The JI
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     
 

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Milner, J. D.
Right arrow Articles by Hafler, D. A.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Milner, J. D.
Right arrow Articles by Hafler, D. A.
The Journal of Immunology, 1999, 163: 2522-2529.
Copyright © 1999 by The American Association of Immunologists

Differential Responses of Invariant V{alpha}24J{alpha}Q T Cells and MHC Class II-Restricted CD4+ T Cells to Dexamethasone1

Joshua D. Milner2,3,*, Sally C. Kent3,*, Timothy A. Ashley*, S. Brian Wilson{dagger}, Jack L. Strominger{dagger} and David A. Hafler4,*

* Center for Neurologic Diseases, Brigham and Women’s Hospital and Harvard Medical School, Boston, MA 02115; and {dagger} Department of Molecular and Cellular Biology, Harvard University, Cambridge, MA 02138, and Cancer Immunology and AIDS, Dana Farber Cancer Institute and Harvard Medical School, Boston, MA 02115


   Abstract
Top
 Abstract
Introduction
 Methods and Materials
 Results
 Discussion
 References
 
NK T cells are a T cell subset in the human that express an invariant {alpha}-chain (V{alpha}24invt T cells). Because of the well-described immunomodulation by glucocorticoids on activation-induced cell death (AICD), the effects of dexamethasone and anti-CD3 stimulation on V{alpha}24invt T cell clones and CD4+ T cell clones were investigated. Dexamethasone significantly enhanced anti-CD3-mediated proliferation of V{alpha}24invt T cells, whereas CD4+ T cells were inhibited. Addition of neutralizing IL-2 Ab partially abrogated dexamethasone-induced potentiation of V{alpha}24invt T cell proliferation, indicating a role for autocrine IL-2 production in corticosteroid-mediated proliferative augmentation. Dexamethasone treatment of anti-CD3-stimulated V{alpha}24invt T cells did not synergize with anti-Fas blockade in enhancing proliferation or preventing AICD. The V{alpha}24invt T cell response to dexamethasone was dependent on the TCR signal strength. In the presence of dexamethasone, lower doses of anti-CD3 inhibited proliferation of V{alpha}24invt T cells and CD4+ T cells; at higher doses of anti-CD3, which caused inhibition of CD4+ T cells, the V{alpha}24invt T cell clones proliferated and were rescued from AICD. These results demonstrate significant differences in TCR signal strength required between V{alpha}24invt T cells and CD4+ cells, and suggest important immunomodulatory consequences for endogenous and exogenous corticosteroids in immune responses.


   Introduction
Top
 Abstract
 Introduction
Methods and Materials
 Results
 Discussion
 References
 
T cells expressing the invariant V{alpha}24J{alpha}Q (V{alpha}24invt T cells) TCR with canonical rearrangements without N region additions are a recently described lymphocyte population in humans that are analogous to the murine V{alpha}14J{alpha}281 NK1 T cell (1, 2). They are restricted by CD1d, an Ag-presenting surface molecule that can present glycolipids to these invariant V{alpha}24invt T cells. Relatively high frequencies of V{alpha}24invt T cells, up to 1:100 of the total circulating T cell repertoire, can be observed in normal humans. They are of particular interest because of their rapid secretion of IL-4 and IFN-{gamma} within hours of TCR engagement, suggesting an important role in immunoregulation and possibly in Th1/Th2 differentiation (3, 4, 5, 6). Although the role of NK1 T cells in Th1/Th2 differentiation is under investigation, CD1 knockout mice lacking invariant V{alpha}14J{alpha}281 T cells exhibit alterations in anti-CD3-induced IL-4 secretion, although they are still capable of mounting a Th2 response (7).

There are a number of observations suggesting that invariant NK T cells may function in an important regulatory role in several models of autoimmunity (8, 9, 10, 11, 12). In humans with insulin-dependent diabetes mellitus (IDDM),5 a decrease in the frequency of V{alpha}24invt T cells in monozygotic twins discordant for IDDM was observed. Moreover, while the V{alpha}24invt T cell clones from nondiabetic siblings secreted IL-4 and IFN-{gamma}, the V{alpha}24invt T cells from subjects with IDDM secreted IFN-{gamma}, but not IL-4 (9). These alterations of V{alpha}24invt T cells clones from identical twins discordant for IDDM suggested that environmental events contributed to the alterations in cytokine secretion. Thus, it was of interest to examine nongenetic factors regulating the function of V{alpha}24invt, CD161a+ (NKR-P1A) T cells as compared with MHC class II-restricted CD4 T cells recognizing peptide Ags.

Among the major humoral factors that T cells encounter are endogenous glucocorticoids. Glucocorticoids have been shown to modulate Th1/Th2 cytokine secretion of T cells (13, 14, 15) and inhibit proinflammatory cytokines (16), resulting in an inhibition of proliferation (17, 18) with a wide range of effects on T cell activation. Glucocorticoids have been shown to significantly affect the disease course in several models of autoimmunity (19). For example, adrenalectomy of Lewis rats susceptible to experimental autoimmune encephalomyelitis leads to a progressive, fatal disease, whereas elevated plasma levels of glucocorticoids have been associated with spontaneous recovery from experimental autoimmune encephalomyelitis (20, 21). Thus, glucocorticoids have a clear influence on regulation of T cell function.

Unstimulated T cells normally undergo apoptosis in the presence of the synthetic glucocorticoid, dexamethasone (22). Paradoxically, AICD induced by TCR activation and dexamethasone-induced apoptosis antagonize each other in several systems (23, 24, 25, 26, 27). However, there appear to be discrepancies as to the effects of dexamethasone on activated T cells. Numerous groups have shown that dexamethasone significantly inhibits the proliferation of stimulated T cells (18, 28, 29, 30). Using T cell hybridomas, it was suggested that the reason proliferation was observed in some systems and inhibition in others was because of differences in the strength of TCR cross-linking (24). Stronger signals resulted in decreased cell death in the presence of increased amounts of dexamethasone, whereas weaker signals led to the opposite effect, proliferation.

Several mechanisms for the antagonism of dexamethasone-induced and AICD have been postulated involving both inhibition and induction of specific genes by dexamethasone. Dexamethasone has been shown to inhibit Fas ligand (31, 32), IL-2 production (17), and other activation-related genes. Additionally, GILZ, a leucine-zipper family gene, was identified as a dexamethasone-induced gene that, when transfected into T cell hybridomas, inhibited anti-CD3-induced apoptosis (33).

Finally, dexamethasone induction of cytokine receptors, notably IL-2R (26), has been proposed as an explanation for increased sensitivity to IL-2 during dexamethasone treatment, and hence increased proliferation in response to anti-CD3 in the presence of dexamethasone.

In a panel of CD4+, class II-restricted clones and V{alpha}24invt T clones derived from twins discordant for IDDM and additional subjects, proliferation of the CD4+ clones stimulated with anti-CD3 was inhibited by dexamethasone, whereas proliferation of the V{alpha}24invt T cell clones was augmented. This observation may be partially explained by markedly different sensitivities to AICD via strength of signal through the TCR between the two cell types, and may have implications for understanding peripheral modulation of the immune system by glucocorticoids.


