Annexin I and dexamethasone effects on phospholipase and cyclooxygenase activity in human synoviocytes

Abstract

Annexin I is a glucocorticoid-induced mediator with anti-inflammatory activity in animal models of arthritis. We studied the effects of a bioactive annexin I peptide, ac 2–26, dexamethasone (DEX), and interleukin-1β (IL-1β) on phospholipase A₂ (PLA₂) and cyclooxygenase (COX) activities and prostaglandin E₂ (PGE₂) release in cultured human fibroblast-like synoviocytes (FLS). Annexin I binding sites on human osteoarthritic (OA) FLS were detected by ligand binding flow cytometry. PLA₂ activity was measured using ³H-arachidonic acid release, PGE₂ release and COX activity by ELISA, and COX2 content by flow cytometry. Annexin I binding sites were present on human OA FLS. Annexin I peptide ac 2–26 exerted a significant concentration-dependent inhibition of FLS constitutive PLA₂ activity, which was reversed by IL-1β. In contrast, DEX inhibited IL-1β-induced PLA₂ activity but not constitutive activity. DEX but not annexin I peptide inhibited IL-1β-induced PGE₂ release. COX activity and COX2 expression were significantly increased by IL-1β. Annexin I peptide demonstrated no inhibition of constitutive or IL-1β-induced COX activity. DEX exerted a concentration-dependent inhibition of IL-1β-induced but not constitutive COX activity. Uncoupling of inhibition of PLA₂ and COX by annexin I and DEX support the hypothesis that COX is rate-limiting for PGE₂ synthesis in FLS. The effect of annexin I but not DEX on constitutive PLA₂ activity suggests a glucocorticoid-independent role for annexin I in autoregulation of arachidonic acid production. The lack of effect of annexin I on cytokine-induced PGE₂ production suggests PGE₂-independent mechanisms for the anti-inflammatory effects of annexin I in vivo.