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The Polyunsaturated Fatty Acids Arachidonic Acid and Docosahexaenoic Acid Induce Mouse Dendritic Cells Maturation but Reduce T-Cell Responses In Vitro.

Carlsson JA, Wold AE, Sandberg AS, Östman SM - PLoS ONE (2015)

Bottom Line: Here, we measured the effects of omega-3 (n-3), n-6 and n-9 fatty acids on the interaction between dendritic cells (DCs) and naïve T cells.Fatty acids were taken up by the DCs, as shown by gas chromatography analysis.However, fewer T cells co-cultured with these DCs proliferated (CellTrace Violet low) and expressed CD69 or CD25, while more were necrotic (7AAD+).

View Article: PubMed Central - PubMed

Affiliation: Department of Infectious Diseases, Institute of Biomedicine, the Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden.

ABSTRACT
Long-chain polyunsaturated fatty acids (PUFAs) might regulate T-cell activation and lineage commitment. Here, we measured the effects of omega-3 (n-3), n-6 and n-9 fatty acids on the interaction between dendritic cells (DCs) and naïve T cells. Spleen DCs from BALB/c mice were cultured in vitro with ovalbumin (OVA) with 50 μM fatty acids; α-linolenic acid, arachidonic acid (AA), eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), linoleic acid or oleic acid and thereafter OVA-specific DO11.10 T cells were added to the cultures. Fatty acids were taken up by the DCs, as shown by gas chromatography analysis. After culture with arachidonic acid or DHA CD11c+ CD11b+ and CD11c+ CD11bneg DCs expressed more CD40, CD80, CD83, CD86 and PDL-1, while IAd remained unchanged. However, fewer T cells co-cultured with these DCs proliferated (CellTrace Violet low) and expressed CD69 or CD25, while more were necrotic (7AAD+). We noted an increased proportion of T cells with a regulatory T cell (Treg) phenotype, i.e., when gating on CD4+ FoxP3+ CTLA-4+, CD4+ FoxP3+ Helios+ or CD4+ FoxP3+ PD-1+, in co-cultures with arachidonic acid- or DHA-primed DCs relative to control cultures. The proportion of putative Tregs was inversely correlated to T-cell proliferation, indicating a suppressive function of these cells. With arachidonic acid DCs produced higher levels of prostaglandin E2 while T cells produced lower amounts of IL-10 and IFNγ. In conclusion arachidonic acid and DHA induced up-regulation of activation markers on DCs. However arachidonic acid- and DHA-primed DCs reduced T-cell proliferation and increased the proportion of T cells expressing FoxP3, indicating that these fatty acids can promote induction of regulatory T cells.

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Viability of T cells after co-culture.T cells were analyzed with 7AAD and Annexin V after 6 days of co-culture with dendritic cells (DCs) previously supplemented with fatty acids (50 μM); α-linolenic acid (ALA), arachidonic acid (AA), eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), linoleic acid (LA), oleic acid (OA) or ethanol only (Ctrl). (A) Stepwise gating procedure to eliminate debris. (B) Representative dot plots showing gating of live cells (Annexin V- 7AAD-), apoptotic cells (Annexin V+ 7AAD-) and necrotic and late apoptotic cells (Annexin V+ 7AAD+). (C) Proportion of live, (D) necrotic and late apoptotic and (E) apoptotic CD4+ T cells. Each dot represents one individual. Black dots denote samples supplemented with fatty acid while white dots with black borders denote control (ethanol only). Horizontal solid black lines show median value. The median from the control group has been extended with a dotted line for easy comparison to the other groups. Statistical mean difference was compared to the control group. Data are representative of two independent experiments. p-values: ** <0.01, *** <0.001, **** <0.0001.
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pone.0143741.g007: Viability of T cells after co-culture.T cells were analyzed with 7AAD and Annexin V after 6 days of co-culture with dendritic cells (DCs) previously supplemented with fatty acids (50 μM); α-linolenic acid (ALA), arachidonic acid (AA), eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), linoleic acid (LA), oleic acid (OA) or ethanol only (Ctrl). (A) Stepwise gating procedure to eliminate debris. (B) Representative dot plots showing gating of live cells (Annexin V- 7AAD-), apoptotic cells (Annexin V+ 7AAD-) and necrotic and late apoptotic cells (Annexin V+ 7AAD+). (C) Proportion of live, (D) necrotic and late apoptotic and (E) apoptotic CD4+ T cells. Each dot represents one individual. Black dots denote samples supplemented with fatty acid while white dots with black borders denote control (ethanol only). Horizontal solid black lines show median value. The median from the control group has been extended with a dotted line for easy comparison to the other groups. Statistical mean difference was compared to the control group. Data are representative of two independent experiments. p-values: ** <0.01, *** <0.001, **** <0.0001.

Mentions: We determined the levels of apoptosis and necrosis in the T cells by staining with Annexin V and 7AAD. The gating strategy is shown in Fig 7A and 7B. Briefly, to identify debris, live cells (Annexin V- 7AAD-) were gated and on those cells we put a non-debris gate from which CD4+ T cells were gated for Annexin V and 7AAD staining. There was a non-significantly lower proportion of live T cells in the DC: T cell co-cultures with arachidonic acid-primed DCs compared to control cultures (70% vs 80%, Fig 7C) and a corresponding increase in late stage apoptotic/necrotic cells (Annexin V+ 7AAD+, ≈ 20% compared to 5%, Fig 7D), but no differences in early stage apoptotic T cells (Annexin V+ 7AAD-, Fig 7E).