   Methods and Materials
Top
 Abstract
 Introduction
 Methods and Materials
Results
 Discussion
 References
 
Cell culture

V{alpha}24invt. T cell clones (Me.10, Fc.13, Gw.4, Sb4.13) and CD4+ clones (Ob1.A12 and Ob3.E3) were generated as previously described (9, 34). CD4+ clone Sb3.D5 was generated from peripheral blood of subject Sb as a glutamic acid decarboxylase 65 (GAD)-specific clone in a manner similar to Wucherpfennig et al. (34). Briefly, PBMC purified by Ficoll-Hypaque separation were placed in media. Media was 10% autologous, heat-inactivated serum in RPMI 1640 with 10 mM HEPES buffer, 2 mM L-glutamine, and 100 U/100 µg/ml penicillin/streptomycin (all from BioWhittaker, Walkersville, MD) were pulsed with 30 µg/ml GAD for 3 h, and cells were plated at 200,000 cells/well. Whole, soluble human rGAD was generously provided by Dr. S. B. Wilson (Cancer Immunology and AIDS, Dana Farber Cancer Institute, Boston, MA). On day 7, autologous PBMC were pulsed with GAD, as described above, irradiated (5000 rad), and added at 75,000 cells/well. On day 9, 10 U/ml of human rIL-2 (Teceleukin, National Cancer Institute, Frederick, MD) was added to each well. On day 14, each well was split into four, and nonpulsed or GAD-pulsed autologous, irradiated PBMC (75,000/well) were added to two wells each of the split well. After 48 h, 1 µCi/well of [3H]thymidine was added and the wells were harvested after an additional 18 h. Wells judged positive for GAD reactivity had a stimulation index >3 and a {delta} cpm >1000. Positive wells were expanded on Ag, as described above for two to four more cycles. Cells were then cloned from positive wells at 0.3 cell/well with 100,000 irradiated, allogeneic feeders and 1 µg/ml PHA-P (Murex Biotech, Dartford, U.K.). Clone Sb.DP1 was generated from peripheral blood of subject Sb in an Ag-nonspecific manner by single cell-sorting CD4+ cells and grown as described (9). All clones were maintained (as described) in media that was changed every 2 to 3 days with addition of IL-2 (10 U/ml) and IL-7 (10 U/ml; Boehringer Mannheim, Indianapolis, IN). All clones were stimulated in the presence of 1 µg/ml PHA-P, IL-2, and IL-7 (10 U/ml each), irradiated, allogeneic feeders (50,000/well) and irradiated 722.221 lymphoblastoid cells (5,000/well). All experiments were performed 12 days after the last stimulation of the clones.

Proliferation assays

Clones were harvested and washed 12 days poststimulation. They were then plated at a density of 2–5 x 104 cells/well (three-six replicates of each condition, depending on the experiment) in 10% FCS in 96-well plates (Costar, Cambridge, MA) on plate-bound anti-CD3 (PharMingen, San Diego, CA) at 1 µg/ml, or in dilutions from 0.001–25 µg/ml of anti-CD3. For other experiments, cells were incubated on various concentrations of plate-bound anti-CD3 with additional factors added simultaneously to the cultures: concentrations (0.005–5000 nM in some experiments) of dexamethasone (Sigma, St. Louis, MO), anti-Fas-blocking mAb (1 µg/ml, 1/23; Boehringer Ingelheim, Ridgefield, CT), anti-IL-2-neutralizing Ab (2 µg/ml; R&D Systems, Minneapolis, MN), or IL-2 (100 U/ml). Supernatants were removed at 48 h, and cells were pulsed with 1 µ Ci/well [3H]thymidine and harvested and counted 18 h later on a beta scintillation counter (Wallace, Gaithersburg, MD).

Cell surface expression of TCR-{alpha}ß, Fas, and FasL

V{alpha}24invt clones and CD4+ T cell clones were assayed for TCR-{alpha}ß, Fas, and FasL expression by flow cytometry 12 days after restimulation. Cells (V{alpha}24invt clone, Gw.4 and Sb4.13; CD4+ T cell clones, Ob.3E3, Sb.DP1, and SbGAD.3D5) were stained with a pan {alpha}ß-TCR mAb or isotype-matched control Ab (Abs from Coulter-Immunotech, Miami, FL and PharMingen, respectively). V{alpha}24invt clone, Gw.4, and CD4+ T cell clone, Ob.3E3, were examined for Fas and FasL expression by flow cytometry. Twelve days after restimulation, clones were treated with 5 µg/ml soluble anti-CD3 (PharMingen) and 5 µg/ml F(ab')2 goat anti-mouse Ig (Sigma) with and without 5 nM dexamethasone and then stained for Fas expression (PharMingen) on day 0 (pretreatment), day 1, and day 2. For FasL expression, Gw.4 and Ob.3E3 T cell clones were treated with anti-CD3 and dexamethasone, as described for Fas expression, and with matrix metalloproteinase inhibitor KB8301 (PharMingen) at 10 µM for 2 h before staining. Cells were stained with biotin-mouse anti-human FasL and streptavidin-PE (PharMingen). Cells were analyzed by flow cytometry (FacSort; Becton Dickinson, San Jose, CA).

Cell death studies

V{alpha}24invt clones and CD4+ T cell clones were assayed for AICD by analyzing for DNA content by flow cytometry (35, 36). Cells were incubated with 10 µg/ml plate-bound anti-CD3 at a density of 5 x 104 cells/well in 10% FCS/RPMI. Dexamethasone (5 nM) and/or 2 µg/ml anti-Fas mAb were added simultaneously. After 48 h, cells were washed and resuspended in 0.3% saponin, 50 µg/ml DNase-free RNase, 500 mM EDTA, and 5 µg/ml propidium iodide (all from Sigma) for 30 min at room temperature. Cells were washed and DNA content was analyzed using flow cytometry. Cells with a DNA content of less than that seen in the G1 cell cycle peak were considered apoptotic.


   Results
Top
 Abstract
 Introduction
 Methods and Materials
 Results
Discussion
 References
 
V{alpha}24invt T cell proliferation induced by anti-CD3 is enhanced by dexamethasone, whereas CD4+ clones are inhibited

A panel of T cell clones that were either CD1d restricted and expressed the invariant V{alpha}24J{alpha}Q TCR {alpha}-chain or were CD4+, MHC class II restricted, and expressed noninvariant TCR sequences was generated. This panel included: three IL-4-secreting V{alpha}24invt T cell clones derived from individuals without IDDM; one non-IL-4-secreting V{alpha}24invt T cell clone from a subject with IDDM; two MBP-reactive, CD4+ clones; one GAD-reactive CD4+ T cell clone; and one non-Ag-specific CD4+ clone. The latter two clones and one of the IL-4-secreting V{alpha}24invt T cell clones were generated from the same patient without IDDM to show that this phenomenon was not the result of interindividual differences. The T cell clones were maintained and stimulated identically.

