Occurrence and biological activity of palmitoleic acid isomers in phagocytic cells.

Recent studies have highlighted the role of palmitoleic acid [16:1n-7 (cis-9-hexadecenoic acid)] as a lipid hormone that coordinates cross-talk between liver and adipose tissue and exerts anti-inflammatory protective effects on hepatic steatosis and insulin signaling in murine models of metabolic disease. More recently, a 16:1n-7 isomer, cis-7-hexadecenoic acid (16:1n-9), that also possesses marked anti-inflammatory effects, has been described in human circulating monocytes and monocyte-derived macrophages. By using gas chromatographic/mass spectrometric analyses of dimethyl disulfide derivatives of fatty acyl methyl esters, we describe in this study the presence of a third 16:1 isomer, sapienic acid [16:1n-10 (6-cis-hexadecenoic acid)], in phagocytic cells. Cellular levels of 16:1n-10 appear to depend not only on the cellular content of linoleic acid, but also on the expression level of fatty acid desaturase 2, thus revealing a complex regulation both at the enzyme level, via fatty acid substrate competition, and directly at the gene level. However, unlike 16:1n-7 and 16:1n-9, 16:1n-10 levels are not regulated by the activation state of the cell. Moreover, while 16:1n-7 and 16:1n-9 manifest strong anti-inflammatory activity when added to the cells at low concentrations (10 µM), notably higher concentrations of 16:1n-10 are required to observe a comparable effect. Collectively, these results suggest the presence in phagocytic cells of an unexpected variety of 16:1 isomers, which can be distinguished on the basis of their biological activity and cellular regulation.

Cellular Plasmalogen Content Does Not Influence Arachidonic Acid Levels or Distribution in Macrophages: A Role for Cytosolic Phospholipase A2γ in Phospholipid Remodeling.

Availability of free arachidonic acid (AA) constitutes a rate limiting factor for cellular eicosanoid synthesis. AA distributes differentially across membrane phospholipids, which is largely due to the action of coenzyme A-independent transacylase (CoA-IT), an enzyme that moves the fatty acid primarily from diacyl phospholipid species to ether-containing species, particularly the ethanolamine plasmalogens. In this work, we examined the dependence of AA remodeling on plasmalogen content using the murine macrophage cell line RAW264.7 and its plasmalogen-deficient variants RAW.12 and RAW.108. All three strains remodeled AA between phospholipids with similar magnitude and kinetics, thus demonstrating that cellular plasmalogen content does not influence the process. Cell stimulation with yeast-derived zymosan also had no effect on AA remodeling, but incubating the cells in AA-rich media markedly slowed down the process. Further, knockdown of cytosolic-group IVC phospholipase A2γ (cPLA2γ) by RNA silencing significantly reduced AA remodeling, while inhibition of other major phospholipase A2 forms such as cytosolic phospholipase A2α, calcium-independent phospholipase A2ß, or secreted phospholipase A2 had no effect. These results uncover new regulatory features of CoA-IT-mediated transacylation reactions in cellular AA homeostasis and suggest a hitherto unrecognized role for cPLA2γ in maintaining membrane phospholipid composition via regulation of AA remodeling.

Essential Role for Ethanolamine Plasmalogen Hydrolysis in Bacterial Lipopolysaccharide Priming of Macrophages for Enhanced Arachidonic Acid Release.

Due to their high content in esterified arachidonic acid (AA), macrophages provide large amounts of eicosanoids during innate immune reactions. Bacterial lipopolysaccharide (LPS) is a poor trigger of AA mobilization in macrophages but does have the capacity to prime these cells for greatly increased AA release upon subsequent stimulation. In this work, we have studied molecular mechanisms underlying this phenomenon. By using mass spectrometry-based lipidomic analyses, we show in this work that LPS-primed zymosan-stimulated macrophages exhibit an elevated consumption of a particular phospholipid species, i.e., the ethanolamine plasmalogens, which results from reduced remodeling of phospholipids via coenzyme A-independent transacylation reactions. Importantly however, LPS-primed macrophages show no changes in their capacity to directly incorporate AA into phospholipids via CoA-dependent acylation reactions. The essential role for ethanolamine plasmalogen hydrolysis in LPS priming is further demonstrated by the use of plasmalogen-deficient cells. These cells, while responding normally to zymosan by releasing quantities of AA similar to those released by cells expressing normal plasmalogen levels under the same conditions, fail to show an LPS-primed response to the same stimulus, thus unambiguously demonstrating a cause-effect relationship between LPS priming and plasmalogen hydrolysis. Collectively, these results suggest a hitherto unrecognized role for ethanolamine plasmalogen hydrolysis and CoA-independent transacylation reactions in modulating the eicosanoid biosynthetic response.