prostaglandin-h2 and 1-benzylimidazole

Bovine lung thromboxane synthase was immobilized on phenyl-Sepharose beads by adsorption. The immobilized enzyme was catalytically active and synthesized both TXA2 and HHT. The production of both products was inhibited by 1-benzylimidazole and furegrelate. Multiple additions of PGH2 dramatically reduced the ability of the enzyme to synthesize TXA2, but did not effect the synthesis of HHT. In addition, 1-benzylimidazole did not protect thromboxane synthase from inactivation with multiple additions of PGH2. When the enzyme was incubated with PGH2 in the presence of 1-benzylimidazole, the synthesis of TXA2 was inhibited. When the inhibitor was removed the enzyme had still been inactivated by PGH2 in the presence of 1-benzylimidazole. Thus the substrate inactivation of the enzyme does not require the production of TXA2. Our data suggests that the synthesis of TXA2 and HHT can be differentially inactivated and may occur at different sites on the enzyme.

Topics: Animals; Cattle; Enzyme Activation; Imidazoles; Lung; Microsomes; Prostaglandin Endoperoxides, Synthetic; Prostaglandin H2; Prostaglandins H; Substrate Specificity; Thromboxane A2; Thromboxane-A Synthase

To test the hypothesis that prostacyclin (PGI2) is formed via a biochemical interaction between platelets and lymphocytes, we measured eicosanoids by high performance liquid chromatography (HPLC) and radioimmunoassay (RIA). A distinct 6-keto-prostaglandin F1 alpha (6KPGF1 alpha) peak was noted when [14C]arachidonic acid ([14C]AA) was added to the mixed cell preparations which was increased by pretreating platelets with 1-benzylimidazole (1-BI). Lymphocytes prelabeled with [14C]AA failed to form 6KPGF1 alpha when stimulated with phytohemagglutinin (PHA) or ionophore A23187. When the prelabeled platelets were suspended together with aspirin-treated lymphocytes and stimulated with ionophore, thrombin, or collagen, a 6KPGF1 alpha peak was detected and enhanced by 1-BI. These results were supported by quantifying the 6KPGF1 alpha content in the HPLC-purified fraction by RIA. Adding prostaglandin H2 (PGH2) directly to lymphocytes led to 6KPGF1 alpha production. Platelet aggregation and release were inhibited by lymphocytes in a dose-related manner. We conclude that lymphocytes possess PGI2 synthase activity which is capable of converting platelet-derived PGH2 into PGI2. PGI2 formed is sufficient to inhibit platelet function.

Topics: 6-Ketoprostaglandin F1 alpha; Arachidonic Acid; Arachidonic Acids; Blood Platelets; Epoprostenol; Humans; Imidazoles; Lymphocytes; Platelet Aggregation; Prostaglandin Endoperoxides, Synthetic; Prostaglandin H2; Prostaglandins H; Serotonin

Production of eicosanoids by deendothelialized aorta in response to continuous infusions of arachidonic acid and platelet suspensions was determined in a rabbit aorta perfusion model. 6-keto-PGF1 alpha production was stimulated by AA infusion in a dose-related manner. Infusion of AA at 4 micrograms/ml/min led to an initial production rate of 0.64 +/- 0.29 ng/min which gradually increased to 0.93 +/- 0.11 ng/min at the 20th min of infusion. When the concentration of AA infusion was increased to 10 micrograms/ml/min, 6-keto-PGF1 alpha production increased to 1.14 +/- 0.86 ng/min initially but declined with time. PGE2 production in response to AA 10 micrograms/min/ml was steady at around 5 ng/min while PGF2 alpha and TXB2 production were only slightly above the control. Perfusion of rabbit washed platelet suspensions at a rate of 3 X 10(8) plt/ml/min raised 6KPGF1 alpha production. The production was further increased when platelets were pretreated with 1-benzylimidazole (5 mM), along with a concurrent reduction in TXB2 release. Pretreatment of platelets with aspirin, on the other hand, abolished the increase in 6KPGF1 alpha production. Our data indicated that the vascular smooth muscle cells can efficiently utilize PGH2 produced by platelets to synthesize PGI2.

Topics: Animals; Aorta; Blood Platelets; Blood Vessels; Dinoprost; Dinoprostone; Dose-Response Relationship, Drug; Eicosanoic Acids; Endothelium; Epoprostenol; Imidazoles; In Vitro Techniques; Perfusion; Platelet Transfusion; Prostaglandin Endoperoxides, Synthetic; Prostaglandin H2; Prostaglandins E; Prostaglandins F; Prostaglandins H; Rabbits; Thromboxane B2