The adherence to collagen of rabbit platelets labeled in vivo with (35)SO(4) (=) has been studied both in vitro and in vivo. The young platelets are labeled with (35)SO(4) (=) 2-3 days after administration of the isotope to the animals. We exposed platelet-rich plasma (ethylenediamine-tetraacetate, EDTA, as anticoagulant), prepared from blood taken from rabbits 54 hr after giving the (35)SO(4) (=), to collagen in vitro. There was a fall in the specific radioactivity of the nonadherent platelets which indicated a selective adhesion of young platelets to the collagen. In experiments designed to have most of the (35)S label in the oldest platelets it was found that exposure of plasma containing these platelets to collagen resulted in an increase in the specific radioactivity of the nonadherent platelets. Similar observations were obtained when glycine-(14)C was used as a platelet label. However, when DF(32)P (di-isopropyl phosphorofluoridate-(32)P), which is thought to label platelets of all ages equally, was used, the adherence of platelets to collagen did not result in any changes in the specific activity of the nonadherent platelets. In in vivo studies in which we infused a collagen suspension into rabbits 54 hr after giving (35)SO(4) (=) we found that the specific radioactivity of the platelets remaining in the circulation fell. This did not occur when we infused the collagen 96 hr after giving the (35)SO(4) (=). The results from these studies indicate that young platelets adhere to collagen more readily than older platelets.

Adenine Nucleotides↗
PubMed1998

Activity of platelet in patients with high level of LDL and the effect of LDL on platelet glycoproteins.

OBJECTIVE: To investigate the activity of platelet in patients with high level of low density lipoproteins (LDL) and the effect of LDL on platelet glycoproteins (GP). METHODS: Platelet glycoproteins, whose platelet were activated or unactivated, were measured by flow cytometry. RESULTS: The amount of the platelet membrane GP II b/III a in patients was not significantly different from that of the control when platelet was unactivated (P > 0.05); when platelet was activated by adenosine diphosphate (ADP), the amount of patients' platelet GP II b/III a was increased markedly in comparison with that in the control (P < 0.01). Having been preincubated with LDL, the platelets were activated by ADP and the amounts of GP II b/III a of patients group and the control were all increased obviously as compared with those in the platelets which were not incubated with LDL (P < 0.01; P < 0.01). There was no significant difference of the amount of granule membrane protein-140 (GMP-140) between patients and the control when platelet was activated by thrombin (P > 0.05). Having been preincubated with LDL, platelet was activated by thrombin, the amounts of GMP-140 on patients' and control platelet were all increased markedly (P < 0.01, P < 0.01). CONCLUSION: The activity of platelet in patients with high level of LDL is increased significantly. LDL can increase the expression of platelet glycoproteins.

Adult↗
PubMed1987

Thrombin interaction with platelets. Influence of a platelet protease nexin.

A fraction of the 125I-thrombin that binds to human platelets is taken into a sodium dodecyl sulfate-resistant 77 kDa complex with a platelet factor (Bennett, W. F., and Glenn, K. C. (1980) Cell 22, 621-627). Here we show that this platelet factor is in several respects similar to protease nexin I (PNI), a fibroblast thrombin inhibitor. The complexes are of the appropriate size, bind to Sepharose that has been derivatized with anti-PNI antibody, do not form when the thrombin active site has been blocked with diisopropylphosphofluoridate, and do not appear on platelets when heparin is present. However, the platelet factor does not bind urokinase, indicating that this "platelet PN" may be distinct from PNI. Following brief incubation with 125I-thrombin, platelet PN X 125I X thrombin complexes are found both associated with the platelets and free in the binding medium. 125I-Thrombin has a higher affinity for platelet PN than for platelet receptors. In 30-s binding incubations carried out with thrombin at concentrations below 0.3 nM, formation of the 77-kDa complex accounts for most of the platelet specific binding of 125I-thrombin. Subtracting this large contribution to 125I-thrombin-specific binding reveals that the reversible binding of 125I-thrombin to platelet receptors exhibits sigmoidal thrombin dose-dependence. Thrombin stimulation of platelet [14C]serotonin release exhibits similar thrombin dose dependence. These results indicate that platelets may possess a mechanism for suppressing their interaction with active thrombin at thrombin doses below 0.3 nM. It is possible that platelet PN carries out this function by capturing thrombin before thrombin binds to its signal-transmitting receptors.

Amyloid beta-Protein Precursor↗
PubMedDate not supplied

Effect of methods of platelet resuspension on stored platelets.