To examine whether culture of T cells with dexamethasone had effects on proliferation, as measured by thymidine incorporation, cultures were stimulated with 1 µg/ml anti-CD3 in the presence or absence of 5 nM dexamethasone. As expected, the proliferation of all MHC class II-restricted CD4+ clones was significantly inhibited by dexamethasone (Fig. 1GoA). In marked contrast, anti-CD3-induced proliferation of all CD1d-restricted V{alpha}24invt clones was potentiated by the dexamethasone (Fig. 1GoB). There were no significant differences in response to dexamethasone between IL-4- and non-IL-4-secreting V{alpha}24invt T cell clones. The effect of varying dexamethasone concentrations was examined on CD1d-restricted V{alpha}24invt T cell and MHC class II-restricted T cell clones. Again, while dexamethasone significantly inhibited proliferation of the MHC class II-restricted CD4+ clone, it significantly augmented the proliferation of the CD1d-restricted V{alpha}24invt T cell clone in a dose-dependent manner (Fig. 2Go). To ensure that lower levels of endogenous glucocorticoids present in serum did not contribute to the T cell response, the experiments were repeated in serum-free media with concentrations of dexamethasone as low as 5 x 10-12 M. The results were identical to that obtained with serum in the media (data not shown).



View larger version (25K):
[in this window]
[in a new window]
  		FIGURE 1. Dexamethasone augments V{alpha}24invt T cell proliferation, but inhibits CD4+ proliferation. A, Four CD4+ T cell clones were derived from a nonprogressor IDDM- individual (Sb.DP1, Sb3.D5) and a control (Ob1.A12, Ob3.E3). Ob1.A12 and Ob3.E3 are specific for MBP p85–99, Sb3.D5 for GAD65, and Sb.DP1 was cloned in an Ag-nonspecific manner. Sb4.13, Sb.DP1, and Sb3.D5 clones all come from the same individual. These clones and four V{alpha}24invt T cell clones (B) from nonprogressor IDDM- (Fc.13, Gw.4, Sb4.13) and IDDM+ (Me.10) individuals were stimulated with 1 µg/ml plate-bound anti-CD3 (50,000/well) in the absence or presence of 5 nM dexamethasone for 48 h, then pulsed for 18 h with [3H]thymidine to measure proliferation. All anti-CD3-induced V{alpha}24invt T cell proliferation was augmented by dexamethasone, whereas the anti-CD3-induced proliferation of the MHC class II-restricted CD4+ T cell clones was inhibited by dexamethasone. This is a representative experiment of three similar experiments; SEs are less than 10% for each data point and not shown.

 


View larger version (15K):
[in this window]
[in a new window]
  		FIGURE 2. Dexamethasone inhibition of CD4+ T cells and augmentation of proliferation of V{alpha}24invt T cells are dose dependent. As in Fig. 1Go, clones were stimulated in the presence of 0.05–5000 nM dexamethasone on 1 µg/ml plate-bound anti-CD3. As a control for each dexamethasone dilution, the V{alpha}24invt T cell clone (Gw.4) was incubated in equal amounts of ethanol, as ethanol was the vehicle for dexamethasone. The proliferation of the CD4+ T cell clone (Ob1.A12) was inhibited at all dexamethasone concentrations, whereas the anti-CD3-induced proliferation of the V{alpha}24invt T cell clone was augmented at high dexamethasone concentrations. This is a representative experiment of three similar experiments; SEs are less than 10% and not shown. The same responses were seen with several other CD4+ and V{alpha}24invt T cell clones (data not shown).

 
Autocrine IL-2 secretion is not the sole mechanism responsible for augmentation of proliferation of V{alpha}24invt T cells by dexamethasone

A potential mechanism for the dexamethasone-induced augmentation of proliferation by CD1d-restricted V{alpha}24invt T cells vs the inhibition of proliferation by MHC class II-restricted CD4+ T cells may involve autocrine IL-2 secretion (26). The rescue of MHC class II-restricted CD4+ T cells from dexamethasone-induced inhibition of proliferation by the addition of exogenous IL-2 to the culture was examined (Fig. 3Go). Although there was a marked enhancement in TCR-mediated proliferation without dexamethasone in the CD4+ T cell clone with addition of exogenous IL-2, significant inhibition of proliferation was observed when increasing concentrations of dexamethasone were added to the culture. In contrast, there was no significant response of CD1d-restricted V{alpha}24invt T cells to exogenous IL-2 with increasing concentrations of dexamethasone.



View larger version (20K):
[in this window]
[in a new window]
  		FIGURE 3. Exogenous IL-2 protects CD4+ T cells from inhibition of proliferation by dexamethasone and has no effect on dexamethasone-augmented proliferation of V{alpha}24invt T cells. V{alpha}24invt (Gw.4) and CD4+ T cell clones (Ob.3E3) were incubated on 1 µg/ml plate-bound anti-CD3 and concentrations of dexamethasone (as in Fig. 1Go) in the presence or absence of 100 U/ml IL-2. The CD4+ T cell clone was protected from anti-CD3/dexamethasone-induced proliferation inhibition by IL-2, but exogenous IL-2 had no effect on the dexamethasone-enhanced proliferation of the V{alpha}24invt T cell clone. This is a representative experiment of three similar experiments; SEs are less than 10% and not shown.

 
V{alpha}24invt T cells secrete large amounts of IL-2, which may protect them from dexamethasone-induced apoptosis (37, 38, 39). Anti-IL-2-neutralizing Ab was added to the culture of T cells stimulated by TCR cross-linking at either 0.1 µg/ml (Fig. 4GoA) or 1 µg/ml of anti-CD3 (Fig. 4GoB) with increasing concentrations of dexamethasone. The proliferation of V{alpha}24invt T cells induced with 1 µg/ml of anti-CD3 was moderately inhibited by anti-IL-2, although dexamethasone still moderately potentiated proliferation. Interestingly, at the lower concentration of 0.1 µg/ml anti-CD3, neutralization of autocrine-secreted IL-2 resulted in suppression of proliferation at increasing concentrations of dexamethasone (Fig. 4Go). To determine whether increased autocrine secretion of IL-2 affected the efficiency of IL-2 neutralization, IL-2 production by V{alpha}24invt T cell clones stimulated at both 0.1 and 1 µg/ml anti-CD3 was measured by ELISA. Although increased amounts of IL-2 produced at 1 µg/ml anti-CD3 that was not neutralized by anti-IL-2 mAb could be a potential explanation for sustained dexamethasone-mediated proliferation, the Ab concentrations used exceed the neutralization capacity of the IL-2 concentrations produced in these experiments (data not shown). These data suggest that at lower TCR signal strength, IL-2 may be required to potentiate dexamethasone-induced proliferation, whereas at higher TCR signal strength, enhancement of proliferation by dexamethasone is less IL-2 dependent.



View larger version (22K):
[in this window]
[in a new window]
  		FIGURE 4. Autocrine IL-2 production alone does not appear to be responsible for V{alpha}24invt T cells’ response to dexamethasone at higher levels, but does at lower levels of anti-CD3 stimulation. V{alpha}24invt T cell clones (Gw.4) were stimulated with 0.1 µg/ml (A) or 1 µg/ml (B) plate-bound anti-CD3 and concentrations of dexamethasone (as in Fig. 1Go) in the presence or absence of excess (2 µg/ml) anti-IL-2-neutralizing Ab. At 0.1 µg/ml CD3, addition of anti-IL-2 inhibited dexamethasone-augmented proliferation of the V{alpha}24invt T cell clone, whereas at 1 µg/ml anti-CD3, addition of anti-IL-2 did not completely inhibit dexamethasone-augmented V{alpha}24invt T cell clone proliferation. This is a representative experiment of four similar experiments; SEs are less than 10% and not shown.

