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Although amrinone produces thrombocytopenia, no information is available regarding the acute effects of milrinone on platelets. Therefore, we evaluated the effects of milrinone on platelet number and function in cardiac surgical patients. Twenty-seven patients were studied during cardiac surgery requiring cardiopulmonary bypass (CPB). Patients were randomized to receive no milrinone (n = 10), or milrinone (n = 17) at a loading dose of 50-75 micrograms/kg in the CPB circuit followed by 0.5-0.75 micrograms.kg-1.min-1 for 12-24 h. Bleeding times and blood samples for coagulation studies were obtained prior to induction, and at 2 and 24 h after CPB. In both groups, platelet counts decreased significantly from the baseline at 2 and 24 h after CPB, and bleeding time increased significantly from the baseline at 2 and 24 h after CPB. No significant thromboelastoplasty (TEG) changes were observed in either group, and there were no significant differences in platelet aggregation or chest tube drainage between the groups. Acute milrinone administration did not cause significant changes in platelet number or function in patients undergoing cardiac operations requiring CPB, beyond the usual adverse effects of cardiac surgery and CPB.

After incubation with [1-14C]-arachidonic acid, washed platelets from selenium deficient rats produced a sevenfold greater amount of 12-hydroperoxytetraenoic acid than platelets from control animals. When stimulated with either arachidonic acid or t-butyl-hydroperoxide, antimycin-A1 treated platelets from the deficient rats also converted markedly lower amounts of [1-14C]-glucose to [14C]-CO2 than platelets from control rats. These results indicate a significant role for platelet selenium-dependent glutathione peroxidase in the enzymatic reduction of platelet-produced hydroperoxides.

Oral administration of onion and garlic reportedly decreases platelet aggregation in both human and animal subjects. An oily chloroform extract of onion (Allium Cepa) was prepared and separated by column chromatography on silicic acid into six fractions by elution with solvents of increasing polarity. The least polar fraction contained most of the inhibitory activity towards platelet aggregation induced by either ADP or arachidonic acid. Further purification was afforded by thin-layer chromatography. The specific activity of this major active fraction (I50 per ml of PRP) was approximately 7 units per milligram. Platelets incubated in the presence of onion inhibitor and (1-14C)-arachidonic acid showed striking changes in the pattern of arachidonic acid metabolites formed. Thromboxane B2 synthesis was almost completely suppressed without significant decreases in total hydroxy fatty acid formation. It was concluded that the observed antiplatelet activity of onion relates to the presence of a non-polar, heat stable inhibitor of thromboxane synthesis. This appears to be the first demonstration of this type of inhibitor present in significant quantities in a common foodstuff.

Washed human platelets incubated with 1-14C-arachidonic acid (1mM) produced a new metabolite which migrated on thin layer chromatography close to thromboxane B2, but which was identified by mass spectrometry as a trihydroxy fatty acid. The mass spectrum was consistent with the assigned structure, 8,11,12-trihydroxy-5,9,14-eicosatrienoic acid (THETE). Platelet THETE synthesis from arachidonate was not inhibited by preincubation with asprin or indomethacin but was blocked by 5,8,11,14-eicosatetraynoic acid. Therefore, THETE appears to arise via the platelet lipoxygenase pathway rather than via the prostaglandin cyclooxygenase. Two proposed structures, including a novel dihydro-hydroxy-pyran cyclic intermediate, which could give rise to THETE are presented.

Feverfew has been used since antiquity to treat fevers and other inflammatory conditions. Feverfew extracts were found to inhibit ADP, thrombin, or collagen-induced aggregation of human platelets, but significantly, did not affect aggregation induced by arachidonic acid. Synthesis of thromboxane B2 from exogenous 14C-arachidonic acid was also not inhibited. Washed platelets prelabelled with 14C-AA responded normally to thrombin by releasing 14C-TXB2. This was completely blocked by feverfew. A purified platelet phospholipase A2 was inhibited by the material with an I50 of 0.1 antiplatelet units. The pharmacological properties of feverfew may thus be due to an inhibitor of cellular phospholipases, which prevents release of arachidonic acid in response to appropriate physiological stimuli.

