Limits...
Persistence of peptide-induced CD4+ T cell anergy in vitro.

Ryan KR, Evavold BD - J. Exp. Med. (1998)

Bottom Line: The anergic state was characterized by blunted proliferation and interleukin (IL) 2 production upon restimulation with Hb(64-76), and was not the result of impaired TCR/CD3 downmodulation.Interestingly, persistence of the anergic phenotype was observed despite biweekly restimulation with antigen, APCs, and IL-2.Thus, T cell unresponsiveness induced by a peptide produced a stable, persistent anergic state in a Th0 clone that was not reversible by stimulation with IL-2 or -12.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Emory University, Atlanta, Georgia 30322, USA.

ABSTRACT
Clonal T cell unresponsiveness, or anergy, has been proposed as a mechanism of peripheral tolerance in vivo, and as a potential means of curbing unwanted T cell responses. In this study, anergy was induced in a T helper cell (Th) clone reactive to hemoglobin (Hb) peptide 64-76 by coculture of the T cells with live antigen-presenting cells (APCs) and 74L, a peptide analog of Hb(64-76) that contains a single amino acid substitution of leucine for glycine at position 74, or with a low concentration of the agonist ligand. The anergic state was characterized by blunted proliferation and interleukin (IL) 2 production upon restimulation with Hb(64-76), and was not the result of impaired TCR/CD3 downmodulation. The addition of exogenous IL-12 transiently restored proliferation of the anergic lines, but removal of IL-12 from culture returned the T cells to their nonproliferative state. Interestingly, persistence of the anergic phenotype was observed despite biweekly restimulation with antigen, APCs, and IL-2. Thus, T cell unresponsiveness induced by a peptide produced a stable, persistent anergic state in a Th0 clone that was not reversible by stimulation with IL-2 or -12.

Show MeSH
(a) The addition of IL-12 increases proliferation of clone I to  Hb(64–76) and 74L. Proliferation assay was performed as in Fig. 1, with or  without the addition of 10 ng/ml rIL-12. (b) Culture of the unresponsive  clones with IL-12 restores their proliferation. IL-12 was added to the culture  of the unresonsive clones during one of the biweekly restimulations. After 2  wk the IL-12 was washed away and proliferation of the clones was measured  as before. (c) The IL-12–treated clones return to their unresponsive state  upon removal of the IL-12 from culture. 2 wk after the treatment with IL-12, the IL-12 was washed away and the clones were passaged in culture without IL-12. After another 2 wk, proliferation was determined by DNA incorporation of [3H]thymidine.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2199180&req=5

Figure 6: (a) The addition of IL-12 increases proliferation of clone I to Hb(64–76) and 74L. Proliferation assay was performed as in Fig. 1, with or without the addition of 10 ng/ml rIL-12. (b) Culture of the unresponsive clones with IL-12 restores their proliferation. IL-12 was added to the culture of the unresonsive clones during one of the biweekly restimulations. After 2 wk the IL-12 was washed away and proliferation of the clones was measured as before. (c) The IL-12–treated clones return to their unresponsive state upon removal of the IL-12 from culture. 2 wk after the treatment with IL-12, the IL-12 was washed away and the clones were passaged in culture without IL-12. After another 2 wk, proliferation was determined by DNA incorporation of [3H]thymidine.

Mentions: The addition of exogenous costimulation has been used to overcome anergy in most of the model systems (9, 24, 25), but costimulation does not alter the anergic phenotype induced with partial agonist peptides (10, 17). Previous work with a Th2 clone demonstrated that a soluble factor (IL-1) could restore the proliferative response to a partial agonist peptide (3), and we have similarly observed that IL-12 boosts the proliferation of clone I to antigen (Fig. 6 A). In fact, a significant proliferative response was observed with both agonist peptide (0.1 μM) and partial agonist (10 μM 74L) by the addition of IL-12. To determine the effects of exogenous IL-12 on our anergic Th0 cells, 10 ng/ml recombinant IL-12 was added to the biweekly restimulations, and proliferation to Hb(64– 76) was assessed (Fig. 6 B). Inclusion of IL-12 in culture with the anergic T cells (but not in the proliferation assay) restored the proliferation of these lines to a similar level as clone I. However, the effect of IL-12 was transient, as the T cells retained their low proliferative response to Hb(64– 76) after subsequent passage (antigen, APCs, and IL-2) in the absence of exogenous IL-12 (Fig. 6 C). The increased proliferation resulting from IL-12 costimulation was not sufficient to reverse the anergic phenotype.