 
Dexamethasone inhibited apoptosis of CD1d-restricted V{alpha}24invt T cells and induced apoptosis of MHC class II-restricted CD4+ cells

Increases in proliferation induced by dexamethasone in the V{alpha}24invt T cells may have been due to inhibition of cell death. This was examined by measuring the percentage of T cells undergoing cell death after TCR cross-linking, as determined by propidium iodide staining, in the presence of increasing concentrations of dexamethasone. As shown in Fig. 5Go, signals through the TCR with anti-CD3 mAb induced apoptosis of V{alpha}24invt T cells, which was blocked by anti-Fas mAb blockade, indicating AICD. The inhibition of AICD by dexamethasone was examined (Fig. 5Go). Increasing concentrations of dexamethasone blocked the induction of AICD in the V{alpha}24invt T cells while inducing significant levels of apoptotic cell death in the CD4+ T cell clone, and this was not affected by Fas blockade. Increasing concentrations of dexamethasone in the presence of anti-Fas blockade did not further inhibit apoptosis in the CD4+ T cell clone.



View larger version (18K):
[in this window]
[in a new window]
  		FIGURE 5. Dexamethasone rescues V{alpha}24invt T cells from AICD, but causes cell death in CD4+ T cell clones. V{alpha}24invt (Gw.4) and CD4+ T cell clones (Ob.3E3) were stimulated with 10 µg/ml plate-bound anti-CD3 with increasing amounts of dexamethasone in the presence or absence of anti-Fas (2 µg/ml)-blocking mAb. Cells were harvested after 48 h, washed, and stained with propidium iodide to detect cell death. Percentage represents fraction of gated cells stained at <2n DNA content. This experiment was repeated three times and a representative experiment is shown; SEs are less than 10% and not shown.

 
CD1d-restricted V{alpha}24invt T cells are highly sensitive to AICD; dexamethasone does not enhance the increase in proliferation caused by Fas blockade

A potential mechanism for the antagonism of AICD by dexamethasone is the down-regulation of FasL by dexamethasone (32). If the AICD-preventative action of dexamethasone acted exclusively through a mechanism independent of the Fas-mediated pathway, one would expect to see a synergistic effect on proliferation between Fas blockade and dexamethasone. The addition of anti-Fas-blocking mAb produced a marked increase in proliferation of anti-CD3-stimulated V{alpha}24invt T cell clones, but the combination of anti-Fas mAb and dexamethasone was not as effective as Fas blockade alone in augmenting proliferation (Fig. 6Go). In fact, in the presence of anti-Fas mAb, the V{alpha}24invt T cell clone proliferation was inhibited at higher dexamethasone concentrations. Similarly, anti-Fas-blocking Ab prevented AICD of V{alpha}24invt T cells, and this effect was not potentiated by dexamethasone (Fig. 5Go). The addition of anti-Fas blockade affected the proliferative capacity of the MHC class II-restricted CD4+ T cells only at lower concentrations of dexamethasone (0.05 and 0.5 nM). The proliferation of V{alpha}24invt T cells in the presence of anti-Fas mAb alone and anti-Fas mAb and dexamethasone was dramatically augmented (Fig. 7Go, A and B).



View larger version (15K):
[in this window]
[in a new window]
  		FIGURE 6. Dexamethasone and Fas blockade did not synergize to enhance V{alpha}24invt T cell proliferation in the presence of anti-CD3. V{alpha}24invt T cell clones (Gw.4) were stimulated with increasing concentrations of plate-bound anti-CD3 in the presence or absence of 5 nM dexamethasone and/or an excess (1 µg/ml) of anti-Fas-blocking Ab. The combination of anti-Fas and dexamethasone did not synergize to enhance proliferation of the V{alpha}24invt T cell clone. Anti-CD3 dose response of V{alpha}24invt T cell clone to dexamethasone/anti-Fas indicated that Fas blockade enhanced proliferation of the V{alpha}24invt T cell clone at high doses of anti-CD3, and dexamethasone inhibits this effect. This is a representative experiment of four similar experiments; SEs are less than 10% and not shown.

 


View larger version (24K):
[in this window]
[in a new window]
  		FIGURE 7. V{alpha}24invt T cell proliferation is enhanced in the presence of dexamethasone and anti-Fas mAb, but CD4+ T cell clone proliferation is not. At 1 µg/ml plate-bound anti-CD3 and increasing amounts of dexamethasone, the proliferation of the V{alpha}24invt T cell clone (Gw.4) is enhanced by Fas ligation (A), whereas Fas ligation could not maintain proliferation of the CD4+ T cell clone (Ob.3E3) in the presence of dexamethasone (B). This is a representative experiment of two similar experiments; SEs are less than 10% and not shown.

 
We then directly examined Fas and FasL expression levels under various stimulation conditions on V{alpha}24invt T cells and CD4+ cells (Fig. 8Go). Anti-CD3 stimulation in the absence or presence of dexamethasone had no effect on Fas expression of either CD4+ T cells or V{alpha}24invt T cells (Fig. 8GoA). In contrast, anti-CD3 stimulation resulted in an increase of FasL expression of both CD4+ and V{alpha}24invt T cells, which was down-regulated by dexamethasone (Fig. 8GoB). Taken together these data suggested that dexamethasone is exerting its effect at least in part by disrupting the Fas-mediated AICD pathway.



View larger version (46K):
[in this window]
[in a new window]
  		FIGURE 8. Effect of anti-CD3 cross-linking and dexamethasone on Fas and FasL expression on V{alpha}24invt clones and CD4+ T cell clones. V{alpha}24invt clone Gw.4 and CD4+ T cell clone Ob.3E3 were examined for Fas and FasL expression by flow cytometry. Twelve days after restimulation, clones were treated with 5 µg/ml soluble anti-CD3 and 5 µg/ml F(ab')2 goat anti-mouse Ig with and without 5 nM dexamethasone and then stained for Fas expression or FasL expression, as described in Materials and Methods. A, Both the V{alpha}24invt T cell clone (Gw.4) and the CD4+ T cell clone (Ob.3E3) express Fas (bold line) similarly; addition of anti-CD3/dexamethasone does not effect Fas expression (isotype control is non-bolded line). B, FasL is not expressed at rest, but becomes up-regulated in the presence of anti-CD3 and down-regulated in the presence of anti-CD3 and dexamethasone in both clones. This is a representative experiment of five similar experiments.

