Phospholipid Arachidonic Acid Remodeling During Phagocytosis in Mouse Peritoneal Macrophages

Abstract

Macrophages contain large amounts of arachidonic acid (AA), which distributes differentially across membrane phospholipids. This is largely due to the action of coenzyme A-independent transacylase (CoA-IT), which transfers the AA primarily from diacyl choline-containing phospholipids to ethanolamine-containing phospholipids. In this work we have comparatively analyzed glycerophospholipid changes leading to AA mobilization in mouse peritoneal macrophages responding to either zymosan or serum-opsonized zymosan (OpZ). These two phagocytic stimuli promote the cytosolic phospholipase A2-dependent mobilization of AA by activating distinct surface receptors. Application of mass spectrometry-based lipid profiling to identify changes in AA-containing phospholipids during macrophage exposure to both stimuli revealed significant decreases in the levels of all major choline phospholipid molecular species and a major phosphatidylinositol species. Importantly, while no changes in ethanolamine phospholipid species were detected on stimulation with zymosan, significant decreases in these species were observed when OpZ was used. Analyses of CoA-IT-mediated AA remodeling revealed that the process occurred faster in the zymosan-stimulated cells compared with OpZ-stimulated cells. Pharmacological inhibition of CoA-IT strongly blunted AA release in response to zymosan but had only a moderate effect on the OpZ-mediated response. These results suggest a hitherto undescribed receptor-dependent role for CoA-independent AA remodeling reactions in modulating the eicosanoid biosynthetic response of macrophages. Our data help define novel targets within the AA remodeling pathway with potential use to control lipid mediator formation

Choline Glycerophospholipid-Derived Prostaglandins Attenuate TNFα Gene Expression in Macrophages via a cPLA2α/COX-1 Pathway

“…These cells have been found to contain high amounts of AA in membrane phospholipids; 20–25% of total fatty acid content is AA [ 71 , 72 ]. Importantly, macrophages also exhibit a characteristic distribution of AA among phospholipids, with PE, not PC, constituting the richest AA containing class, and PI containing much lesser amounts than PE or PC [ 18 , 21 ]. Among molecular species, the ethanolamine plasmalogens are markedly enriched with AA [ 56 , 73 ].…”

“…In mouse resident peritoneal macrophages, PE constitutes the major AA-containing class (45–50% of total cellular AA), followed by PC (35–40%), and PI (5–15%) [ 18 , 21 ]. Note that the 3 H/ 14 C ratio for extracellular free AA is considerably higher than that of the prostaglandins, thus suggesting a substantial contribution of PE, in addition to PC and PI, to AA release.…”

“…We demonstrated the involvement of several phospholipase A 2 forms, each acting on distinct phospholipid pools and releasing different fatty acids [ 15 , 18 , 19 ]. Also, we showed the importance of coenzyme A-independent transacylation reactions to shape the cellular AA pools and their involvement in some macrophage responses [ 20 , 21 , 22 ]. In the current work we have utilized advanced mass spectrometry-based lipidomic approaches to characterize the immediate generation of PG and time-dependent changes of AA-containing phospholipid species in zymosan-stimulated mouse peritoneal macrophages.…”

The Roles of Phospholipase A2 in Phagocytes

“…Eicosanoids are a broad family of oxygenated lipid compounds that include prostaglandins, thromboxanes, and leukotrienes. Eicosanoids are generated via non-enzymatic oxidation of ARA or by the action of enzymes, such as cyclooxygenase (COX), lipooxygenase (LOX), and cytochrome P450 (CYP) ( Dennis and Norris, 2015 ; Gil-de-Gómez et al, 2020 ). Eicosanoids exert immunomodulatory functions and, depending on the species of eicosanoids, can have pro- or anti-inflammatory effects ( Dennis and Norris, 2015 ).…”

The Impact of the Ca2+-Independent Phospholipase A2β (iPLA2β) on Immune Cells

“…The inhibition of secretory PLA 2 s (sPLA 2 s) GIIA, GV, and GX or cPLA 2 α did not alter the classically activated macrophage production of eicosanoids, suggesting a select role for iDLs on macrophage polarization. However, other groups have reported a role for sPLA 2 or cPLA 2 α in eicosanoid production from macrophages [ 142 , 143 , 144 , 145 , 146 , 147 ]. Those studies were performed with macrophage-like cells (RAW264.7 and P388D1) or peritoneal macrophages from rats and mice, with only LPS (10-1000 ng/mL) or zymosan stimulation for 6–48 h, and the involvement of various PLA 2 s was discerned with chemical inhibitors, some of which effected multiple PLA 2 s. In contrast, our studies were performed with primary mouse peritoneal macrophages, which have been demonstrated to behave differently from the macrophage-like cells [ 36 ], from wild-type and genetically- modified iPLA 2 β-deficient mice, and importantly, treated with LPS+IFNγ, conditions that induce a M1 macrophage phenotype.…”