A functional role for eicosanoid-lysophospholipids in activating monocyte signaling.

Recently, eicosanoid-lysophospholipids were identified as novel metabolites generated from the direct cyclooxygenase- or lipoxygenase-catalyzed oxidation of 2-arachidonoyl-lysophospholipids produced from either phospholipase A-mediated hydrolysis of diacyl arachidonoyl-phospholipids or through the cytochrome -catalyzed oxidative hydrolysis of the vinyl ether linkage of arachidonoyl-plasmalogens. Although the metabolic pathways generating eicosanoid-lysophospholipids have been increasingly appreciated, the signaling functions of eicosanoid-lysophospholipids remain largely unknown. Herein, we demonstrate that 2-12()-HETE-lysophospholipids as well as nonesterified 12()-HETE are potent lipid mediators that activate THP-1 human monocytic cells to generate tumor necrosis factor α (TNFα) and interleukin 8 (IL8). Remarkably, low nanomolar concentrations of 12()-HETE-lysophospholipids, but not other oxidized signaling lipids examined activated THP-1 cells resulting in the production of large amounts of TNFα. Moreover, TNFα release induced by 12()-HETE-lysophospholipids was inhibited by the TNFα converting enzyme inhibitor TAPI-0 indicating normal processing of TNFα in THP-1 cells stimulated with these agonists. Western blotting analyses revealed that 12()-HETE-lysophospholipids activated the phosphorylation of NFκB p65, suggesting activation of the canonical NFκB signaling pathway. Importantly, activation of THP-1 cells to release TNFα was stereoselective with 12()-HETE favored over 12()-HETE. Furthermore, the EC of 2-12()-HETE-lysophosphatidylcholine in activating THP-1 cells was 2.1 nm, whereas the EC of free 12()-HETE was 23 nm Additionally, lipid extracts of activated platelets were separated by RP-HPLC demonstrating the coelution of 12()-HETE with fractions initiating TNFα release. Collectively, these results demonstrate the potent signaling properties of 2-12()-HETE-lysophospholipids and 12()-HETE by their ability to release TNFα and activate NFκB signaling thereby revealing a previously unknown role of 2-12()-HETE-lysophospholipids in mediating inflammatory responses.