Human platelets metabolize arachidonic acid via cyclooxygenase (E.C. 1.14.99.1) to thromboxane A2 and the 12-lipoxygenase to 12-hydroxyeicosatetraenoic acid (12-HETE). Aspirin inhibits cyclooxygenase while the neutrophil product 15-Hydroxyeicosatetraenoic acid (15-HETE) is a selective inhibitor of platelet 12-lipoxygenase. The unexpected observation was made that the platelet 12-lipoxygenase of individuals who had ingested aspirin showed up to a twenty-fold increase in sensitivity to inhibition by 15-HETE. This observation was confirmed in platelets treated with aspirin in vitro. Aspirin pretreatment consistently resulted in a decrease in the I50 for 15-HETE from an average of 21.5 microM to only 5.2 +/- 1.5 microM, indicating a probable interaction between the cyclooxygenase and lipoxygenase pathways.

During allergic and inflammatory reactions, arachidonic acid is oxidized by lipoxygenases to a variety of biologically active products, including leukotrienes. The mechanisms for regulation of the different lipoxygenase activities are not well defined. We report here that [14C]arachidonic acid metabolism by the 5- and 15-lipoxygenase activities in rabbit leukocytes and the 12-lipoxygenase in rabbit platelets is inhibited by various hydroxyeicosatetraenoic acids (HETEs). 15-HETE was the most effective inhibitor of the 5- and 12-lipoxygenases, whereas similar inhibitory potencies were observed for 5-HETE and 12-HETE acting on the 15-lipoxygenase. These three enzyme pathways were all least sensitive to their own products HETEs. To determine which structural characteristics of 15-HETE are essential for inhibition of the 5-lipoxygenase, various derivatives were prepared and purified by high pressure liquid chromatography, and their structures were confirmed by gas chromatography-mass spectrometry. The inhibitory potencies of 15-HETE analogs with different degrees of unsaturation were in the order of three double bonds greater than 4 greater than 2 greater than 0. 15-Hydroperoxy-5,8,11,13-eicosatetraenoic acid (15-HPETE) was four times more potent than 15-HETE. The 15-acetoxy, 15-keto and methyl ester derivatives were of comparable activity to 15-HETE, and the 15-acetoxy methyl ester derivative was less potent. Based upon the observed patterns of inhibition, we postulate that complex interregulatory relationships exist between the various lipoxygenases, and that cells containing these lipoxygenases may interact with each other via their lipoxygenase metabolites.

The functional interrelationships between the cyclooxygenase and lipoxygenase systems for arachidonic acid metabolism in platelets have not yet been clarified. Although a number of specific inhibitors of the cyclooxygenase, such as aspirin and indomethacin, have been described, few inhibitors of the lipoxygenase have been found. Several hydroxy and hydroperoxy derivatives of arachidonic acid were prepared and purified by high performance liquid chromatography, and their structures were confirmed by gas chromatography-mass spectrometry. The effect of these compounds on oxygenation of [1-14C]arachidonic acid by platelet cyclooxygenase and lipoxygenase enzymes was monitored both with an oxygen electrode and by analysis of radioactive products formed. It was found that 15-hydroxy-5,8,11,13-eicosatetraenoic acid selectively inhibited platelet lipoxygenase activity at micromolar concentrations (I50 = 8 microM) without inhibiting the cyclooxygenase. The 15-hydroperoxy analog, although more potent, was less selective, whereas neither 12-hydroxy-5,8,10,14-eicosatetraenoic acid nor 12-hydroxy-9-octadecenoic acid (ricinoleic acid) appeared to affect either enzyme. The high degree of selectivity of the 15-hydroxy derivative of eicosatetraenoic acid makes it the most suitable inhibitor so far discovered for studying the functions of the platelet lipoxygenase system.