Docosahexaenoic Acid Decreases Pro-Inflammatory Mediators in an In Vitro Murine Adipocyte Macrophage Co-Culture Model

Abstract

Paracrine interactions between adipocytes and macrophages contribute to chronic inflammation in obese adipose tissue. Dietary strategies to mitigate such inflammation include long-chain polyunsaturated fatty acids, docosahexaenoic (DHA) and eicosapentaenoic (EPA) acids, which act through PPARγ-dependent and independent pathways. We utilized an in vitro co-culture model designed to mimic the ratio of macrophages:adipocytes in obese adipose tissue, whereby murine 3T3-L1 adipocytes were cultured with RAW 264.7 macrophages in direct contact, or separated by a trans-well membrane (contact-independent mechanism), with 125 µM of albumin-complexed DHA, EPA, palmitic acid (PA), or albumin alone (control). Thus, we studied the effect of physical cell contact versus the presence of soluble factors, with or without a PPARγ antagonist (T0070907) in order to elucidate putative mechanisms. After 12 hr, DHA was the most anti-inflammatory, decreasing MCP1 and IL-6 secretion in the contact system (−57%, −63%, respectively, p≤0.05) with similar effects in the trans-well system. The trans-well system allowed for isolation of cell types for inflammatory mediator analysis. DHA decreased mRNA expression (p<0.05) of Mcp1 (−7.1 fold) and increased expression of the negative regulator, Mcp1-IP (+1.5 fold). In macrophages, DHA decreased mRNA expression of pro-inflammatory M1 polarization markers (p≤0.05), Nos2 (iNOS; −7 fold), Tnfα (−4.2 fold) and Nfκb (−2.3 fold), while increasing anti-inflammatory Tgfβ1 (+1.7 fold). Interestingly, the PPARγ antagonist co-administered with DHA or EPA in co-culture reduced (p≤0.05) adiponectin cellular protein, without modulating other cytokines (protein or mRNA). Overall, our findings suggest that DHA may lessen the degree of MCP1 and IL-6 secreted from adipocytes, and may reduce the degree of M1 polarization of macrophages recruited to adipose tissue, thereby decreasing the intensity of pro-inflammatory cross-talk between adipocytes and macrophages in obese adipose tissue.

Effect of dietary Schizochytrium microalga oil on selected markers of low‐grade inflammation in rats

“…d) agrees with conclusion of De Boer et al. () that transcriptionally active NF‐ κ B subunit may not be a useful endpoint to measure the inflammatory status of cells.…”

A decade of progress in adipose tissue macrophage biology

“…The fish oil‐derived eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) strongly promote the alternative activation of macrophages. In vitro treatment of macrophages with EPA and DHA results in an M2‐like phenotype, exhibiting reduced pro‐inflammatory gene expression and increased anti‐inflammatory gene expression . We and others have shown that supplementation of mice with dietary fish oils reduced AT expression of pro‐inflammatory genes while increasing adiponectin expression.…”

Dietary Fatty Acids Differentially Regulate Secretion of Adiponectin and Interleukin‐6 in Primary Canine Adipose Tissue Culture

“…It is likely that the FA affected the secretion of IL6 by resident macrophages within the undifferentiated SVC used in this study (Fain, ). PAM has also been shown to induce secretion of IL6 from murine adipocytes (Chakrabarti et al, ) and adipocyte cell line (Ajuwon & Spurlock, ) as well as human endothelial cells (Staiger et al, ), while DHA decreased IL6 secretion from human subcutaneous adipocytes differentiated in culture (Bjork et al, ) and from 3T3L‐1 cells (De Boer, Monk, & Robinson, ). It is possible that PAM and EPA affected secretion of IL6 from preadipocytes and endothelial cells within the undifferentiated SVC in this study as well, contributing to the overall secretion of IL6 from the SVC in this study.…”