 
Strength of signal determines susceptibility to dexamethasone

A major functional attribute of the CD1d-restricted V{alpha}24invt T cells is rapid secretion of cytokines with TCR cross-linking as compared with MHC class II-restricted CD4+ T cells (40). This suggested that a very strong strength of signal was mediated through the V{alpha}24invt TCR, as compared to CD4+ T cells. One factor affecting TCR signal strength is TCR density. A panel of CD4+ clones and V{alpha}24invt T cell clones was assayed for TCR-{alpha}ß expression on day 0 (12 days after restimulation). TCR-{alpha}ß expression on the two types of T cell clones was similar (Fig. 9Go). This suggested that the strength of the TCR-mediated signal of the V{alpha}24invt T cells may be greater as compared with CD4+ T cells; this difference may explain the effect of dexamethasone on the enhancement of proliferation and protection from cell death in the V{alpha}24invt T cells. This hypothesis was directly investigated by examining the effect of dexamethasone on proliferation with different concentrations of anti-CD3 mAb. As predicted for MHC class II-restricted CD4+ T cell clones, increasing concentrations of anti-CD3 induced greater thymidine incorporation, which was inhibited by increasing concentrations of dexamethasone (Fig. 10GoA). Surprisingly, the effect of dexamethasone on proliferation of V{alpha}24invt T cells at lower concentrations of anti-CD3 duplicated the inhibitory effects of dexamethasone observed on MHC class II-restricted CD4 T cells. Thus, in the presence of dexamethasone, at 0.01 µg/ml anti-CD3, V{alpha}24invt T cells were inhibited. The 1.0 µg/ml of anti-CD3, which allowed dexamethasone-mediated inhibition of proliferation of the CD4+ T cell clone, was in the range of maximal proliferative response to anti-CD3 for the V{alpha}24invt T cell clone. In general, higher doses of anti-CD3 resulted in high dose suppression of V{alpha}24invt T cells, presumably by AICD, whereas lower doses did not induce any proliferative response (Fig. 10GoB). This suggests that for a given anti-CD3 stimulus, the strength of signal via theV{alpha}24invt TCR is significantly stronger than those in CD4+ TCR and is unrelated to the cell surface density of the TCR.



View larger version (24K):
[in this window]
[in a new window]
  		FIGURE 9. TCR expression levels on V{alpha}24invt clones and CD4+ T cell clones. V{alpha}24invt clones (Gw.4, Sb4.13) and CD4+ T cell clones (Ob.3E3, Sb.DP1, SbGAD.3D5) were assayed for TCR-{alpha}ß expression by flow cytometry 12 days after restimulation. Cells were stained with a pan {alpha}ß-TCR mAb (bold line) or isotype-matched control Ab (non-bolded line). The TCR-{alpha}ß expression at rest among the V{alpha}24invt clone (Gw.4) and the CD4+ T cell clone (Ob.3E3) is similar. The TCR-{alpha}ß expression of V{alpha}24invt clone SB4.13 was greater than other clones tested. Upon anti-CD3 cross-linking, all clones down-regulate TCR-{alpha}ß, as expected (not shown). This is a representative experiment of five similar experiments.

 


View larger version (27K):
[in this window]
[in a new window]
  		FIGURE 10. Dexamethasone only augments proliferation of V{alpha}24invt T cells undergoing high dose suppression. The CD4+ T cell clone Ob.3E3 (A) and the V{alpha}24invt T cell clone Gw.4 (B) were stimulated with increasing amounts of plate-bound anti-CD3 and increasing amounts of dexamethasone (as in Fig. 1Go). V{alpha}24invt T cells proliferate at lower anti-CD3 concentrations (0.01 µg/ml) than did the CD4+ T cell clone, with no dexamethasone added, and ~2 logs more dexamethasone was required to inhibit proliferation of the V{alpha}24invt T cell clone as compared with the CD4+ T cell clone. At anti-CD3 concentrations of 0.1 and 1 µg/ml, addition of dexamethasone inhibited proliferation of the CD4+ T cell clone, whereas the V{alpha}24invt T cell clone is enhanced for proliferation. This is a representative experiment of three similar experiments; SEs are less than 10% and not shown.

 

   Discussion
Top
 Abstract
 Introduction
 Methods and Materials
 Results
 Discussion
References
 
CD1d-restricted V{alpha}24invt T cells with invariant TCR {alpha}-chains represent a potentially major functional population of T cells in humans. Unlike MHC class II-restricted CD4+ T cells, V{alpha}24invt T cells circulate in an activated state, as evidenced by medium affinity IL-2R, and rapidly secrete cytokines with TCR cross-linking (41). This suggests that CD1d-restricted V{alpha}24invt T cells have an important early function in regulating immune responses. Corticosteroids are rapidly secreted into the circulation with infections and are a major early regulator of inflammatory responses (42). Thus, the response of V{alpha}24invt T cells to corticosteroids may be important in directing the immune response. In this study, the biologic effects of the synthetic corticosteroid, dexamethasone, on V{alpha}24invt T cells were compared with MHC class II-restricted CD4+ T cells. Although dexamethasone markedly inhibited proliferation by enhancing CD3-mediated AICD in CD4+ T cell clones, dexamethasone enhanced proliferation by inhibiting CD3-mediated AICD in V{alpha}24invt T cells. The mechanism of enhanced proliferation in V{alpha}24invt T cells was in part related to higher strengths of signals provided through the TCR.

Dexamethasone can either induce or protect from cell death, and a balance of factors is likely to dictate the response of a stimulated T cell to dexamethasone. These factors include the signal delivered through the TCR, induction of genes involved in AICD, apoptotic signals induced by dexamethasone, and interference with AICD genes by dexamethasone. These counterbalancing factors are all likely to be involved in determining the outcome of T cell activation in the presence of corticosteroids. Based on our observations, it is possible to conclude that the discrepancies in the literature regarding the effects of dexamethasone on stimulated freshly isolated T cells, hybridomas, or splenocytes may be explained by the differences in activation states, or individual responses to TCR engagement.

The augmented V{alpha}24invt T cell proliferation induced by dexamethasone with anti-CD3 was observed only in the signal strength range of stimulation that induced TCR-mediated AICD. In contrast, CD4+ clones were inhibited by dexamethasone at concentrations of anti-CD3 250 times those that resulted in dexamethasone augmentation for V{alpha}24invt T cell clones. Markedly increased signals through the TCR antagonize the dexamethasone-induced cell death in hybridomas (24), and this antagonism is maximal in cells undergoing vigorous AICD. In those studies, weaker signals through the TCR were incapable of overriding the dexamethasone-mediated death pathway, similar to the results of our own investigation of MHC class II-restricted CD4+ clones and in V{alpha}24invt T cell clones at the very lowest doses of anti-CD3. It remains unclear whether increased proliferation of stimulated V{alpha}24invt T cells in the presence of dexamethasone is directly linked to the inhibition of AICD, or whether the presence of AICD without dexamethasone is simply an accurate predictor of increased proliferation with dexamethasone addition.

Because the CD4+ and NK T cell clones were cloned, maintained, and stimulated identically and predominantly from the same individual, variations among individual sources and cloning techniques that could affect activation states were controlled. Although dexamethasone-induced enhancement of proliferation mediated by protection from AICD was demonstrated, as related to the strength of signal through the TCR, the biochemical signals regulating the protective effect are as yet unknown. Dexamethasone can indirectly inhibit AICD through induction of the protective gene GILZ, and dexamethasone can regulate FasL expression (31, 32, 33). This may occur through the transcription factors, Egr-3 (43) and Egr-2 (44); these factors may play a role in T cell survival in the presence of glucocorticoids. Future experiments will examine these pathways of activation.

