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-Chain in Regulating IL-2-Dependent, Activation-Induced CD8+ T Cell Death1
The Carlos and Marguerite Mason Transplantation Research Center, Renal Division, Department of Medicine, Veterans Affairs Medical Center and Emory University, Atlanta, GA 30033
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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, or IL-2Rß inhibited AICD. In contrast,
blocking the common cytokine receptor 
-chain (c) prevented Bcl-2
induction and augmented AICD. IL-2 up-regulated Fas ligand (FasL)
and down-regulated c expression on activated 2C cells in vitro and
in vivo. Adult IL-2 gene-knockout mice displayed exaggerated c
expression on their CD8+, but not on their
CD4+, T cells. IL-4, IL-7, and IL-15, which do not promote
AICD, did not influence FasL or c expression. These data provide
evidence that IL-2 prepares CD8+ T lymphocytes for AICD by
at least two mechanisms: 1) by up-regulating a pro-apoptotic molecule,
FasL, and 2) by down-regulating a survival molecule,
c. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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, IL-2Rß, and the
common cytokine receptor -chain
(c)3
(1). Paradoxically, IL-2 also programs T lymphocytes for
activation-induced cell death (AICD) (2) after
repeated antigenic stimulation (2, 3, 4, 5, 6, 7). The AICD of mature
CD4+ T lymphocytes is mediated by Fas-Fas ligand
(FasL) interactions (8, 9, 10, 11). IL-2 prepares these cells for
AICD by up-regulating FasL expression and down-regulating the
transcription of FLIP (IL-1ß-converting enzyme-like protease-like
inhibitory protein), a protein that inhibits Fas-mediated apoptosis
(12). The AICD of CD8+ T lymphocytes
is mediated mainly by the TNFR cell death pathway
(13, 14, 15), but Fas plays a critical role in the AICD of
autoreactive CD8+ T cells (16). The
cellular mechanisms by which IL-2 prepares CD8+ T
lymphocytes for AICD are not known. These mechanisms may differ from
those in the CD4+ population, because efficient
elimination of effector CD8+ T cells is necessary
to avoid nonspecific injury of tissues in which the inciting Ag, for
example a viral protein, persists.
c is crucial for CD8+ T cell survival. It is
expressed on naive and activated CD4+ and
CD8+ T cells; is a shared subunit of the IL-2,
IL-4, IL-7, IL-9, and IL-15 receptors; and is central to
cytokine-mediated T cell proliferation (17, 18). c
mutations in humans result in severe combined immunodeficiency
characterized by a profound decrease in circulating T lymphocytes
(19). c gene-knockout mice exhibit severely defective
lymphoid development (20, 21, 22). At birth, NK cells are
absent, and mature B and T lymphocytes are markedly diminished,
indicating that c is indispensable for the development of all murine
lymphocyte classes. Although activated
CD4+ T cells accumulate over time in
c-deficient mice, the CD8+
population remains extremely small (20, 23). These
observations suggest that mature CD4+ T cells
respond to c-independent mitogens, whereas mature
CD8+ T cells are critically dependent on c for
proliferation and survival. The finding that female c-deficient mice
made transgenic (tg) for a TCR specific for the HY male Ag lack mature
CD8+ TCR-tg T lymphocytes in their peripheral
lymphoid organs despite efficient positive selection of these cells in
the thymus further indicates that c provides essential survival
signals to CD8+ T cells (24).
The presence of multiple surface proteins that influence
CD8+ T lymphocyte survival raises the possibility
that IL-2 could prepare these cells for AICD by up-regulating
death-promoting molecules such as FasL and down-regulating
survival-promoting molecules such as c. To test this
hypothesis, we established an Ag-specific, IL-2-dependent AICD model
using CD8+ TCR-tg T cells (2C), which recognize
the Ld murine MHC class I Ag (25).
In this model, 2C cells primed in vivo with
Ld-expressing splenocytes underwent apoptosis
upon cross-linking of their tg TCR in vitro with a clonotypic Ab (1B2).
AICD occurred only if IL-2 was present in the medium. Using this model,
we provide evidence that IL-2 sensitizes CD8+ T
cells to AICD by at least two feedback mechanisms: 1) by up-regulating
FasL expression, and 2) by down-regulating c expression on activated
CD8+ T lymphocytes.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Wild-type and IL-2 gene knockout (IL-2-/-) (26) C57BL6 mice were purchased from The Jackson Laboratory (Bar Harbor, ME). 2C TCR-tg mice (C57BL6 background) (27) were provided by Dr. Dennis Loh (Washington University, St. Louis, MO) and were bred at the Veterans Administration Medical Center/Emory University animal facility. Screening for the 2C transgene was performed by PCR.
