thromboxane-a2 and Hemorrhagic-Disorders

A platelet disorder characterized by the absence of thromboxane A2 (TXA2)-induced platelet aggregation is a new clinical entity of platelet dysfunction. The platelets of three patients had the ability to bind exogenous TXA2, but synthetic TXA2 mimetic-induced postreceptor biochemical events, such as IP3 formation, Ca2+ mobilization, phosphatidic acid formation, and GTPase activities, were selectively defective, suggesting impaired coupling between the TXA2 receptor and phospholipase C activation. Gene analysis of the TXA2 receptor showed a substitution of Leu for Arg60 in the first cytoplasmic loop in all patients, and this mutation seemed to be responsible for this platelet disorder.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Amino Acid Substitution; Animals; Blood Platelets; Calcimycin; Calcium Signaling; CHO Cells; Codon; Cricetinae; Cricetulus; DNA, Complementary; Enzyme Activation; GTP Phosphohydrolases; Hemorrhagic Disorders; Humans; Phosphatidic Acids; Phosphatidylinositol Diacylglycerol-Lyase; Platelet Aggregation; Point Mutation; Receptors, Thromboxane; Recombinant Proteins; Sodium Fluoride; Thromboxane A2; Transfection; Type C Phospholipases

In this study, gram-positive Staphylococcus aureus lipoteichoic acid (LTA) dose-dependently (0.1-1.0 microg/ml) and time-dependently (10-60 min) inhibited platelet aggregation in human platelets stimulated by agonists. LTA also dose-dependently inhibited phosphoinositide breakdown and intracellular Ca+2 mobilization in human platelets stimulated by collagen. LTA (0.5 and 1.0 microg/ml) also significantly inhibited thromboxane A2 formation stimulated by collagen in human platelets. Moreover, LTA (0.1-1.0 microg/ml) dose-dependently decreased the fluorescence of platelet membranes tagged with diphenylhexatrience. Rapid phosphorylation of a platelet protein of Mr. 47,000 (P47), a marker of protein kinase C activation, was triggered by PDBu (30 nM). This phosphorylation was markedly inhibited by LTA (0.5 and 1.0 microg/ml) within a 10-min incubation period. These results indicate that the antiplatelet activity of LTA may be involved in the following pathways: LTA's effects may initially be due to induction of conformational changes in the platelet membrane, leading to a change in the activity of phospholipase C, and subsequent inhibition of phosphoinositide breakdown and thromboxane A2 formation, thereby leading to inhibition of both intracellular Ca+2 mobilization and phosphorylation of P47 protein. Therefore, LTA-mediated alteration of platelet function may contribute to bleeding diathesis in gram-positive septicemic and endotoxemic patients.

Topics: Calcium Signaling; Cell Membrane; Collagen; Cytosol; Dose-Response Relationship, Drug; Endotoxemia; Enzyme Activation; Gram-Negative Bacterial Infections; Gram-Positive Bacterial Infections; Hemorrhagic Disorders; Humans; L-Lactate Dehydrogenase; Lipopolysaccharides; Membrane Fluidity; Membrane Lipids; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Peptides; Phorbol 12,13-Dibutyrate; Phosphatidylinositols; Phosphorylation; Platelet Aggregation; Platelet Aggregation Inhibitors; Platelet Glycoprotein GPIIb-IIIa Complex; Protein Kinase C; Protein Processing, Post-Translational; Sepsis; Shock, Septic; Staphylococcus aureus; Teichoic Acids; Thromboxane A2; Thromboxane B2

