5-Lipoxygenase activating protein (FLAP) dependent leukotriene biosynthesis inhibition (MK591) attenuates Lipid A endotoxin-induced inflammation.

Abstract

The Lipid A moiety of endotoxin potently activates TLR-4 dependent host innate immune responses. We demonstrate that Lipid-A mediated leukotriene biosynthesis regulates pathogen-associated molecular patterns (PAMP)-dependent macrophage activation. Stimulation of murine macrophages (RAW264.7) with E. coli 0111:B4 endotoxin (LPS) or Kdo2-lipid A (Lipid A) induced inflammation and Lipid A was sufficient to induce TLR-4 mediated macrophage inflammation and rapid ERK activation. The contribution of leukotriene biosynthesis was evaluated with a 5-lipoxygenase activating protein (FLAP) inhibitor, MK591. MK591 pre-treatment not only enhanced but also sustained ERK activation for up to 4 hours after LPS and Lipid A stimulation while inhibiting cell proliferation and enhancing cellular apoptosis. Leukotriene biosynthesis inhibition attenuated inflammation induced by either whole LPS or the Lipid A fraction. These responses were regulated by inhibition of the key biosynthesis enzymes for the proinflammatory eicosanoids, 5-lipoxygenase (5-LO), and cyclooxygenase-2 (COX-2) quantified by immunoblotting. Inhibition of leukotriene biosynthesis differentially regulated TLR-2 and TLR-4 cell surface expression assessed by flow cytometry, suggesting a close mechanistic association between TLR expression and 5-LO associated eicosanoid activity in activated macrophages. Furthermore, MK591 pre-treatment enhanced ERK activation and inhibited cell proliferation after LPS or Lipid A stimulation. These effects were regulated in part by increased apoptosis and modulation of cell surface TLR expression. Together, these data clarify the mechanistic association between 5-lipoxygenase activating protein-mediated leukotriene biosynthesis and 5-LO dependent eicosanoid metabolites in mediating the TLR-dependent inflammatory response after endotoxin exposure typical of bacterial sepsis.

• Authors: Wen-Feng Fang, Ivor S Douglas, Chin-Chou Wang, Hsu-Ching Kao, Ya-Ting Chang, Chia-Cheng Tseng, Kuo-Tung Huang, Huang-Chih Chang, Meng-Chih Lin
• Journal: PloS one (PLoS One) Vol. 9 Issue 7 Pg. e102622 ( 2014) ISSN: 1932-6203 [Electronic] United States
• PMID: 25025775 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• 5-Lipoxygenase-Activating Protein Inhibitors
• 5-Lipoxygenase-Activating Proteins
• Alox5ap protein, mouse
• Chemokine CXCL2
• Cxcl2 protein, mouse
• Indoles
• Leukotrienes
• Lipid A
• Quinolines
• Tlr2 protein, mouse
• Tlr4 protein, mouse
• Toll-Like Receptor 2
• Toll-Like Receptor 4
• MK 0591
• Arachidonate 5-Lipoxygenase

Topics

• 5-Lipoxygenase-Activating Protein Inhibitors (pharmacology)
• 5-Lipoxygenase-Activating Proteins (physiology)
• Animals
• Apoptosis
• Arachidonate 5-Lipoxygenase (biosynthesis)
• Cell Line
• Cell Proliferation
• Cell Survival (drug effects)
• Chemokine CXCL2 (metabolism)
• Indoles (pharmacology)
• Leukotrienes (biosynthesis)
• Lipid A (pharmacology)
• Macrophages (drug effects, immunology, metabolism)
• Mice
• Quinolines (pharmacology)
• Toll-Like Receptor 2 (metabolism)
• Toll-Like Receptor 4 (metabolism)