Passage state affects arachidonic acid content and eicosanoid release in porcine aortic endothelial cells

Porcine aortic endothelial cells were cultured through four passages from primary cultures. The arachidonic acid content of individual phospholipid classes and the release of 6-keto-prostaglandin F1 alpha and 15-hydroxyeicosatetraenoic acid in response to 1 microM ionophore A23187 were assayed at each passage. The content of arachidonic acid in phosphatidylinositol and diacyl phosphatidylethanolamine remained constant at passage 1 but declined at passage 2 by approximately 29% and at passage 4 by approximately 59%. The release of 6-keto-prostaglandin F1 alpha was also unchanged at passage 1 but decreased by 60% at passage 2 and by 82% from its original value at passage 4. In contrast, the arachidonic acid content of diacyl phosphatidylcholine and of alkenyl phosphatidylethanolamine decreased with each passage, by 34% at passage 1, 59% at passage 2, 71% at passage 3, and 76% of the original value at passage 4. Stimulation with arachidonic acid reversed the passage effect. The release of 15-hydroxyeicosatetraenoic acid decreased by 82% at passage 1 and diminished to a 97% decrement from the original value by passage 4. When stimulated with arachidonic acid, 15-hydroxyeicosatetraenoic acid steadily decreased by approximately 70% at passages 3 and 4. The data indicate that passage state strikingly and nonuniformly affects phospholipid class arachidonic acid content and eicosanoid release in response to agonist stimulation.