n-3 PUFA improves fatty acid composition, prevents palmitate-induced apoptosis, and differentially modifies B cell cytokine secretion in vitro and ex vivo - PubMed (original) (raw)

n-3 PUFA improves fatty acid composition, prevents palmitate-induced apoptosis, and differentially modifies B cell cytokine secretion in vitro and ex vivo

Benjamin Drew Rockett et al. J Lipid Res. 2010 Jun.

Abstract

n-3 polyunsaturated fatty acids (PUFAs) modify T-cell activation, in part by remodeling lipid composition; however, the relationship between n-3 PUFA and B-cell activation is unknown. Here we tested this relationship in vitro and ex vivo by measuring upregulation of B-cell surface molecules, the percentage of cells activated, and cytokine secreted in response to lipopolysaccharide (LPS) activation. In vitro, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) improved the membrane n-6/n-3 PUFA ratio, and DHA lowered interleukin (IL)-6 secretion; overall, n-3 PUFAs did not suppress B-cell activation compared with BSA, oleate, or elaidate treatment. Palmitate treatment suppressed the percentage of B cells activated through lipoapoptosis, which was differentially prevented by cosupplementing cells with MUFAs and PUFAs. Ex vivo, we tested the hypothesis with mice fed a control or high-fat saturated, hydrogenated, MUFA or n-3 PUFA diets. n-3 PUFAs had no effect on the percentage of B cells activated. Unexpectedly, the n-3 PUFA diet increased B-cell CD69 surface expression, IL-6 and IFNgamma secretion, and it significantly increased body weight gain. Overall, we propose that changes in lipid composition with n-3 PUFA and suppression of lymphocyte activation is not universal. The study highlights that high-fat n-3 PUFA diets can promote pro-inflammatory responses, at least from one cell type.

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Figures

Fig. 1.

Fig. 1.

Differential effects of fatty acids on the percentage of B cells activated, upregulation of surface molecules, and cytokine secretion. (A) Sample flow cytometry histograms to show staining of the B-cell activation marker CD69. (B) Percentage of B220+ B cells activated with LPS at 48 h after treatment with BSA or 25–50 µM PA, OA, ELA, EPA, and DHA. (C) Surface expression of B cell activation markers CD69, MHC class II, and CD80 after 48 h of treatment with 50 µM fatty acids. (D) Cytokine secretion from B220+ B cells treated with BSA or 50 µM PA, OA, ELA, EPA, and DHA treated for 48 h. Values are mean ± SEM, n = 3–10. Asterisks indicate difference from BSA, *P < 0.05, **P <0.01, ***P < 0.001. DHA, docosahexaenoic acid; ELA, elaidic acid; EPA, eicosapentaenoic acid; LPS, lipopolysaccharide; OA, oleic acid; PA, palmitic acid.

Fig. 2.

Fig. 2.

PA treatment promotes B cell lipoapoptosis. (A) Sample dot plots of Annexin V/Sytox Blue staining of B220+ B cells treated with the BSA control, PA, or EPA. (B) Percentage of cells that are Annexin V+ at 48 h of treatment with the BSA control or 50 µM PA, OA, ELA, EPA, or DHA. Values are mean ± SEM, n = 4. Asterisk indicates difference from BSA, *P < 0.05. DHA, docosahexaenoic acid; ELA, elaidic acid; EPA, eicosapentaenoic acid; OA, oleic acid; PA, palmitic acid.

Fig. 3.

Fig. 3.

Cosupplementing PA-treated cells with n-3 PUFAs prevents the effects of PA on B-cell phenotype and lipoapoptosis by increasing cell size. (A) Percentage of B cells activated and (B) percentage of apoptotic cells at 48 h of treatment with 50 µM fatty acids. (C) IL-6 secretion from B220+ B cells treated with 50 µM PA, and 50 µM PA + 50 µM OA, ELA, EPA, or DHA for 48 h. (D) MFI values for Nile Red staining at 48 h of treatment with 50 µM fatty acid. (E) Median forward scatter values for cells treated with 50 µM fatty acids at 48 h. Values are mean ± SEM, n = 3–5. Asterisks indicate difference from PA (A–C) or BSA (D–E), *P < 0.05, **P < 0.01, ***P < 0.001. DHA, docosahexaenoic acid; ELA, elaidic acid; EPA, eicosapentaenoic acid; MFI, median fluorescence intensity; OA, oleic acid; PA, palmitic acid.

Fig. 4.

Fig. 4.

Changes in fatty acid composition as a function of time for splenocytes isolated from mice fed different diets. Levels of (A) 18:1t (B) 18:3 n-3 (C) 20:4 n-6 (D) 20:5 n-3 (E) 22:5 n-3 (F) 22:6 n-3 and (G) n-6/n-3 PUFA ratio as a function of time after feeding mice ND or high-fat diets enriched in SFA, HFA, MUFA, and n-3 PUFA. (A–F) values (mean ± SEM) are percentage of the total fatty acids, n = 3 for 1 and 17 days, and n = 4 for 98 days. Asterisks indicate difference from ND, *P < 0.05, **P < 0.01, ***P < 0.001. Values on the Y axis vary from plot to plot, and for some values, the error bars are smaller than the data points. For simplicity, only fatty acids that showed differences relative to the control are presented. HFA, hydrogenated fatty acid; MUFA, monounsaturated fatty acid ND, normal diet; PUFA; polyunsaturated fatty acid SFA, saturated fatty acid.

Fig. 5.

Fig. 5.

High-fat n-3 PUFA diet does not modify the percentage of activated B cells but increases CD69 surface expression and secretion of IL-6 and IFNγ. (A) Percentage of B cells activated after 24 h of activation and (B) MFI of CD69, MHC class II, and CD80 of B cells isolated from mice fed ND, SFA, HFA, MUFA, and n-3 PUFA diets. (C) Cytokine secretion from B220+ B cells isolated from diet modified mice. Data are n = 3 (A) and n = 4 (B–C) per diet. Asterisks indicate different from ND, *P < 0.05, **P < 0.01. HFA, hydrogenated fatty acid; MUFA; monunsaturated fatty acid ND, normal diet; PUFA; polyunsaturated fatty acid SFA, saturated fatty acid.

Fig. 6.

Fig. 6.

High-fat n-3 PUFA diet increases body weight gain and induces splenomegaly. (A) Body weight as a function of time for mice fed ND or high-fat diets enriched in SFA, HFA, MUFA, and n-3 PUFA. Average daily (B) food and (C) kcal intake of mice fed the different diets as a function of time. The kcal values are normalized to the body weight. (D) Spleen weight of mice fed the different diets after 98 days of feeding. Values are mean ± SEM, n = 6 per diet. Asterisks indicate difference from ND, *P < 0.05, **P < 0.01, ***P < 0.001. HFA, hydrogenated fatty acid; MUFA; monunsaturated fatty acid ND, normal diet; PUFA; polyunsaturated fatty acid SFA, saturated fatty acid.

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