Dietary n-3 polyunsaturated fatty acids modulate purified murine T-cell subset activation

Abstract

Studies in humans and murine disease models have clearly shown dietary fish oil to possess anti-inflammatory properties, apparently mediated by the n-3 polyunsaturated fatty acids, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). To determine the mechanisms by which dietary EPA and DHA modulate mouse T-cell activation, female C57BL/6 mice were fed diets containing either 2% safflower oil (SAF), 2% fish oil (FO), or a 2% purified EPA/DHA ethyl ester mixture for 14 days. Splenic CD4 T cells ( approximately 90% purity) or CD8 T cells ( approximately 85% purity) were incubated with agonists which act at the plasma membrane receptor level [anti(alpha)-CD3/anti(alpha)-CD28], the intracellular level (PMA/Ionomycin), or at both the receptor and intracellular levels (alphaCD3/PMA). CD4 T cells stimulated with alphaCD3/alphaCD28 or PMA/Ionomycin proliferated and produced principally IL-2 (i.e. a Th1 phenotype), whereas the proliferation of CD4 T cells stimulated with alphaCD3/PMA was apparently driven principally by IL-4 (i.e. a Th2 phenotype). The IL-4 driven proliferation of putative Th2 CD4 cells was enhanced by dietary n-3 fatty acids (P = 0.02). Conversely, IL-2 production by alphaCD3/alpha CD28-stimulated CD4 T cells was reduced in FO-fed animals (P < 0.0001). The alphaCD3/alphaCD28-stimulated CD8 cells cultured from FO-fed animals exhibited a significant decrease (P < 0.05) in proliferation. There were no dietary effects seen in alphaCD3/PMA-stimulated CD8 cells, which produced both IL-2 and IL-4, or in PMA/Ionomycin-stimulated CD8 cells, which produced principally IL-2. These data suggest that dietary n-3 fatty acids down-regulated IL-2 driven CD4 and CD8 activation, while up-regulating the activation of the Th2 CD4 T-cell subset. Thus, the anti-inflammatory effects of n-3 fatty acids may result in both the direct suppression of IL-2-induced Th1 cell activation and the indirect suppression of Th1 cells by the enhanced cross-regulatory function of Th2 cells.

Fig. 1

Effect of dietary fatty acids on CD4 T-cell proliferation. Mice were fed diets enriched in SAF (black bars), EPA/DHA (grey bars) or FO (white bars) for 14 days. Splenic CD4 T lymphocytes were isolated and stimulated with various agonists as described in Materials and methods. Values (n = 5) represent the mean ± s.e.m. of net thymidine uptake (DPM). Different letters denote significant differences within each agonist group (P < 0·05). Abs, αCD3/αCD28 antibodies; 3/P, αCD3/PMA; P/I, PMA/Ionomycin.

Fig. 2

Effect of dietary fatty acids on CD8 T-cell proliferation. Mice were fed diets enriched in SAF (black bars), EPA/DHA (grey bars) or FO (white bars) for 14 days. Splenic CD8 T cells were isolated and stimulated with various agonists as described in Materials and methods. Values (SAF, n = 5; EPA/DHA, n = 5; FO, n = 6) represent the mean ± s.e.m. of net thymidine uptake (DPM). Different letters denote significant differences within each agonist group (P < 0·05). Abs, αCD3/αCD28 antibodies; 3/P, αCD3/PMA; P/I, PMA/Ionomycin.

Fig. 3

Effect of dietary fatty acids on IL-2 (a) and IL-4 (b) production by purified murine splenic CD4 T cells. Mice were fed diets enriched in SAF (black bars), EPA/DHA (grey bars) or FO (white bars) for 14 days, and purified splenic CD4 T cells were cultured with various agonists for 48 h. IL-2 and IL-4 in culture supernatant fluids were quantified by ELISA as described in Materials and methods. Values (n = 4) represent the mean ± s.e.m. in pg/200 000 cells. Different letters denote significant differences within each agonist group (P < 0·05). R, RPMI; Abs, αCD3/αCD28; 3/P, αCD3/PMA; P/I, PMA/Ionomycin.

Fig. 4

Effect of dietary fatty acids on IL-2 (a) and IL-4 (b) production by purified murine splenic CD8 T cells. Mice were fed diets enriched in SAF (black bars), EPA/DHA (grey bars) or FO (white bars) for 14 days, and purified splenic CD8 T cells were cultured with various agonists for 48 h. IL-2 and IL-4 in the culture supernatant fluids were quantified by ELISA as described in Materials and methods. Values (n = 5 or 6) represent the mean ± s.e.m. in pg/200 000 cells. Different letters denote significant differences within each agonist group (P < 0·05). R, RPMI; Abs, αCD3/αCD28; 3/P, αCD3/PMA; P/I, PMA/Ionomycin.