Valproate (VPA) is an extensively used drug in the therapy of epilepsies. One of the most frequently reported side effects of VPA is hemorrhagic diathesis. Some authors emphasized the decreased platelet count as the basis of VPA-induced hemorrhagic diathesis, but some reports suggested that a significant proportion of patients with normal platelet count may still have an altered platelet function. The mechanism of the VPA-induced platelet dysfunction has not yet been elucidated. A determining element of platelet functions is the arachidonate cascade. Present ex vivo experiments were designed to determine whether a relation exists between the incidence of hemostasis caused by VPA and the effect of this drug on the arachidonate cascade of platelets.. Platelets were isolated from patients receiving long-term VPA treatment (serum level, 36.04+/-16.12 microg/ml; n = 10) or carbamazepine (CBZ) treatment (serum level, 5.24+/-2.67 microg/ml; n = 10) and were labeled with [14C]arachidonic acid. (CBZ-treated patients were chosen as a control group, because CBZ causes blood dyscrasias similar to those elicited by VPA, but there has been no report that CBZ induces a platelet dysfunction.) The 14C-eicosanoids were separated by means of overpressure thin-layer chromatography and determined quantitatively by liquid scintillation.. Even when the mean plasma concentration of the drug was low, VPA treatment reduced the activity of the arachidonate cascade in platelets. VPA effectively inhibited the cyclooxygenase pathway and the synthesis of the strong platelet aggregator thromboxane A2.. Inhibition of the platelet arachidonate cascade may contribute to the platelet-function alterations caused by VPA.

Topics: Adult; Arachidonic Acids; Blood Coagulation Tests; Blood Platelets; Epilepsy; Fasting; Hemorrhagic Disorders; Humans; Male; Platelet Activation; Platelet Aggregation; Thromboxane A2; Valproic Acid

We reported that aspirin (ASA) abnormally prolongs bleeding time (BT) in uremia. The present study was designed to investigate whether the abnormally prolonged post-ASA BT in uremia is due to different ASA pharmacokinetics and bioavailability that might be a consequence of uremic condition, platelet cyclooxygenase is peculiarly sensitive to ASA in uremia, and ASA affects primary hemostasis in uremia by a mechanism independent of cyclooxygenase inhibition. Our results showed that in patients with uremia, but not in normal subjects, ASA markedly prolongs the BT. This effect is transient and depends on the presence of ASA in the blood. The observed differences in ASA kinetic parameters are not an explanation of the exaggerated effect of ASA on primary hemostasis in uremia. The sensitivity of platelet cyclooxygenase to ASA inhibition is comparable in uremics and in normal subjects. The temporal dissociation between ASA-induced prolongation of BT and the effect on platelet thromboxane A2 generation suggests that ASA inhibits platelet function in uremia by a mechanism distinct from cyclooxygenase blocking. This possibility is strengthened by the observation that ibuprofen at a dose that fully inhibits platelet cyclooxygenase activity does not significantly prolong BT.

Topics: Acetylation; Adult; Aged; Aspirin; Bleeding Time; Blood Platelets; Cyclooxygenase Inhibitors; Female; Hemorrhagic Disorders; Humans; Ibuprofen; Kinetics; Male; Middle Aged; Platelet Function Tests; Thromboxane A2; Uremia

The authors describe a patient with a longstanding bleeding disorder associated with impaired platelet aggregation and secretion despite normal granule contents. Thrombin-induced platelet thromboxane A2 production, measured using a radioimmunoassay for thromboxane B2, was markedly decreased or undetectable in platelet-rich plasma and whole blood serum. However, significant amounts of thromboxane B2 were detected on thrombin stimulation of platelets suspended in albumin-free salt medium. Malondialdehyde and 14C-hydroxyheptadecatrienoic acid production was undetectable in the patient's platelets. Liberation of free 14C-arachidonic acid from phospholipids during stimulation of prelabeled platelets was normal, indicating normal phospholipase activity. These observations indicate an albumin-dependent partial deficiency in thromboxane production resulting from a defect either in cyclooxygenase or thromboxane synthetase. Further, the authors studied the effect of albumin on arachidonic acid metabolism in normal platelets. These studies indicate that albumin enhances liberation of arachidonic acid from phospholipids but has an overall inhibitory effect on thromboxane synthesis.

Topics: Adult; Albumins; Arachidonic Acid; Arachidonic Acids; Bleeding Time; Blood Platelet Disorders; Blood Platelets; Chromatography, Gel; Female; Hemorrhagic Disorders; Humans; Malondialdehyde; Partial Thromboplastin Time; Phospholipids; Platelet Aggregation; Platelet Count; Serotonin; Thrombin; Thromboxane A2; Thromboxane B2; Thromboxanes