Formation of an Intermediate in Prostaglandin Biosynthesis and Its Association with the Platelet Release Reaction

276

A B S T R A C T When thrombin is added to washed human platelets, one of its actions results in activation of a phospholipase that hydrolyzes arachidonic acid from phospholipids. The arachidonate is converted to the cyclic endoperoxides (prostaglandin G2 and prostaglandin H2) by fatty acid cyclo-oxygenase. These compounds are then converted to thromboxane A2, also called rabbit aorta-contracting substance, by thromboxane synthetase. These labile, pharmacologically active compounds then break down to inactive products including thromboxane B2 and malonaldehyde. Incubation of platelets with either dibutyryl cyclic adenosine 3',5'-monophosphate (dBcAMP) or prostaglandin E, (PGE,) before thrombin addition blocks the subsequent formation of oxygenated products of arachidonic acid including thromboxane A2, thromboxane B2, and malonaldehyde. In contrast, when arachidonic acid is added directly to platelets, prior incubation with dBcAMP or PGE, does not inhibit production of the prostaglandins or their metabolites. Thrombin treatment of platelets also blocks the acetylation of cyclo-oxygenase by aspirin since the hydrolyzed arachidonic acid competes with aspirin for the active site on cyclo-oxygenase. Prior treatment of platelets with dBcAMP or PGE, reverses the thrombin inhibition of the acetylation of cyclo-oxygenase. We conclude that agents which elevate platelet cAMP levels inhibit the hydrolysis of arachidonic acid from platelet phospholipids. We also find that prostaglandin synthesis can be dissociated, in part, from platelet aggregation and release, and that cAMP has separate actions on these processes. Higher thrombin concentrations are required to stimu-

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Cyclic adenosine 3',5'-monophosphate inhibits the availability of arachidonate to prostaglandin synthetase in human platelet suspensions.

Minkes¹,

Stanford²,

M³

et al. 1977

276

“…Cause and mechanism of this phenomenon have recently been elucidated primarily by the studies of Hamberg et al (3,4), Smith et al (5), and Willis et al (6,7). The rapid synthesis of prostaglandins during platelet aggregation (8) involves the transient formation of endoperoxide intermediates, extremely potent inducers of platelet aggregation.…”

Section: Introductionmentioning

confidence: 99%

Vitamin E. An inhibitor of the platelet release reaction.

Steiner¹,

Anastasi²

1976

206

A B S T R A C T Formation of lipid peroxides rises sharply when platelets undergo the release reaction. In this study the in vitro effect of vitamin E on platelet aggregation was investigated. a-Tocopherol, an antioxidant of known inhibitory action on lipid peroxidation, was added to platelet suspensions in concentrations up to 1.5 mM. A dose-dependent reduction in platelet aggregation was observed, with complete inhibition of the secondary wave of aggregation at > 0.9 mM atocopherol. The inhibitory effect of a-tocopherol on the platelet release reaction was further documented by the decrease in aggregation-induced release of [14C]5-hydroxytryptamine from prelabeled platelets and by the reduction of N-acetylglucosaminidase activity released into the medium. The sharp rise in lipid peroxides normally associated with platelet aggregation was markedly reduced by a-tocopherol and also by acetylsalicylic acid, a known inhibitor of the platelet release reaction.In vivo studies examined the effect of oral vitamin E administration (1,200-2,400 IU daily) on plasma and platelet levels of a-tocopherol. Up to 1,800 IU daily, increasing dosages of vitamin E resulted in increasing concentrations of a-tocopherol in plasma and platelets, but intake of vitamin E in excess of this dosage failed to show any further increase in plasma or platelet levels.

“…The effect of creatine phosphate and creatine phosphokinase or of apyrase. To confirm the suggestion that ADP was not responsible for the PAF-induced aggregation, reagents which would consume or destroy any ADP which was released were employed (20)(21)(22)(23)(24)(25)(26)(27)(28)(29)(30). Fig.…”

Section: Paf-induced Aggregationmentioning

confidence: 97%

“…Agents which destroy or consume ADP have been used by a number of investigators to show that the aggregation induced by many stimuli, including collagen, or even thrombin, may result in part from the release of platelet ADP, which then causes the aggregation (20)(21)(22)(23). In addition, the recent discovery of potent pharmacologic intermediates of the prostaglandin synthesis pathway, has suggested that these may represent the common platelet-activating material and that many stimuli activate by initiating prostaglandin synthesis (24)(25)(26).…”

Section: Introductionmentioning

confidence: 99%

Activation of Rabbit Platelets by Platelet-Activating Factor Derived from IgE-Sensitized Basophils

Henson¹

1977

A B S T R A C T Platelet-activating factor (PAF) is liberated from antigen-stimulated, IgE-sensitized rabbit basophils and induces aggregation of platelets and secretion of their content of vasoactive amines. Experiments were performed to determine the relationship between these two platelet responses to this stimulus.(a) Aggregation was initiated by appreciably lower concentrations of PAF than were required to produce even minimal release of constituents. PAF-induced aggregation was also resistant to agents which destroy ADP, such as creatine phosphate/creatine phosphokinase or apyrase, and occurred with platelets made unresponsive (refractory) to ADP. It was concluded that PAF can induce aggregation by a mechanism which is distinguishable from the release reaction and from the aggregating effect of ADP.(b) Secretion can probably occur independently of aggregation because incubation without agitation resulted in secretion without detectable aggregation.(c) Despite the suggestion that the two platelet responses can be independent, they may be induced by the same stimulus. This was indicated by experiments in which platelets specifically desensitized to PAF-induced secretion were also found to be unresponsive to PAF-stimulated aggregation. Moreover, identical levels of inhibition and identical inhibition profiles were obtained for both responses with serine esterase inhibitor, diisopropylphosphofluoridate, and with amino acid esters. It was concluded that the same stimulus-specific activable protease (esterase) was most likely involved in PAF-induced aggregation and secretion.