Although our data are consistent with previous work demonstrating that dexamethasone acts upon the Fas-mediated death pathway (31, 32), these experiments do not exclude actions by dexamethasone on other death or antiapoptosis pathways. Dexamethasone treatment alone of anti-CD3-stimulated V{alpha}24invt T cells does not reach the levels of proliferation or AICD inhibition induced by anti-Fas blockade. Additionally, dexamethasone treatment does not synergize with Fas blockade, in fact they appear to be somewhat antagonistic (Figs. 6Go and 7Go). There are two possible explanations for these observations: 1) the effects of dexamethasone are exclusive to the Fas-induced AICD pathway either at the level of FasL production or further downstream and 2) dexamethasone may also be acting on other AICD pathways with its effects not as pronounced as Fas blockade; any synergy would be obscured by the effects of Fas blockade. Thus, concurrent inhibition of cellular growth or induction of other apoptotic genes by dexamethasone may explain the observed antagonism between dexamethasone and Fas blockade. Further studies examining the role of these mechanisms in V{alpha}24invt T cell activation are important, as they may differ from those found in MHC class II-restricted CD4+ cells.

Another potential mechanism for the contrasting responses of MHC class II-restricted CD4+ T cells and CD1d-restricted V{alpha}24invt T cells to dexamethasone was autocrine IL-2 production (26, 37, 38, 39). Neutralization of IL-2 production with anti-IL-2 mAb affected the augmentation of proliferation by dexamethasone only when the V{alpha}24invt T cells were stimulated at low concentrations of anti-CD3. This may suggest that autocrine IL-2 production plays a role in weakly stimulated V{alpha}24invt T cells. However, while exogenous IL-2 addition provided the CD4+ T cell clones protection from dexamethasone-induced inhibition, it did not cause dexamethasone-mediated enhancement of proliferation. Whether these observations were results of differential susceptibilities of the cell types to dexamethasone-mediated death and AICD, or due to other protective autocrine-secreted molecules is unknown.

The increased activation state of V{alpha}24invt T cells resulting in relative ease of activation and AICD is consistent with other phenotypic characteristics described of NK T cells. Peripheral NK T cells express an array of activation markers (45), very rapidly produce cytokines when stimulated in vivo (40), and are clonally expanded, possessing a canonical TCR {alpha} rearrangement at high frequencies in the peripheral blood. This clonal expansion can occur in germfree conditions (46), consistent with the possibility that there is tonic activation of NK1.1+ cells in the periphery. It remains to be confirmed whether this higher state of activation is due to increased tonic stimulation, or to a stronger signal sent through the V{alpha}24invt TCR as compared with other TCRs for a given stimulus. Regardless, increased susceptibility of V{alpha}24invt T cells to dexamethasone augmentation of proliferation further suggests an in vivo mechanism for the clonal expansion of these cells.

Recently, it was demonstrated that injection of dexamethasone into mice results in increased frequency and number of NK T cells. The results were in part due to the fact that NK T cells are more resistant to dexamethasone- and radiation-induced cell death (47). In those studies, NK T cell proliferation, as measured by BrdU (5-bromo-2'-deoxyuridine) incorporation, could not explain the increased frequency and raw number of NK T cells derived from the liver after dexamethasone injection. The authors suggest this may be due to migration of cells to the liver. Proliferation may be taking place at a separate site such as the spleen or lymph nodes, resulting in increased numbers when these cells reach the liver. It was also shown that Fas-mediated apoptosis is irrelevant to the preferential survival of murine NK1 T cells by showing that Fas levels on NK1+ and NK1- cells are unchanged with dexamethasone treatment. These results are not surprising because FasL expression on lymphocytes, not Fas, is most affected in vitro studies (31, 32).

Previous investigations of T cell selection in the thymus may provide a model for the shaping of the peripheral T cell repertoire by glucocorticoids (24). Increased TCR signals result in an increased probability that glucocorticoids will enhance proliferation. Therefore, during acute stressful events such as infection when glucocorticoid levels increase, there is a positive selection for the highest affinity T cells and a negative selection for the lower affinity T cells, providing an efficient repertoire to clear infection. Tonic glucocorticoid stimulation without adequate Ag presentation would result in general T cell inhibition, which may explain why chronic infection can impair the immune response.

Our results may help further explain important aspects of immune regulation as they pertain to tolerance and autoimmunity. Increases or decreases in V{alpha}24invt T cell frequency may have direct impact on the control of immune response and prevention of autoimmunity (9, 10, 11, 12, 45, 46, 47). Because clonal expansion of CD1d-stimulated V{alpha}24invt T cells is augmented by glucocorticoids, glucocorticoid-mediated expansion of high affinity CD4+ T cells potentially would be accompanied by expansion of recently activated V{alpha}24invt T cells at a sufficient rate to potentially regulate autoreactive T cell activity. In contrast, the presence of circulating corticosteroids in the absence of CD1d activation of V{alpha}24invt T cells could potentially reduce the frequency of these regulatory T cells. Thus, circulating corticosteroids in situations of inflammation where there is increased CD1d stimulation of V{alpha}24invt T cells may potentially regulate the nature of the T cell-mediated immune response.


   Acknowledgments
 
We thank Dr. Betty Diamond and Dr. Byron Waksman for critical reading of the manuscript.


   Footnotes
 
1 These studies were supported by grants (to D.A.H.) from the National Institutes of Health (R01 AI39229-01, P01 AI39671, and R01 AI44447-01) and by a grant (to D.A.H.) from the Juvenile Diabetes Foundation International (1-1998-124). J.D.M. was funded by a Howard Hughes Medical Institute Medical Student Research Training Fellowship. Back

2 Current address: Albert Einstein College of Medicine, Bronx, NY 10461. Back

3 J.D.M. and S.C.K. contributed equally to this manuscript. Back

4 Address correspondence and reprint requests to Dr. David A. Hafler, Center for Neurologic Diseases, 77 Avenue Louis Pasteur, Boston, MA 02115. E-mail address: Back

5 Abbreviations used in this paper: IDDM, insulin-dependent diabetes mellitus; AICD, activation-induced cell death; GAD, glutamic acid decarboxylase 65. Back

Received for publication December 11, 1998. Accepted for publication June 16, 1999.


