Esterified eicosanoids are acutely generated by 5-lipoxygenase in primary human neutrophils and in human and murine infection

5-Lipoxygenase (5-LOX) plays key roles in infection and allergic responses. Herein, four 5-LOX derived lipids comprising 5-hydroxyeicosatetraenoic acid (HETE) attached to phospholipids (PL), either phosphatidylethanolamine (PE) or phosphatidylcholine (PC) (18:0p/5-HETE-PE, 18:1p/5-HETE-PE, 16:0p/5-HETE-PE and 16:0a/5-HETE-PC), were identified in primary human neutrophils. They formed within 2 min in response to serum-opsonized Staphylococcus epidermidis (S. epidermidis) or fMLP, with priming by LPS, GM-CSF or cytochalasin D. Levels generated were similar to free 5-HETE (0.37 ± 0.14 ng vs 0.55 ± 0.18 ng/106 cells, esterified vs. free 5-HETE, respectively). They remained cell-associated, localizing to nuclear and extra-nuclear membrane, and were formed by fast esterification of newly synthesized free 5-HETE. Generation also required Ca2+, PLC, cPLA2, sPLA2, 5-LOX activating protein (FLAP) and MEK1. 5-HETE-PLs were detected in murine S. epidermidis peritonitis, paralleling neutrophil influx, and in effluent from Gram+ve human bacterial peritonitis. Formation of neutrophil extracellular traps (NETs) was significantly enhanced by 5-LOX inhibition, but attenuated by HETE-PE, while 5-HETE-PE enhanced superoxide and IL-8 generation. Thus, new molecular species of oxidized phospholipid formed by human neutrophils during bacterial infection are identified and characterized