An Optimized Ex Vivo n–3 PUFA Supplementation Strategy for Primary Human Macrophages Shows That DHA Suppresses Prostaglandin E2 Formation

• Published in: Molecular nutrition & food research Vol. 69; no. 1; pp. e202400716 - n/a
• Main Authors: Kirchhoff, Rebecca, Kampschulte, Nadja, Rothweiler, Carina, Rohwer, Nadine, Weylandt, Karsten‐Henrich, Schebb, Nils Helge
• Format: Journal Article
• Language: English
• Published: Germany Wiley Subscription Services, Inc 01.01.2025; John Wiley and Sons Inc
• Subjects: Adult; Arachidonic acid; arachidonic acid cascade; blood; Cells, Cultured; cytokines; Cytokines - metabolism; Dietary Supplements; Dinoprostone - metabolism; Docosahexaenoic acid; Docosahexaenoic Acids - pharmacology; Fatty acids; Fatty Acids, Omega-3 - pharmacology; Female; food research; Humans; Immune system; inflammation; Inflammatory diseases; lipid mediators; Lipopolysaccharides - pharmacology; Macrophages; Macrophages - drug effects; Macrophages - metabolism; Male; nutrition; omega-3 fatty acids; omega-6 fatty acids; oxylipins; Oxylipins - pharmacology; Polyunsaturated fatty acids; Prostaglandin E2; Prostaglandins; Supplements; arachidonic acid cascade; lipid mediators; oxylipins; fatty acids; inflammation
• ISSN: 1613-4125; 1613-4133; 1613-4133
• DOI: 10.1002/mnfr.202400716

ABSTRACT Evidence suggests beneficial effects of long‐chain n–3 polyunsaturated fatty acids (PUFAs) in inflammatory diseases. However, the underlying mechanisms are still subject of research. For this purpose, we developed an ex vivo n–3 PUFA supplementation strategy. M2‐like macrophages were supplemented for 2–3 days with 20–40 µM docosahexaenoic acid (DHA) during differentiation. Quality parameters include <3% oxylipins for PUFA‐preparation, total fatty acids (FAs) <10 mM, and low oxylipins in plasma, n–3 PUFA <0.25 mM for the selection of donors of plasma as well as %n–6 in highly unsaturated fatty acids (HUFAs) ≥70% for donors of cells. Following supplementation, PUFA pattern of cells was shifted toward one described for blood and tissue from subjects with higher n–3 and lower n–6 PUFAs. This was accompanied by a decrease of arachidonic acid‐derived oxylipins in a dose‐ and time‐dependent manner in favor of n–3 PUFA ones. Stimulation with LPS resulted in decreased levels of pro‐inflammatory prostaglandins in the DHA‐supplemented cells, but no changes in cytokines. In vitro supplementation studies with n–3 PUFA need rigorous controls to exclude the background formation of oxylipins. By accounting for these possible confounders the described approach allows the mechanistic investigation of n–3 PUFAs in primary human immune cells, offering an alternative for intervention studies. This presented ex vivo n–3 polyunsaturated fatty acid (PUFA) supplementation strategy is an important tool for the mechanistic investigation of n–3 PUFA mediated effects in human immune cells. Fatty acid (FA) pattern from monocytes derived from subjects with a low n–3 PUFA status was shifted toward one reflecting the pattern of subjects with a high n–3 PUFA status, accompanied by remarkable changes in the oxylipin profile. Lower levels of pro‐inflammatory prostaglandins were found in the supplemented macrophages, which may be one of the major mechanisms of how the anti‐inflammatory effect of n–3 PUFA is mediated.