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An A2B Adenosine Receptor Agonist Promotes Th17 Autoimmune Responses in Experimental Autoimmune Uveitis (EAU) via Dendritic Cell Activation.

Chen M, Liang D, Zuo A, Shao H, Kaplan HJ, Sun D - PLoS ONE (2015)

Bottom Line: We have recently reported that, although adenosine receptor (AR) agonists have a suppressive effect on Th1 autoreactive T cells, their effect on Th17 autoreactive T cells and γδ T cells is stimulatory and this effect is mainly mediated via A2A adenosine receptors (A2ARs).In the present study, we showed that exposure of DCs to A2BR agonist facilitated γδ T cell activation, leading to augmented Th17 responses and progressive EAU development.Our results further support our previous finding that AR agonists have distinct effects on Th1 and Th17 autoimmune responses.

View Article: PubMed Central - PubMed

Affiliation: Doheny Eye Institute and Department of Ophthalmology, University of California, Los Angeles, CA90033, United States of America.

ABSTRACT
We have recently reported that, although adenosine receptor (AR) agonists have a suppressive effect on Th1 autoreactive T cells, their effect on Th17 autoreactive T cells and γδ T cells is stimulatory and this effect is mainly mediated via A2A adenosine receptors (A2ARs). In this study, we further demonstrate that treatment of C57BL/6 (B6) mice with a selective A2B adenosine receptor (A2BR) agonist greatly enhanced the development of experimental autoimmune uveitis (EAU), whereas treatment with an A2BR antagonist significantly ameliorated severity of EAU. The A2BR agonist-treated mice showed augmented Th17, but not Th1, responses. Mechanistic studies showed that the A2BR agonist-induced enhancement of the Th17 response was significantly lower when TCR-δ-/- mice received the same treatment and that transfer of γδ T cells into TCR-δ-/- mice partially restored this effect. We also showed that dendritic cells (DCs) from A2BR agonist-treated mice showed a significantly increased ability to activate γδ T cells and Th17 autoreactive T cells. Thus, our previous studies have shown that, in EAU, activated γδ T cells possess greatly increased ability to enhance Th17 autoimmune responses. In the present study, we showed that exposure of DCs to A2BR agonist facilitated γδ T cell activation, leading to augmented Th17 responses and progressive EAU development. Our results further support our previous finding that AR agonists have distinct effects on Th1 and Th17 autoimmune responses.

No MeSH data available.


Related in: MedlinePlus

γδ T cells in A2BR agonist-treated mice were more activated in vivo and possesed increased Th17-promoting activity.A) Proportional numbers of γδ T cells and their activation status were compared among γδ T cells in naive, immunized, and BAY60-6538-treated, immunized mice. Splenic T cells were isolated from pooled mice, stained with anti-αβ, anti-γδ or CD44 antibodies, and followed FACS analysis. B) Cytokine production of freshly isolated γδ T cells was examined after cultured for 48 h in vitro, in the absence of additional stimulation. C) In vivo reconstitution study showing that γδ T cells isolated from BAY60-6538-treated mice have increased Th17-promoting activity when injected to TCR-δ-/- mice. Groups of TCR-δ-/- mice (n = 6) immunized with the uveitogenic peptide (IRBP1-20/CFA) were injected i.p. at the day of immunization with 105 γδ T cells isolated from BAY60-6538-treated or untreated mice, before they were immunized with IRNP/CFA. 13 days post-immunization, splenic T cells were enriched and stimulated with immunizing peptide under Th1- and Th17-polarized conditions, and the activated T cells were stained for intracellular expression of IL-17 (top row) or IFN-g (bottom row). Results of one single experiment are shown, which were repeated for at least 3 times. D) 48-hour culture supernatants of the cells cultured in (C) were assessed for IL-17 and IFN-γ by ELISA. Data are from one single experiment, which are representative of three independent experiments **p<0.05.
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pone.0132348.g004: γδ T cells in A2BR agonist-treated mice were more activated in vivo and possesed increased Th17-promoting activity.A) Proportional numbers of γδ T cells and their activation status were compared among γδ T cells in naive, immunized, and BAY60-6538-treated, immunized mice. Splenic T cells were isolated from pooled mice, stained with anti-αβ, anti-γδ or CD44 antibodies, and followed FACS analysis. B) Cytokine production of freshly isolated γδ T cells was examined after cultured for 48 h in vitro, in the absence of additional stimulation. C) In vivo reconstitution study showing that γδ T cells isolated from BAY60-6538-treated mice have increased Th17-promoting activity when injected to TCR-δ-/- mice. Groups of TCR-δ-/- mice (n = 6) immunized with the uveitogenic peptide (IRBP1-20/CFA) were injected i.p. at the day of immunization with 105 γδ T cells isolated from BAY60-6538-treated or untreated mice, before they were immunized with IRNP/CFA. 13 days post-immunization, splenic T cells were enriched and stimulated with immunizing peptide under Th1- and Th17-polarized conditions, and the activated T cells were stained for intracellular expression of IL-17 (top row) or IFN-g (bottom row). Results of one single experiment are shown, which were repeated for at least 3 times. D) 48-hour culture supernatants of the cells cultured in (C) were assessed for IL-17 and IFN-γ by ELISA. Data are from one single experiment, which are representative of three independent experiments **p<0.05.

