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Infectious tolerance: human CD25(+) regulatory T cells convey suppressor activity to conventional CD4(+) T helper cells.

Jonuleit H, Schmitt E, Kakirman H, Stassen M, Knop J, Enk AH - J. Exp. Med. (2002)

Bottom Line: Coactivation of CD25(+) Treg cells with Treg cell-depleted CD4(+) T cells results in anergized CD4(+) T cells that in turn inhibit the activation of conventional, freshly isolated CD4(+) T helper (Th) cells.The induction of suppressive properties in conventional CD4(+) Th cells represents a mechanism underlying the phenomenon of infectious tolerance.This explains previously published conflicting data on the role of TGF-beta in CD25(+) Treg cell-induced immunosuppression.

View Article: PubMed Central - PubMed

Affiliation: Department of Dermatology, University of Mainz, 55101 Mainz, Germany. jonuleit@hautklinik.klinik.uni-mainz.de

ABSTRACT
Regulatory CD4(+)CD25(+) T cells (Treg) are mandatory for maintaining immunologic self-tolerance. We demonstrate that the cell-cell contact-mediated suppression of conventional CD4(+) T cells by human CD25(+) Treg cells is fixation resistant, independent from membrane-bound TGF-beta but requires activation and protein synthesis of CD25(+) Treg cells. Coactivation of CD25(+) Treg cells with Treg cell-depleted CD4(+) T cells results in anergized CD4(+) T cells that in turn inhibit the activation of conventional, freshly isolated CD4(+) T helper (Th) cells. This infectious suppressive activity, transferred from CD25(+) Treg cells via cell contact, is cell contact-independent and partially mediated by soluble transforming growth factor (TGF)-beta. The induction of suppressive properties in conventional CD4(+) Th cells represents a mechanism underlying the phenomenon of infectious tolerance. This explains previously published conflicting data on the role of TGF-beta in CD25(+) Treg cell-induced immunosuppression.

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The suppressive activity of activated CD25+ Treg cells is fixation-resistant but the activation is sensitive to treatment with monensin or cycloheximide. (A) Freshly isolated CD4+ T cells (105 cells per well) and CD25+ Treg cells (105 cells per well) or a combination of both (1:1) were activated with anti-CD3 (1 μg/ml) and anti-CD28 mAb (2 μg/ml). In addition, CD4+ T cells were coactivated with CD25+ Treg cells or conventional CD4+ T cells that were immediately fixed after isolation (1% paraformaldehyde, 10 min) or were fixed after preactivation with 0.5 μg anti-CD3 mAb plus 10 U/ml IL-2 for 20 h (black bars). (B) CD4+ T cells were coactivated either with preactivated and fixed CD25+ Treg cells or with CD25+ Treg cells treated with monensin (1 μg/ml) or cycloheximide (10 μg/ml) during preactivation before fixation. 3[H]Tdr was added after 3 d of culture for the final 16 h.
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fig2: The suppressive activity of activated CD25+ Treg cells is fixation-resistant but the activation is sensitive to treatment with monensin or cycloheximide. (A) Freshly isolated CD4+ T cells (105 cells per well) and CD25+ Treg cells (105 cells per well) or a combination of both (1:1) were activated with anti-CD3 (1 μg/ml) and anti-CD28 mAb (2 μg/ml). In addition, CD4+ T cells were coactivated with CD25+ Treg cells or conventional CD4+ T cells that were immediately fixed after isolation (1% paraformaldehyde, 10 min) or were fixed after preactivation with 0.5 μg anti-CD3 mAb plus 10 U/ml IL-2 for 20 h (black bars). (B) CD4+ T cells were coactivated either with preactivated and fixed CD25+ Treg cells or with CD25+ Treg cells treated with monensin (1 μg/ml) or cycloheximide (10 μg/ml) during preactivation before fixation. 3[H]Tdr was added after 3 d of culture for the final 16 h.

Mentions: To analyze the suppressive mechanism in more detail, we stimulated CD4+ T cells in the presence of fixed CD25+ Treg cells. In coculture experiments, freshly isolated human CD25+ Treg cells inhibited the proliferation and cytokine production of coactivated conventional CD4+ T cells in a dose-dependent manner. If CD25+ Treg cells were fixed directly after isolation, no suppressive activity could be detected (Fig. 2 A). However, if CD25+ Treg cells were preactivated overnight with anti-CD3 antibodies (0.5 μg/ml) and 10 U/ml IL-2 before fixation, they showed a comparable suppressive capacity for CD4+ T cells as unfixed CD25+ Treg cells. In contrast, Treg-depleted and activated conventional CD4+ T helper cells did not exert any suppressive activity, although such cells express comparable amounts of TGF-β (Fig. 1 A). These data suggest that the inhibitory function of human CD25+ Treg cells is activation dependent. Additional experiments revealed that the induction of suppressor activity requires protein synthesis as it can be inhibited by the presence of cycloheximide or monensin (Fig. 2 B). However, once activated, the suppressive activity of human CD25+ Treg cells is fixation-resistant (Fig. 2 A). These data also strongly corroborate our findings that the inhibitory function of human CD25+ Treg cells is independent of soluble mediators (6). Furthermore, the activation of suppressor function of human CD25+ Treg is also independent of costimulation, since the addition of mature DCs or soluble anti-CD28 antibodies during overnight preactivation with anti-CD3 antibodies did not alter/enhance the functional activities of human CD25+ Treg cells (unpublished data).