Platelets are prepared from whole blood by differential centrifugation. Following their isolation as a platelet button, platelets are allowed to rest for a short period in the residual plasma before resuspension. In this study, the feasibility of resuspending platelets without this rest period is studied. A total of 35 platelet concentrates (PC) were prepared from blood collected in CPDA-1 and resuspended by one of the following four methods: (1) no resting period, PC placed on a rotator immediately after preparation, (2) a 1.5 hour rest period and gentle shaking prior to rotation, (3) no rest period and immediate gentle shaking prior to rotation, and (4) a 1.5 hour resting phase and no shaking prior to rotation. Following the previous processing, all platelet concentrates were stored for 72 hours on an elliptical platelet rotator at 20 to 24 degrees C to provide continuous agitation. A number of in vitro tests were used as indicators of platelet viability during storage. These included platelet morphology, pO2, pCO2, pH, osmotic recovery, number of platelets in the concentrate, and platelet volume distribution. Our findings demonstrate that platelets are of similar quality after storage in all of the four groups described. Our studies suggest that the resting phase is unnecessary for platelet preparation. The elimination of the resting phase would allow platelet concentrates to be available sooner and improve cost-effectiveness of platelet preparation.

Blood Platelets↗
PubMed1992

Deficiency of P62, a putative collagen receptor, in platelets from a patient with defective collagen-induced platelet aggregation.

Recently, we described a platelet antibody against a putative collagen receptor (P62), which was found in a patient with idiopathic thrombocytopenic purpura (ITP) (Blood 69:1712). We now report a deficiency of the P62 receptor in a young man whose platelets showed defective collagen-induced platelet aggregation. He had a mild bleeding tendency and slight thrombocytopenia. The results of coagulation and fibrinolysis studies were normal. The patient's platelets were partially unresponsive to collagen, although aggregation in response to ADP, thrombin, ristocetin, and calcium ionophore (A23187) was almost normal. Adhesion of his platelets to bovine collagen was markedly reduced. Addition of collagen caused no synthesis of thromboxane (TX)B2 in platelet rich plasma (PRP) from this patient. Furthermore, collagen produced no rise of cytosolic free calcium ([Ca2+]i) in fura2-loaded platelets. In contrast, thrombin caused TXB2 formation and an increase of [Ca2+]i in his platelets. These results suggest defective interaction between the platelets and collagen. The IgG from the ITP-patient induced irreversible aggregation in normal PRP, but caused no aggregation of the young man's platelets. Immunoblot studies showed that normal platelets had antigens with a molecular weight of 62 KDa under reducing conditions and of 57 KDa under nonreducing conditions. In contrast, the young man's platelets had no P62 band, although GPIa/IIa and thrombospondin were normally present. These results indicate that impaired collagen-induced aggregation in the patient's platelets was due to a deficiency of P62 and confirm that P62 may play a crucial role as a collagen receptor in platelet activation.

Adenosine Triphosphate↗
PubMed1994

1-Deamino-8-D-arginine vasopressin (DDAVP) increases platelet membrane expression of glycoprotein Ib in patients with disorders of platelet function and after cardiopulmonary bypass.

1-deamino-8-D-Arginine vasopressin (DDAVP) shortens the bleeding time in some patients with platelet dysfunction and decreases blood loss in some cardiopulmonary bypass patients. We studied platelet membrane glycoproteins in patients with von Willebrand disease (vWD), disorders of platelet function, and in cardiopulmonary bypass patients after infusion of 0.3 microgram/kg of DDAVP. Platelets from 8 cardiopulmonary bypass patients, receiving DDAVP immediately after surgery, were compared to those of 14 patients not receiving DDAVP. We also studied 12 patients with vWD, and 8 patients with platelet dysfunction receiving DDAVP. Fixed platelets, stained with monoclonal fluorescein (FITC)-labeled antibodies directed against GPIb (CD42b antigen), GPIb/IX, GPIIb/IIIa (CD41a antigen), CD63 antigen (a platelet activation protein), and P-selectin (CD62 antigen) were studied by flow cytometry. Binding of CD42b monoclonal antibody (MoAb) and anti-GPIb/IX to platelets from both groups of bypass patients increased during the 18-20 hr after surgery, but the group receiving DDAVP showed the greater increase (P = 0.032). Platelets from patients receiving DDAVP for vWD or for platelet dysfunction, had increases in CD42b MoAb and anti-GPIb/IX binding (P < 0.01) that coincided with shortening of their bleeding time. No changes were seen in binding of other antibodies. When platelets from normal donors were incubated with DDAVP for 20 hr, there were increases in platelet surface CD42b MoAb binding, while immunogold-stained transmission electron micrographs of permeabilized platelets demonstrated decreases in cytoplasmic CD42b MoAb binding. DDAVP increases platelet membrane GPIb expression in a variety of patients and may account for improvement in hemostasis seen in some studies. Redistribution of GPIb from the cytoplasm to the membrane may account for this increased expression.

Adult↗
PubMed1993

Studies on the role of platelet eicosanoid metabolism and integrin alpha IIb beta 3 in tumor-cell-induced platelet aggregation.