Mentions: We have previously shown that an enhanced Th17 response is closely associated with increased γδ T cell activation [10,11,13]. To determine whether γδ T cells in treated mice were activated and functionally more active, we compared the percentage of γδ T cells in total T cells from naïve B6 mice and immunized B6 mice with or without BAY 60–6538 treatment and examined them for expression of the T cell activation marker CD44, IL-17 production, and their ability to promote a Th17 response. The results showed that the percentage of γδ T cells increased significantly in immunized mice compared to naïve mice and increased further with BAY 60–6538 treatment (Fig 4A, upper panel). A significantly greater percentage (55%) of γδ T cells isolated from BAY 60-6538-treated mice expressed CD44, a cell surface marker of activated T cells, as compared to vehicle-treated mice (31%) (Fig 4A, lower panel). Moreover, freshly isolated γδ T cells from BAY 60-6538-treated mice secreted significantly higher amounts of IL-17 in the absence of in vitro stimulation, as compared to γδ T cells isolated from vehicle-treated immunized mice (Fig 4B). Based on our previous findings that T cells from immunized TCR-δ-/- mice consistently generate low, but significant, numbers of IL-17+ T cells and that addition of γδ T cells from immunized B6 mice to pure responder T cells from immunized TCR-δ-/- mice (a minimum of 2% of the total T cells) during an in vitro response greatly increases the activation of IL-17+ uveitogenic T cells [10,11,13], we examined whether γδ T cells isolated from BAY 60-6538-treated immunized B6 mice showed an increased Th17-promoting effect and found that γδ T cells from BAY 60-6538-treated mice showed a greater ability to increase IL-17 responses, as assessed by either the number of IL-17+ T cells among in vitro activated T cells (Fig 4C) or the amount of IL-17 in activated T cell culture supernatants (Fig 4D).


An A2B Adenosine Receptor Agonist Promotes Th17 Autoimmune Responses in Experimental Autoimmune Uveitis (EAU) via Dendritic Cell Activation.

Chen M, Liang D, Zuo A, Shao H, Kaplan HJ, Sun D - PLoS ONE (2015)

γδ T cells in A2BR agonist-treated mice were more activated in vivo and possesed increased Th17-promoting activity.A) Proportional numbers of γδ T cells and their activation status were compared among γδ T cells in naive, immunized, and BAY60-6538-treated, immunized mice. Splenic T cells were isolated from pooled mice, stained with anti-αβ, anti-γδ or CD44 antibodies, and followed FACS analysis. B) Cytokine production of freshly isolated γδ T cells was examined after cultured for 48 h in vitro, in the absence of additional stimulation. C) In vivo reconstitution study showing that γδ T cells isolated from BAY60-6538-treated mice have increased Th17-promoting activity when injected to TCR-δ-/- mice. Groups of TCR-δ-/- mice (n = 6) immunized with the uveitogenic peptide (IRBP1-20/CFA) were injected i.p. at the day of immunization with 105 γδ T cells isolated from BAY60-6538-treated or untreated mice, before they were immunized with IRNP/CFA. 13 days post-immunization, splenic T cells were enriched and stimulated with immunizing peptide under Th1- and Th17-polarized conditions, and the activated T cells were stained for intracellular expression of IL-17 (top row) or IFN-g (bottom row). Results of one single experiment are shown, which were repeated for at least 3 times. D) 48-hour culture supernatants of the cells cultured in (C) were assessed for IL-17 and IFN-γ by ELISA. Data are from one single experiment, which are representative of three independent experiments **p<0.05.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4492970&req=5