   References
Top
 Abstract
 Introduction
 Methods and Materials
 Results
 Discussion
 References
 

  1. Bendelac, A., O. Lantz, M. E. Quimby, J. W. Yewdell, J. R. Bennink, R. R. Brutkiewicz. 1995. CD1 recognition by mouse NK1+ T lymphocytes. Science 268:863.[Abstract/Free Full Text]
  2. Porcelli, S., C. E. Yockey, M. B. Brenner, S. P. Balk. 1993. Analysis of T cell antigen receptor (TCR) expression by human peripheral blood CD4-8- {alpha}/ß T cells demonstrates preferential use of several Vß genes and an invariant TCR {alpha} chain. J. Exp. Med. 178:1.[Abstract/Free Full Text]
  3. Rooney, J., T. Hoey, L. Glimcher. 1995. Coordinate and cooperative roles for NF-AT and AP-1 in the regulation of the murine IL-4 gene. Immunity 2:473.[Medline]
  4. Tamada, K., M. Harada, K. Abe, T. Li, H. Tada, Y. Onoe, K. Nomoto. 1997. Immunosuppressive activity of cloned natural killer (NK1.1+) T cells established from murine tumor-infiltrating lymphocytes. J. Immunol. 158:4846.[Abstract]
  5. Yoshimoto, T., A. Bendelac, C. Watson, J. Hu-Li, W. E. Paul. 1995. Role of NK1.1+ T cells in a Th2 response and in immunoglobulin E production. Science 270:1845.[Abstract/Free Full Text]
  6. Yoshimoto, T., A. Bendelac, J. Hu-Li, W. E. Paul. 1995. Defective IgE production by SJL mice is linked to the absence of CD4+, NK1.1+ T cells that promptly produce interleukin 4. Proc. Natl. Acad. Sci. USA 92:11931.[Abstract/Free Full Text]
  7. Brown, D. R., D. J. Fowell, D. B. Corry, T. A. Wynn, N. H. Moskowitz, A. W. Cheever, R. M. Locksley, S. L. Reiner. 1996. ß2-Microglobulin-dependent NK1.1+ T cells are not essential for T helper cell 2 immune responses. J. Exp. Med. 184:1295.[Abstract/Free Full Text]
  8. Sumida, T., A. Sakamoto, H. Murata, Y. Makino, H. Takahashi, S. Yoshida, K. Nishioka, I. Iwamoto, M. Taniguchi. 1995. Selective reduction of T cells bearing invariant V{alpha}24J{alpha}Q antigen receptor in patients with systemic sclerosis. J. Exp. Med. 182:1163.[Abstract/Free Full Text]
  9. Wilson, S. B., S. C. Kent, K. T. Patton, T. Orban, R. A. Jackson, M. Exley, S. Porcelli, D. A. Schatz, M. A. Atkinson, S. P. Balk, et al 1998. Extreme Th1 bias of invariant V{alpha}24J{alpha}Q T cells in type 1 diabetes. Nature 391:177.[Medline]
  10. Mieza, M. A., T. Itoh, J. Q. Cui, Y. Makino, T. Kawano, K. Tsuchida, T. Koike, T. Shirai, H. Yagita, A. Matsuzawa, et al 1996. Selective reduction of V{alpha}14+ NK T cells associated with disease development in autoimmune-prone mice. J. Immunol. 156:4035.[Abstract]
  11. Hammond, K. J. L., L. D. Poulton, L. J. Palmisano, P. A. Silveira, D. I. Godfrey, A. G. Baxter. 1998. {alpha}/ß-T cell receptor (TCR)+CD4-CD8- (NKT) thymocytes prevent insulin-dependent diabetes mellitus in nonobese diabetic (NOD)/Lt mice by the influence of interleukin (IL)-4 and/or IL-10. J. Exp. Med. 187:1047.[Abstract/Free Full Text]
  12. Baxter, A. G., S. J. Kinder, K. J. L. Hammond, R. Scollay, D. I. Godfrey. 1997. Association between {alpha}ßTCR+CD4-CD8- T-cell deficiency and IDDM in NOD/Lt mice. Diabetes 46:572.[Abstract]
  13. Padgett, D., J. Sheridan, R. Loria. 1995. Steroid hormone regulation of a polyclonal TH2 immune response. Ann. NY Acad. Sci. 774:323.[Medline]
  14. Ramirez, F., D. Fowell, M. Puklavec, S. Simmonds, D. Mason. 1996. Glucocorticoids promote a Th2 cytokine response by CD4+ T cells in vitro. J. Immunol. 156:2406.[Abstract]
  15. Dozmorov, I., R. Miller. 1998. Generation of antigen-specific TH2 cells from unprimed mice in vitro: effects of dexamethasone and anti-IL-10 antibody. J. Immunol. 160:2700.[Abstract/Free Full Text]
  16. Rook, G., R. Hernandez-Pando, S. Lightman. 1994. Hormones, peripherally activated prohormones and regulation of the Th1/Th2 balance. Immunol. Today 15:301.[Medline]
  17. Gillis, S., G. Crabtree, K. Smith. 1979. Glucocorticoid-induced inhibition of T cell growth factor production. I. The effect on mitogen-induced lymphocyte proliferation. J. Immunol. 123:1624.[Abstract/Free Full Text]
  18. Almawi, W., M. Lipman, A. Stevens, B. Zanker, E. Hadro, T. Strom. 1991. Abrogation of glucocorticoid-mediated inhibition of T cell proliferation by the synergistic action of IL-1, IL-6 and IFN-{gamma}. J. Immunol. 146:3523.[Abstract]
  19. Wilder, R.. 1995. Neuroendocrine-immune system interactions and autoimmunity. Annu. Rev. Immunol. 13:307.[Medline]
  20. MacPhee, I., F. Antoni, D. Mason. 1989. Spontaneous recovery of rats from experimental allergic encephalomyelitis is dependent on regulation of the immune system by endogenous adrenal corticosteroids. J. Exp. Med. 169:431.[Abstract/Free Full Text]
  21. Mason, D., I. Macphee, F. Antoni. 1990. The role of the neuroendocrine system in determining genetic susceptibility to experimental allergic encephalomyelitis in the rat. Immunology 70:1.[Medline]
  22. Wyllie, A.. 1980. Glucocorticoid-induced thymocyte apoptosis is associated with endogenous endonuclease activation. Nature 284:555.[Medline]
  23. Zacharchuk, C. M., M. Mercep, P. K. Chakraborti, S. S. Simons, J. D. Ashwell. 1990. Programmed T lymphocyte death: cell activation- and steroid- induced pathways are mutually antagonistic. J. Immunol. 145:4037.[Abstract]
  24. Iwata, M., S. Hanaoka, K. Sato. 1991. Rescue of thymocytes and T cell hybridomas from glucocorticoid-induced apoptosis by stimulation via the T cell receptor/CD3 complex: a possible in vitro model for positive selection of the T cell repertoire. Eur. J. Immunol. 21:643.[Medline]
  25. Pelfrey, C. M., L. R. Tranquill, S. A. Boehme, H. F. McFarland, M. J. Lenardo. 1995. Two mechanisms of antigen-specific apoptosis of myelin basic protein (MBP)-specific T lymphocytes derived from multiple sclerosis patients and normal individuals. J. Immunol. 154:6191.[Abstract]
  26. Wiegers, G. J., M. S. Labeur, I. E. M. Stec, W. E. F. Klinkert, F. Holsboer, J. M. H. M. Reul. 1995. Glucocorticoids accelerate anti-T cell receptor-induced T cell growth. J. Immunol. 155:1893.[Abstract]
  27. Piccolella, E., D. Vismara, G. Lombardi, D. Guerritore, M. Piantelli, F. Ranelletti. 1985. Effect of glucocorticoids on the development of suppressive activity in human lymphocyte response to a polysaccharide purified from Candida albicans. J. Immunol. 134:1166.[Abstract]
  28. Elliott, L. H., A. K. Levay, B. Sparks, M. Miller, T. L. Roszman. 1996. Dexamethasone and prostaglandin E2 modulate T-cell receptor signaling through a cAMP-independent mechanism. Cell. Immunol. 169:117.[Medline]
  29. Elliott, L. H., A. K. Levay. 1997. Costimulation with dexamethasone and prostaglandin E2: a novel paradigm for the induction of T-cell anergy. Cell. Immunol. 180:124.[Medline]
  30. Nijhuis, E., B. Hinloopen, J. Odding, L. Nagelkerken. 1994. Abrogation of the suppressive effects of dexamethasone by PKC activation or CD28 triggering. Cell. Immunol. 156:438.[Medline]
  31. Cui, H., D. H. Sherr, M. El-Khatib, K. Matsui, D. J. Panka, A. Marshak-Rothstein, S.-T. Ju. 1996. Regulation of T-cell death genes: selective inhibition of FasL- but not Fas-mediated function. Cell. Immunol. 167:276.[Medline]
  32. Yang, Y., M. Mercep, C. F. Ware, J. D. Ashwell. 1995. Fas and activation-induced Fas ligand mediate apoptosis of T cell hybridomas: inhibition of Fas ligand expression by retinoic acid and glucocorticoids. J. Exp. Med. 181:1673.[Abstract/Free Full Text]
  33. D’Adamio, F., O. Zollo, R. Moraca, E. Ayroldi, S. Bruscoli, A. Bartoli, L. Cannarile, G. Migliorati, C. Riccardi. 1997. A new dexamethasone-induced gene of the leucine zipper family protects T lymphocytes from TCR/CD3-activated cell death. Immunity 7:803.[Medline]
  34. Wucherpfennig, K. W., J. Zhang, C. Witek, M. Matsui, Y. Modabber, K. Ota, D. A. Hafler. 1994. Clonal expansion and persistence of human T cells specific for an immunodominant myelin basic protein peptide. J. Immunol. 152:5581.[Abstract]
  35. Van Parijs, L., A. Biuckians, A. Abbas. 1998. Functional roles of Fas and Bcl-2-regulated apoptosis of T lymphocytes. J. Immunol. 160:2065.[Abstract/Free Full Text]
  36. Aggarwal, S., S. Gupta. 1998. Increased apoptosis of T cell subsets in aging humans: altered expression of Fas (CD95), Fas ligand, Bcl-2, and Bax. J. Immunol. 160:1627.[Abstract/Free Full Text]
  37. Nieto, M., A. Lopez-Rivas. 1989. IL-2 protects T lymphocytes from glucocorticoid-induced DNA fragmentation and cell death. J. Immunol. 143:4166.[Abstract]
  38. Kam, J., S. J. Szefler, W. Surs, E. R. Sher, D. Y. M. Leung. 1993. Combination IL-2 and IL-4 reduces glucocorticoid receptor-binding affinity and T cell response to glucocorticoids. J. Immunol. 151:3460.[Abstract]
  39. Mor, F., I. Cohen. 1996. IL-2 rescues antigen-specific T cells from radiation or dexamethasone-induced apoptosis. J. Immunol. 156:515.[Abstract]
  40. Yoshimoto, T., W. E. Paul. 1994. CD4pos, NK1.1pos T cells promptly produce interleukin 4 in response to in vivo challenge with anti-CD3. J. Exp. Med. 179:1285.[Abstract/Free Full Text]
  41. Vicari, A. P., A. Zlotnik. 1996. Mouse NK1.1+ T cells: a new family of T cells. Immunol. Today 17:71.[Medline]
  42. Sternberg, E., G. Chrousos, R. Wilder, P. Gold. 1992. The stress response and the regulation of inflammatory disease. Ann. Intern. Med. 117:854.
  43. Mittelstadt, P. R., J. D. Ashwell. 1998. Cyclosporin A-sensitive transcription factor Egr-3 regulates Fas ligand expression. Mol. Cell. Biol. 18:3744.[Abstract/Free Full Text]
  44. Mittelstadt, P. R., J. D. Ashwell. 1999. Role of Egr-2 in up-regulation of Fas ligand in normal T cells and aberrant double-negative lpr and gld T cells. J. Biol. Chem. 274:3222.[Abstract/Free Full Text]
  45. Masuda, K., Y. Makino, J. Cui, T. Ito, T. Tokuhisa, Y. Takahama, H. Koseki, K. I. Tsuchida, T. Koike, H. Moriya, et al 1997. Phenotypes and invariant {alpha}ß TCR expression of peripheral V{alpha}14+ NK T cells. J. Immunol. 158:2076.[Abstract]
  46. Adachi, Y., H. Koseki, M. Zijlstra, M. Taniguchi. 1995. Positive selection of invariant V{alpha}14+ T cells by non-major histocompatibility complex-encoded class I-like molecules expressed on bone marrow-derived cells. Proc. Natl. Acad. Sci. USA 92:1200.[Abstract/Free Full Text]
  47. Tamada, K., M. Harada, K. Abe, T. Li, K. Nomoto. 1998. IL-4- producing NK1.1+ T cells are resistant to glucocorticoid-induced apoptosis: implications for the Th1/Th2 balance. J. Immunol. 161:1239.[Abstract/Free Full Text]