Platelet eicosanoid metabolism resulting from tumor-cell-induced platelet aggregation (TCIPA) was examined in a homologous in vitro system. Rat Walker 256 carcinosarcoma cells induced the aggregation of rat platelets via a thrombin-dependent mechanism with concomitant production of eicosanoid metabolites (e.g., 12-HETE, TXA2). TCIPA was dependent on the concentration of tumor cells inducing aggregation, as well as cyclooxygenase and lipoxygenase products. Cyclooxygenase inhibitors, but not lipoxygenase inhibitors, blocked platelet aggregation induced in vitro by a low concentration of agonist. At a high agonist concentration, neither cyclooxygenase nor lipoxygenase inhibitors alone affected platelet aggregation; however, the combined inhibition of both the cyclooxygenase and lipoxygenase pathways resulted in subsequent inhibition of platelet aggregation regardless of agonist concentration. The extent of platelet TXA2 and 12-HETE biosynthesis was likewise dependent on and correlated with agonist concentration. The inhibitors used in this study did not significantly inhibit protein kinase C activity at the doses tested. Platelet surface glycoprotein alpha IIb beta 3 play an important role in platelet aggregation. The effect of platelet cyclooxygenase and lipoxygenase inhibition in regulating alpha IIb beta 3 surface expression was examined by flow cytometric analysis. Thrombin stimulation of washed rat platelets resulted in significantly increased surface expression of platelet alpha IIb beta 3 integrin complex. The enhanced surface expression was not inhibited by a cyclooxygenase inhibitor (aspirin), a thromboxane synthase inhibitor (CGS-14854) or a thromboxane receptor antagonist (SQ 29,548), nor was it stimulated by a thromboxane A2 mimic (pinane-thromboxane A2). However, alpha IIb beta 3 expression was blocked by lipoxygenase inhibition and stereospecifically increased by the platelet lipoxygenase metabolite 12(S)-HETE. These results suggest that both the platelet lipoxygenase and cyclooxygenase pathways are important for TCIPA but that different mechanisms of action are involved.

Animals↗
PubMed1986

Synergistic platelet activation by aggregates of IgG and the phospholipid platelet-activating factor 1-O-alkyl-2-acetyl-SN-glycero-3-phosphorylcholine.

Immune complexes and 1-O-alkyl-2-acetyl-SN-glycero-3-phosphorylcholine (AGEPC) are potent platelet-activating factors which interact with distinct receptors on human platelets. The mechanisms of platelet activation by these two stimuli were investigated by examining the effects of AGEPC on human platelets which had been preexposed to IgG aggregates (IgG-Agg). AGEPC and IgG-Agg in combination activated platelet aggregation synergistically, whereas release of the granular constituent serotonin was not increased. Synergy was maximal within 1 min of exposure of platelets to IgG-Agg and persisted for 5 to 10 min after exposure to IgG-Agg. Synergy was observed at concentrations of IgG-Agg which release platelet granular constituents with minimal aggregation. Monomeric IgG, which did not activate platelets, did not augment the platelet response to AGEPC. Platelet activation by AGEPC is enhanced by both the granular constituent adenosine diphosphate (ADP) and by arachidonic acid, suggesting that one or both agents may contribute to synergy between AGEPC and IgG-Agg. Indomethacin inhibited granule release by IgG-Agg and enhancement of AGEPC-induced platelet aggregation by IgG-Agg. The ADP scavengers creatine phosphate/creatine phosphokinase also blocked synergistic platelet activation. These data suggest that IgG-Agg releases platelet granule ADP by a cyclooxygenase-dependent mechanism and the released ADP, in combination with AGEPC, activates platelets synergistically.

Adenosine Diphosphate↗
PubMed1994

Platelet secretory phospholipase A2 fails to induce rabbit platelet activation and to release arachidonic acid in contrast with venom phospholipases A2.

The ability of platelet secretory phospholipase A2 (sPLA2) to induce platelet activation was investigated. sPLA2 (group II) contained in an activated platelet supernatant, as well as high concentrations of purified recombinant platelet sPLA2, failed to induce platelet activation. Furthermore, sPLA2 did not modify platelet activation induced by various agonists. The possible relationship between the failure of this enzyme to induce platelet activation and its origin (mammalian) or its structural group (group II) was then investigated, using pancreatic PLA2s (group I) and venom PLA2s from groups I, II and III. All venom PLA2s induced platelet activation that was accompanied by the liberation of arachidonic acid and was abolished by aspirin. In contrast, as observed for platelet sPLA2, enzymes from hog or bovine pancreas were unable to induce platelet activation even when used at high concentrations. Interestingly, PLA2 able to induce platelet activation efficiently hydrolyse phosphatidylcholine, while those inactive on platelets did not. Taken together, these results suggest that the catalytic activity of added PLA2 is necessary but not sufficient to induce platelet activation. Moreover, the ability of PLA2 to induce platelet activation is not related to its structural group (I, II, III) but rather to its origin (venom vs. mammalian) and capacity to hydrolyse phosphatidylcholine, the major phospholipid of the outer leaflet of the plasma membrane.

Animals↗