pone.0132348.g004: γδ T cells in A2BR agonist-treated mice were more activated in vivo and possesed increased Th17-promoting activity.A) Proportional numbers of γδ T cells and their activation status were compared among γδ T cells in naive, immunized, and BAY60-6538-treated, immunized mice. Splenic T cells were isolated from pooled mice, stained with anti-αβ, anti-γδ or CD44 antibodies, and followed FACS analysis. B) Cytokine production of freshly isolated γδ T cells was examined after cultured for 48 h in vitro, in the absence of additional stimulation. C) In vivo reconstitution study showing that γδ T cells isolated from BAY60-6538-treated mice have increased Th17-promoting activity when injected to TCR-δ-/- mice. Groups of TCR-δ-/- mice (n = 6) immunized with the uveitogenic peptide (IRBP1-20/CFA) were injected i.p. at the day of immunization with 105 γδ T cells isolated from BAY60-6538-treated or untreated mice, before they were immunized with IRNP/CFA. 13 days post-immunization, splenic T cells were enriched and stimulated with immunizing peptide under Th1- and Th17-polarized conditions, and the activated T cells were stained for intracellular expression of IL-17 (top row) or IFN-g (bottom row). Results of one single experiment are shown, which were repeated for at least 3 times. D) 48-hour culture supernatants of the cells cultured in (C) were assessed for IL-17 and IFN-γ by ELISA. Data are from one single experiment, which are representative of three independent experiments **p<0.05.
Mentions: We have previously shown that an enhanced Th17 response is closely associated with increased γδ T cell activation [10,11,13]. To determine whether γδ T cells in treated mice were activated and functionally more active, we compared the percentage of γδ T cells in total T cells from naïve B6 mice and immunized B6 mice with or without BAY 60–6538 treatment and examined them for expression of the T cell activation marker CD44, IL-17 production, and their ability to promote a Th17 response. The results showed that the percentage of γδ T cells increased significantly in immunized mice compared to naïve mice and increased further with BAY 60–6538 treatment (Fig 4A, upper panel). A significantly greater percentage (55%) of γδ T cells isolated from BAY 60-6538-treated mice expressed CD44, a cell surface marker of activated T cells, as compared to vehicle-treated mice (31%) (Fig 4A, lower panel). Moreover, freshly isolated γδ T cells from BAY 60-6538-treated mice secreted significantly higher amounts of IL-17 in the absence of in vitro stimulation, as compared to γδ T cells isolated from vehicle-treated immunized mice (Fig 4B). Based on our previous findings that T cells from immunized TCR-δ-/- mice consistently generate low, but significant, numbers of IL-17+ T cells and that addition of γδ T cells from immunized B6 mice to pure responder T cells from immunized TCR-δ-/- mice (a minimum of 2% of the total T cells) during an in vitro response greatly increases the activation of IL-17+ uveitogenic T cells [10,11,13], we examined whether γδ T cells isolated from BAY 60-6538-treated immunized B6 mice showed an increased Th17-promoting effect and found that γδ T cells from BAY 60-6538-treated mice showed a greater ability to increase IL-17 responses, as assessed by either the number of IL-17+ T cells among in vitro activated T cells (Fig 4C) or the amount of IL-17 in activated T cell culture supernatants (Fig 4D).

Bottom Line: We have recently reported that, although adenosine receptor (AR) agonists have a suppressive effect on Th1 autoreactive T cells, their effect on Th17 autoreactive T cells and γδ T cells is stimulatory and this effect is mainly mediated via A2A adenosine receptors (A2ARs).In the present study, we showed that exposure of DCs to A2BR agonist facilitated γδ T cell activation, leading to augmented Th17 responses and progressive EAU development.Our results further support our previous finding that AR agonists have distinct effects on Th1 and Th17 autoimmune responses.

View Article: PubMed Central - PubMed

Affiliation: Doheny Eye Institute and Department of Ophthalmology, University of California, Los Angeles, CA90033, United States of America.

ABSTRACT
We have recently reported that, although adenosine receptor (AR) agonists have a suppressive effect on Th1 autoreactive T cells, their effect on Th17 autoreactive T cells and γδ T cells is stimulatory and this effect is mainly mediated via A2A adenosine receptors (A2ARs). In this study, we further demonstrate that treatment of C57BL/6 (B6) mice with a selective A2B adenosine receptor (A2BR) agonist greatly enhanced the development of experimental autoimmune uveitis (EAU), whereas treatment with an A2BR antagonist significantly ameliorated severity of EAU. The A2BR agonist-treated mice showed augmented Th17, but not Th1, responses. Mechanistic studies showed that the A2BR agonist-induced enhancement of the Th17 response was significantly lower when TCR-δ-/- mice received the same treatment and that transfer of γδ T cells into TCR-δ-/- mice partially restored this effect. We also showed that dendritic cells (DCs) from A2BR agonist-treated mice showed a significantly increased ability to activate γδ T cells and Th17 autoreactive T cells. Thus, our previous studies have shown that, in EAU, activated γδ T cells possess greatly increased ability to enhance Th17 autoimmune responses. In the present study, we showed that exposure of DCs to A2BR agonist facilitated γδ T cell activation, leading to augmented Th17 responses and progressive EAU development. Our results further support our previous finding that AR agonists have distinct effects on Th1 and Th17 autoimmune responses.

No MeSH data available.


Related in: MedlinePlus