Abstract
Polyunsaturated fatty acids (PUFAs) are important for immune function. Limited evidence indicates that immune cell activation involves endogenous PUFA synthesis, but this has not been characterised. To address this, we measured metabolism of 18:3n-3 in quiescent and activated peripheral blood mononuclear cells (PBMCs), and in Jurkat T cell leukaemia. PBMCs from men and women (n = 34) were incubated with [1-13C]18:3n-3 with or without Concanavalin A (Con. A). 18:3n-3 conversion was undetectable in unstimulated PBMCs, but up-regulated when stimulated. The main products were 20:3n-3 and 20:4n-3, while 18:4n-3 was undetectable, suggesting initial elongation and Δ8 desaturation. PUFA synthesis was 17.4-fold greater in Jurkat cells than PBMCs. The major products of 18:3n-3 conversion in Jurkat cells were 20:4n-3, 20:5n-3, and 22:5n-3. 13C Enrichment of 18:4n-3 and 20:3n-3 suggests parallel initial elongation and Δ6 desaturation. The FADS2 inhibitor SC26196 reduced PBMC, but not Jurkat cell, proliferation suggesting PUFA synthesis is involved in regulating mitosis in PBMCs. Con. A stimulation increased FADS2, FADS1, ELOVL5 and ELOVL4 mRNA expression in PBMCs. A single transcript corresponding to the major isoform of FADS2, FADS20001, was detected in PBMCs and Jurkat cells. PBMC activation induced hypermethylation of a 470bp region in the FADS2 5′-regulatory sequence. This region was hypomethylated in Jurkat cells compared to quiescent PBMCs. These findings show that PUFA synthesis involving initial elongation and Δ8 desaturation is involved in regulating PBMC proliferation and is regulated via transcription possibly by altered DNA methylation. These processes were dysregulated in Jurkat cells. This has implications for understanding the regulation of mitosis in normal and transformed lymphocytes.
Highlights
Polyunsaturated fatty acids (PUFAs) play key roles in the immune response by acting as substrates for the synthesis of lipid second messengers involved in cell activation, including eicosanoids and for cell membrane biosynthesis [1]
The present findings show that activation of peripheral blood mononuclear cells (PBMCs) induces increased conversion of 18:3n-3 to longer chain PUFA and that this involves increased transcription of key genes involved in this pathway and altered epigenetic regulation of FADS2
Enzyme activity of the PUFA biosynthesis pathway and the epigenetic regulation of FADS2 were dysregulated in Jurkat cells. n-3 PUFA biosynthesis appears to be involved in the control of proliferation of PBMCs, but not of Jurkat cells
Summary
Polyunsaturated fatty acids (PUFAs) play key roles in the immune response by acting as substrates for the synthesis of lipid second messengers involved in cell activation, including eicosanoids and for cell membrane biosynthesis [1]. T cell receptor-stimulated splenocytes from Fads null mice showed that genotype did not alter the effect dietary fatty acids on TNFα, IL-1β, IL-6, or IL-10 synthesis [12] which suggests that PUFA biosynthesis is not involved in the regulation of the production of these mediators. We characterised PUFA biosynthesis in human PBMCs by determining the effect of lectin stimulation on the metabolism of [1-13C]18:3n-3, and on the expression and epigenetic regulation of key genes in the PUFA synthesis pathway. We compared these findings with a spontaneously proliferating human T cell lymphoma cell line
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