Abstract
Human umbilical arteries convert arachidonic acid into three hydroxy-eicosatetraenoic acids as well as 6- ketoprostaglandin F 1 α , prostaglandins E 2, F 2 α and D 2 and thromboxane B 2. Two of these hydroxy derivatives of arachidonic acid were purified by reverse-phase HPLC and identified by GC-MS as 11-hydroxyeicosatetraenoic acid (11-HETE) and 15-hydroxyeicosatetraenoic acid (15-HETE) while a third, presumed dihydroxy derivative has not yet been identified. Both the cyclooxygenase and HETE synthesizing activities were found to be localized mainly in the microsomal fraction (100000 × g pellet) (51 and 61% of total, respectively), and approx. 25% of both activities was found in the 10000 × g pellet. The formation of these HETEs was inhibited by the cyclooxygenase inhibitors indomethacin and aspirin but not by the lipoxygenase inhibitor nordihydroguaiaretic acid. Production of immunoreactive 15-HETE as well as 6-ketoprostaglandin F 1α were also decreased significantly when arterial segments were incubated in the presence of either indomethacin or aspirin. Indomethacin inhibited the formation of both prostanoids and HETEs by microsomes in a concentration-dependent and time-dependent manner. The ID 50, values for indomethacin against HETE synthesizing activity and against cyclooxygenase were 4.5 and 3.8 μM, respectively. The inactivation constants were found to be 0.09 and 0.08 min −1 for HETE synthesizing activity and cyclooxygenase, respectively. These two microsomal activities were solubilized in parallel with Tween-20. Incubation with three distinct monoclonal antibodies against different epitopes on cyclooxygenase precipitated both cyclooxygenase and HETE synthesizing activity. Each of these activities was recovered in the immune pellets. These studies demonstrate that in human umbilical arteries 11-HETE, 15-HETE and a presumed di-HETE are the products of cyclooxygenase.
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Schedule a callMore From: Biochimica et Biophysica Acta (BBA) - Lipids and Lipid Metabolism
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