This article has been cited by other articles:


Home page
 Int ImmunolHome page
M. D. Smithgall, M. R. Comeau, B.-R. Park Yoon, D. Kaufman, R. Armitage, and D. E. Smith
IL-33 amplifies both Th1- and Th2-type responses through its activity on human basophils, allergen-reactive Th2 cells, iNKT and NK Cells
Int. Immunol., August 1, 2008; 20(8): 1019 - 1030.
[Abstract] [Full Text] [PDF]

Home page
 BloodHome page
D. H. Chang, N. Liu, V. Klimek, H. Hassoun, A. Mazumder, S. D. Nimer, S. Jagannath, and M. V. Dhodapkar
Enhancement of ligand-dependent activation of human natural killer T cells by lenalidomide: therapeutic implications
Blood, July 15, 2006; 108(2): 618 - 621.
[Abstract] [Full Text] [PDF]

Home page
 J. Immunol.Home page
S. C. Kent, Y. Chen, S. M. Clemmings, V. Viglietta, N. S. Kenyon, C. Ricordi, B. Hering, and D. A. Hafler
Loss of IL-4 Secretion from Human Type 1a Diabetic Pancreatic Draining Lymph Node NKT Cells
J. Immunol., October 1, 2005; 175(7): 4458 - 4464.
[Abstract] [Full Text] [PDF]

Home page
 J. Exp. Med.Home page
C. Baecher-Allan and D. A. Hafler
Suppressor T Cells in Human Diseases
J. Exp. Med., August 2, 2004; 200(3): 273 - 276.
[Abstract] [Full Text] [PDF]

Home page
 Cancer Res.Home page
H. J. J. van der Vliet, J. W. Molling, N. Nishi, A. J. Masterson, W. Kolgen, S. A. Porcelli, A. J. M. van den Eertwegh, B. M. E. von Blomberg, H. M. Pinedo, G. Giaccone, et al.
Polarization of V{alpha}24+ V{beta}11+ Natural Killer T Cells of Healthy Volunteers and Cancer Patients Using {alpha}-Galactosylceramide-loaded and Environmentally Instructed Dendritic Cells
Cancer Res., July 15, 2003; 63(14): 4101 - 4106.
[Abstract] [Full Text] [PDF]

This Article