Polyunsaturated Fatty Acid Biosynthesis Involving Δ8 Desaturation and Differential DNA Methylation of FADS2 Regulates Proliferation of Human Peripheral Blood Mononuclear Cells

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.

How does polyunsaturated fatty acid biosynthesis regulate T‐lymphocyte function?

“…Elongase (ELOVL) 2, which catalyses conversion of 22:5 n ‐3 to 24:5 n ‐3, which precedes 22:6 n ‐3 synthesis, was not detected (Sibbons et al . ). This is consistent with the lack of detectable enrichment of 22:6 n ‐3 in stimulated cells and suggests truncation of the pathway after 22:5 n ‐3.…”

“…The findings also showed for the first time that treatment with SC26196 decreased both the proportion of lymphocytes that underwent division and the mean number of cell divisions of the original cell population by approximately 10% each (Sibbons et al . ). The partial reduction in proliferation is consistent with the partial inhibition of PUFA synthesis.…”

“…Stimulation of PBMCs also induced a significant increase in methylation of twenty CpG loci in a 611 bp region between 667 and 1278 bp upstream of the FADS2 transcription start site (Sibbons et al . ). DNA methylation is typically associated with suppression of transcription (Kovesdi et al .…”

“…The fatty acid 22:5 n ‐3 did not accumulate in activated cells (Sibbons et al . ), which suggests further conversion to an unidentified metabolite. As expected, treatment of mitogen‐stimulated PBMCs with the Δ6 desaturase inhibitor SC26196 increased the level of 20:3 n ‐3, but reduced the amount of 20:4 n ‐3.…”

“…Alternative pathways of n ‐3 fatty acid synthesis in activated T cells [adapted from our published work in peripheral blood mononuclear cells: Sibbons et al . ()]; the asterisk indicates the main accumulated products of synthesis from 18:3 n ‐3, alpha‐linolenic acid, aLNA. For simplicity, the full names of not all individual fatty acids are given.…”