Persistence of peptide-induced CD4+ T cell anergy in vitro.

Ryan KR, Evavold BD - J. Exp. Med. (1998)

(a) The addition of IL-12 increases proliferation of clone I to  Hb(64–76) and 74L. Proliferation assay was performed as in Fig. 1, with or  without the addition of 10 ng/ml rIL-12. (b) Culture of the unresponsive  clones with IL-12 restores their proliferation. IL-12 was added to the culture  of the unresonsive clones during one of the biweekly restimulations. After 2  wk the IL-12 was washed away and proliferation of the clones was measured  as before. (c) The IL-12–treated clones return to their unresponsive state  upon removal of the IL-12 from culture. 2 wk after the treatment with IL-12, the IL-12 was washed away and the clones were passaged in culture without IL-12. After another 2 wk, proliferation was determined by DNA incorporation of [3H]thymidine.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2199180&req=5

Figure 6: (a) The addition of IL-12 increases proliferation of clone I to Hb(64–76) and 74L. Proliferation assay was performed as in Fig. 1, with or without the addition of 10 ng/ml rIL-12. (b) Culture of the unresponsive clones with IL-12 restores their proliferation. IL-12 was added to the culture of the unresonsive clones during one of the biweekly restimulations. After 2 wk the IL-12 was washed away and proliferation of the clones was measured as before. (c) The IL-12–treated clones return to their unresponsive state upon removal of the IL-12 from culture. 2 wk after the treatment with IL-12, the IL-12 was washed away and the clones were passaged in culture without IL-12. After another 2 wk, proliferation was determined by DNA incorporation of [3H]thymidine.
Mentions: The addition of exogenous costimulation has been used to overcome anergy in most of the model systems (9, 24, 25), but costimulation does not alter the anergic phenotype induced with partial agonist peptides (10, 17). Previous work with a Th2 clone demonstrated that a soluble factor (IL-1) could restore the proliferative response to a partial agonist peptide (3), and we have similarly observed that IL-12 boosts the proliferation of clone I to antigen (Fig. 6 A). In fact, a significant proliferative response was observed with both agonist peptide (0.1 μM) and partial agonist (10 μM 74L) by the addition of IL-12. To determine the effects of exogenous IL-12 on our anergic Th0 cells, 10 ng/ml recombinant IL-12 was added to the biweekly restimulations, and proliferation to Hb(64– 76) was assessed (Fig. 6 B). Inclusion of IL-12 in culture with the anergic T cells (but not in the proliferation assay) restored the proliferation of these lines to a similar level as clone I. However, the effect of IL-12 was transient, as the T cells retained their low proliferative response to Hb(64– 76) after subsequent passage (antigen, APCs, and IL-2) in the absence of exogenous IL-12 (Fig. 6 C). The increased proliferation resulting from IL-12 costimulation was not sufficient to reverse the anergic phenotype.

Bottom Line: The anergic state was characterized by blunted proliferation and interleukin (IL) 2 production upon restimulation with Hb(64-76), and was not the result of impaired TCR/CD3 downmodulation.Interestingly, persistence of the anergic phenotype was observed despite biweekly restimulation with antigen, APCs, and IL-2.Thus, T cell unresponsiveness induced by a peptide produced a stable, persistent anergic state in a Th0 clone that was not reversible by stimulation with IL-2 or -12.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Emory University, Atlanta, Georgia 30322, USA.

ABSTRACT
Clonal T cell unresponsiveness, or anergy, has been proposed as a mechanism of peripheral tolerance in vivo, and as a potential means of curbing unwanted T cell responses. In this study, anergy was induced in a T helper cell (Th) clone reactive to hemoglobin (Hb) peptide 64-76 by coculture of the T cells with live antigen-presenting cells (APCs) and 74L, a peptide analog of Hb(64-76) that contains a single amino acid substitution of leucine for glycine at position 74, or with a low concentration of the agonist ligand. The anergic state was characterized by blunted proliferation and interleukin (IL) 2 production upon restimulation with Hb(64-76), and was not the result of impaired TCR/CD3 downmodulation. The addition of exogenous IL-12 transiently restored proliferation of the anergic lines, but removal of IL-12 from culture returned the T cells to their nonproliferative state. Interestingly, persistence of the anergic phenotype was observed despite biweekly restimulation with antigen, APCs, and IL-2. Thus, T cell unresponsiveness induced by a peptide produced a stable, persistent anergic state in a Th0 clone that was not reversible by stimulation with IL-2 or -12.

Show MeSH