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In recent years, several studies have demonstrated that polyunsaturated fatty acids have strong immunomodulatory properties, altering several functions of macrophages. In the present work, we sought to provide a multi-omic approach combining the analysis of the lipidome, the proteome, and the metabolome of RAW 264.7 macrophages supplemented with phospholipids containing omega-3 (PC 18:0/22:6; ω3-PC) or omega-6 (PC 18:0/20:4; ω6-PC) fatty acids, alone and in the presence of lipopolysaccharide (LPS). Supplementation of macrophages with ω3 and ω6 phospholipids plus LPS produced a significant reprogramming of the proteome of macrophages and amplified the immune response; it also promoted the expression of anti-inflammatory proteins (e.g., pleckstrin). Supplementation with the ω3-PC and ω6-PC induced significant changes in the lipidome, with a marked increase in lipid species linked to the inflammatory response, attributed to several pro-inflammatory signalling pathways (e.g., LPCs) but also to the pro-resolving effect of inflammation (e.g., PIs). Finally, the metabolomic analysis demonstrated that supplementation with ω3-PC and ω6-PC induced the expression of several metabolites with a pronounced inflammatory and anti-inflammatory effect (e.g., succinate). Overall, our data show that supplementation of macrophages with ω3-PC and ω6-PC effectively modulates the lipidome, proteome, and metabolome of these immune cells, affecting several metabolic pathways involved in the immune response that are triggered by inflammation.

international journal of molecular sciences

Multi-Omic Profiling of Macrophages Treated with Phospholipids Containing Omega-3 and Omega-6 Fatty Acids Reveals Complex Immunomodulatory Adaptations at Protein, Lipid and Metabolic Levels