Infectious tolerance: human CD25(+) regulatory T cells convey suppressor activity to conventional CD4(+) T helper cells.

Jonuleit H, Schmitt E, Kakirman H, Stassen M, Knop J, Enk AH - J. Exp. Med. (2002)

The suppressive activity of activated CD25+ Treg cells is fixation-resistant but the activation is sensitive to treatment with monensin or cycloheximide. (A) Freshly isolated CD4+ T cells (105 cells per well) and CD25+ Treg cells (105 cells per well) or a combination of both (1:1) were activated with anti-CD3 (1 μg/ml) and anti-CD28 mAb (2 μg/ml). In addition, CD4+ T cells were coactivated with CD25+ Treg cells or conventional CD4+ T cells that were immediately fixed after isolation (1% paraformaldehyde, 10 min) or were fixed after preactivation with 0.5 μg anti-CD3 mAb plus 10 U/ml IL-2 for 20 h (black bars). (B) CD4+ T cells were coactivated either with preactivated and fixed CD25+ Treg cells or with CD25+ Treg cells treated with monensin (1 μg/ml) or cycloheximide (10 μg/ml) during preactivation before fixation. 3[H]Tdr was added after 3 d of culture for the final 16 h.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2193929&req=5

fig2: The suppressive activity of activated CD25+ Treg cells is fixation-resistant but the activation is sensitive to treatment with monensin or cycloheximide. (A) Freshly isolated CD4+ T cells (105 cells per well) and CD25+ Treg cells (105 cells per well) or a combination of both (1:1) were activated with anti-CD3 (1 μg/ml) and anti-CD28 mAb (2 μg/ml). In addition, CD4+ T cells were coactivated with CD25+ Treg cells or conventional CD4+ T cells that were immediately fixed after isolation (1% paraformaldehyde, 10 min) or were fixed after preactivation with 0.5 μg anti-CD3 mAb plus 10 U/ml IL-2 for 20 h (black bars). (B) CD4+ T cells were coactivated either with preactivated and fixed CD25+ Treg cells or with CD25+ Treg cells treated with monensin (1 μg/ml) or cycloheximide (10 μg/ml) during preactivation before fixation. 3[H]Tdr was added after 3 d of culture for the final 16 h.
Mentions: To analyze the suppressive mechanism in more detail, we stimulated CD4+ T cells in the presence of fixed CD25+ Treg cells. In coculture experiments, freshly isolated human CD25+ Treg cells inhibited the proliferation and cytokine production of coactivated conventional CD4+ T cells in a dose-dependent manner. If CD25+ Treg cells were fixed directly after isolation, no suppressive activity could be detected (Fig. 2 A). However, if CD25+ Treg cells were preactivated overnight with anti-CD3 antibodies (0.5 μg/ml) and 10 U/ml IL-2 before fixation, they showed a comparable suppressive capacity for CD4+ T cells as unfixed CD25+ Treg cells. In contrast, Treg-depleted and activated conventional CD4+ T helper cells did not exert any suppressive activity, although such cells express comparable amounts of TGF-β (Fig. 1 A). These data suggest that the inhibitory function of human CD25+ Treg cells is activation dependent. Additional experiments revealed that the induction of suppressor activity requires protein synthesis as it can be inhibited by the presence of cycloheximide or monensin (Fig. 2 B). However, once activated, the suppressive activity of human CD25+ Treg cells is fixation-resistant (Fig. 2 A). These data also strongly corroborate our findings that the inhibitory function of human CD25+ Treg cells is independent of soluble mediators (6). Furthermore, the activation of suppressor function of human CD25+ Treg is also independent of costimulation, since the addition of mature DCs or soluble anti-CD28 antibodies during overnight preactivation with anti-CD3 antibodies did not alter/enhance the functional activities of human CD25+ Treg cells (unpublished data).

Bottom Line: Coactivation of CD25(+) Treg cells with Treg cell-depleted CD4(+) T cells results in anergized CD4(+) T cells that in turn inhibit the activation of conventional, freshly isolated CD4(+) T helper (Th) cells.The induction of suppressive properties in conventional CD4(+) Th cells represents a mechanism underlying the phenomenon of infectious tolerance.This explains previously published conflicting data on the role of TGF-beta in CD25(+) Treg cell-induced immunosuppression.

View Article: PubMed Central - PubMed

Affiliation: Department of Dermatology, University of Mainz, 55101 Mainz, Germany. jonuleit@hautklinik.klinik.uni-mainz.de

ABSTRACT
Regulatory CD4(+)CD25(+) T cells (Treg) are mandatory for maintaining immunologic self-tolerance. We demonstrate that the cell-cell contact-mediated suppression of conventional CD4(+) T cells by human CD25(+) Treg cells is fixation resistant, independent from membrane-bound TGF-beta but requires activation and protein synthesis of CD25(+) Treg cells. Coactivation of CD25(+) Treg cells with Treg cell-depleted CD4(+) T cells results in anergized CD4(+) T cells that in turn inhibit the activation of conventional, freshly isolated CD4(+) T helper (Th) cells. This infectious suppressive activity, transferred from CD25(+) Treg cells via cell contact, is cell contact-independent and partially mediated by soluble transforming growth factor (TGF)-beta. The induction of suppressive properties in conventional CD4(+) Th cells represents a mechanism underlying the phenomenon of infectious tolerance. This explains previously published conflicting data on the role of TGF-beta in CD25(+) Treg cell-induced immunosuppression.

Show MeSH
Related in: MedlinePlus