The Polyunsaturated Fatty Acids Arachidonic Acid and Docosahexaenoic Acid Induce Mouse Dendritic Cells Maturation but Reduce T-Cell Responses In Vitro.

Carlsson JA, Wold AE, Sandberg AS, Östman SM - PLoS ONE (2015)

Viability of T cells after co-culture.T cells were analyzed with 7AAD and Annexin V after 6 days of co-culture with dendritic cells (DCs) previously supplemented with fatty acids (50 μM); α-linolenic acid (ALA), arachidonic acid (AA), eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), linoleic acid (LA), oleic acid (OA) or ethanol only (Ctrl). (A) Stepwise gating procedure to eliminate debris. (B) Representative dot plots showing gating of live cells (Annexin V- 7AAD-), apoptotic cells (Annexin V+ 7AAD-) and necrotic and late apoptotic cells (Annexin V+ 7AAD+). (C) Proportion of live, (D) necrotic and late apoptotic and (E) apoptotic CD4+ T cells. Each dot represents one individual. Black dots denote samples supplemented with fatty acid while white dots with black borders denote control (ethanol only). Horizontal solid black lines show median value. The median from the control group has been extended with a dotted line for easy comparison to the other groups. Statistical mean difference was compared to the control group. Data are representative of two independent experiments. p-values: ** <0.01, *** <0.001, **** <0.0001.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4664484&req=5

pone.0143741.g007: Viability of T cells after co-culture.T cells were analyzed with 7AAD and Annexin V after 6 days of co-culture with dendritic cells (DCs) previously supplemented with fatty acids (50 μM); α-linolenic acid (ALA), arachidonic acid (AA), eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), linoleic acid (LA), oleic acid (OA) or ethanol only (Ctrl). (A) Stepwise gating procedure to eliminate debris. (B) Representative dot plots showing gating of live cells (Annexin V- 7AAD-), apoptotic cells (Annexin V+ 7AAD-) and necrotic and late apoptotic cells (Annexin V+ 7AAD+). (C) Proportion of live, (D) necrotic and late apoptotic and (E) apoptotic CD4+ T cells. Each dot represents one individual. Black dots denote samples supplemented with fatty acid while white dots with black borders denote control (ethanol only). Horizontal solid black lines show median value. The median from the control group has been extended with a dotted line for easy comparison to the other groups. Statistical mean difference was compared to the control group. Data are representative of two independent experiments. p-values: ** <0.01, *** <0.001, **** <0.0001.
Mentions: We determined the levels of apoptosis and necrosis in the T cells by staining with Annexin V and 7AAD. The gating strategy is shown in Fig 7A and 7B. Briefly, to identify debris, live cells (Annexin V- 7AAD-) were gated and on those cells we put a non-debris gate from which CD4+ T cells were gated for Annexin V and 7AAD staining. There was a non-significantly lower proportion of live T cells in the DC: T cell co-cultures with arachidonic acid-primed DCs compared to control cultures (70% vs 80%, Fig 7C) and a corresponding increase in late stage apoptotic/necrotic cells (Annexin V+ 7AAD+, ≈ 20% compared to 5%, Fig 7D), but no differences in early stage apoptotic T cells (Annexin V+ 7AAD-, Fig 7E).

Bottom Line: Here, we measured the effects of omega-3 (n-3), n-6 and n-9 fatty acids on the interaction between dendritic cells (DCs) and naïve T cells.Fatty acids were taken up by the DCs, as shown by gas chromatography analysis.However, fewer T cells co-cultured with these DCs proliferated (CellTrace Violet low) and expressed CD69 or CD25, while more were necrotic (7AAD+).

View Article: PubMed Central - PubMed

Affiliation: Department of Infectious Diseases, Institute of Biomedicine, the Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden.

ABSTRACT
Long-chain polyunsaturated fatty acids (PUFAs) might regulate T-cell activation and lineage commitment. Here, we measured the effects of omega-3 (n-3), n-6 and n-9 fatty acids on the interaction between dendritic cells (DCs) and naïve T cells. Spleen DCs from BALB/c mice were cultured in vitro with ovalbumin (OVA) with 50 μM fatty acids; α-linolenic acid, arachidonic acid (AA), eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), linoleic acid or oleic acid and thereafter OVA-specific DO11.10 T cells were added to the cultures. Fatty acids were taken up by the DCs, as shown by gas chromatography analysis. After culture with arachidonic acid or DHA CD11c+ CD11b+ and CD11c+ CD11bneg DCs expressed more CD40, CD80, CD83, CD86 and PDL-1, while IAd remained unchanged. However, fewer T cells co-cultured with these DCs proliferated (CellTrace Violet low) and expressed CD69 or CD25, while more were necrotic (7AAD+). We noted an increased proportion of T cells with a regulatory T cell (Treg) phenotype, i.e., when gating on CD4+ FoxP3+ CTLA-4+, CD4+ FoxP3+ Helios+ or CD4+ FoxP3+ PD-1+, in co-cultures with arachidonic acid- or DHA-primed DCs relative to control cultures. The proportion of putative Tregs was inversely correlated to T-cell proliferation, indicating a suppressive function of these cells. With arachidonic acid DCs produced higher levels of prostaglandin E2 while T cells produced lower amounts of IL-10 and IFNγ. In conclusion arachidonic acid and DHA induced up-regulation of activation markers on DCs. However arachidonic acid- and DHA-primed DCs reduced T-cell proliferation and increased the proportion of T cells expressing FoxP3, indicating that these fatty acids can promote induction of regulatory T cells.

Show MeSH
Related in: MedlinePlus