5-Lipoxygenase contributes to PPARγ activation in macrophages in response to apoptotic cells

Abstract

Macrophage polarization to an anti-inflammatory phenotype upon contact with apoptotic cells is a contributing hallmark to immune suppression during the late phase of sepsis. Although the peroxisome proliferator-activated receptor γ (PPARγ) supports this macrophage phenotype switch, it remains elusive how apoptotic cells activate PPARγ. Assuming that a molecule causing PPARγ activation in macrophages originates in the cell membrane of apoptotic cells we analyzed lipid rafts from apoptotic, necrotic, and living human Jurkat T cells which showed the presence of 5-lipoxygenase (5-LO) in lipid rafts of apoptotic cells only. Incubating macrophages with lipid rafts of apoptotic, but not necrotic or living cells, induced PPAR responsive element (PPRE)-driven mRuby reporter gene expression in RAW 264.7 macrophages stably transduced with a 4xPPRE containing vector. Experiments with lipid rafts of apoptotic murine EL4 T cells revealed similar results. To verify the involvement of 5-LO in activating PPARγ in macrophages, Jurkat T cells were incubated with the 5-LO inhibitor MK-866 prior to induction of apoptosis, which failed to induce mRuby expression. Similar results were obtained with lipid rafts of apoptotic EL4 T cells preexposed to the 5-LO inhibitors zileuton and CJ-13610. Interestingly, Jurkat T cells overexpressing 5-LO failed to activate PPARγ in macrophages, while their 5-LO overexpressing apoptotic counterparts did. Our results suggest that during apoptosis 5-LO gets associated with lipid rafts and synthesizes ligands that in turn stimulate PPARγ in macrophages.

Keywords: 5-LO; 5-LO activating protein; 5-Lipoxygenase; 5-lipoxygenase; AC; Alternative activation; CV; Efferocytosis; F5; FF; FLAP; LC; LR; Luc; M; Macrophages; MΦ; N-acetyl-cysteine; NAC; NC; NO; OE; PC; PPAR response element; PPARα/β/γ; PPRE; ROS; Sepsis; apoptotic cell(s); control virus; d/n; dominant negative; firefly; fraction 5; lipid raft(s); living cell(s); luciferase; macrophage(s); marker; methyl-ß-cyclodextrin; mßCDT; necrotic cell(s); nitric oxide; overexpression; peroxisome proliferator activating receptor alpha/delta/gamma; positive control; reactive oxygen species.