Cytokines and Abs
Endotoxin-free recombinant mouse IL-2, mouse IL-4 (3.4 x
107 U/mg), mouse IL-7 (5 x
106 U/mg), and simian IL-15 (2.2 x
108 U/mg) were purchased from Genzyme (Cambridge,
MA). Monoclonal rat anti-mouse IL-2, rat anti-mouse IL-4, and
hamster anti-mouse TNF- were also purchased from Genzyme.
Monoclonal rat anti-mouse c (3E12 and 4G3) (28),
rat anti-mouse IL-2R (3C7), rat anti-mouse IL-2Rß
(TM-ß1), isotype control rat IgG2a and rat IgG2b, and hamster
anti-mouse FasL (MFL3) were purchased from PharMingen (San Diego,
CA). The mouse hybridoma cell line producing the clonotypic Ab 1B2 that
is specific for the 2C tg TCR was provided by Dr. Dennis Loh
(Washington University) (27). Hybridoma supernatant or
isotype control mIgG1 (PharMingen) were used to coat culture wells.
Cell preparation
To study alloantigen-specific responses of CD8+ TCR-tg T lymphocytes, 2C C57BL/6 (H-2b) mice were either left naive or were primed in the footpads and i.p. with 1 x 107 BALB/c (H-2d) splenocytes/injection. Five days later, the mice were sacrificed, and the lymph node and spleen cells were isolated, pooled, and enriched for T lymphocytes by nonadherence to nylon wool columns (Polysciences, Warrington, PA). CD4+ T cells, B cells, monocytes, granulocytes, and NK cells were then eliminated by incubation with monoclonal rat anti-mouse CD4 (GK1.5), mouse anti-mouse CD59 (clone 2B4; PharMingen), and rat anti-mouse HSA (J11D; PharMingen), followed by addition of guinea pig complement (Accurate Chemical & Scientific, Westbury, NY). The remaining cell population was >95% CD8+, of which >98% expressed the 2C tg TCR (1B2+) as determined by flow cytometry. The enriched CD8+1B2+ cells, referred to as 2C cells in this manuscript, were then used in the AICD assay. The CD8+ T cell enrichment protocol was also applied to wild-type (nontransgenic) and IL-2-/- C57BL/6 mice when indicated.
AICD assay
CD8+1B2+ T lymphocytes (1 x 106) isolated from primed 2C mice as described in the previous section were cultured in 24-well plates precoated with hamster anti-mouse CD28 (37.51; PharMingen) and 1B2 hybridoma supernatant. Control wells were precoated with isotype control mouse IgG1 (PharMingen) instead of 1B2. Complete RPMI 1640 medium (10% FCS, 2 mM glutamine, 100 U/ml penicillin, and 100 µg/ml streptomycin) was used. Cytokines, cytokine-neutralizing Abs, or anti-cytokine receptor Abs were added at the beginning of the culture as indicated. Twenty-four hours later, cells were washed, counted, and analyzed for apoptosis or for cell surface markers by flow cytometry as described below. To study AICD in vivo, 2C mice were primed in the footpads with 1 x 107 BALB/c (H-2d) splenocytes on days 0 and 5. Twenty-four hours later, popliteal and inguinal lymph node cells were pooled, enriched for CD8+1B2+ cells as described in the previous section, and analyzed for apoptosis or for cell surface markers by flow cytometry.
Flow cytometry
To detect apoptosis, cells were fixed in 2% paraformaldehyde,
permeabilized with 0.1% Triton X-100 in 0.1% sodium citrate, and
labeled with fluorescein-tagged dUTP by the TUNEL method according to
the manufacturer’s instructions (In Situ Cell Death Detection Kit,
Boehringer Mannheim, Mannheim, Germany). Cells were then analyzed by
single-color flow cytometry on a Becton Dickinson FACScan (Mountain
View, CA). The total lymphocyte population was gated, and apoptosis was
measured by calculating the percentage of TUNEL+
cells. To determine the absolute number of apoptotic cells, the
percentage of TUNEL+ cells was multiplied by the
total number of cells present in each well at the end of the
experiment. To measure cell surface markers, cells were stained with
PE- or FITC-conjugated rat anti-CD4 (H129.19; PharMingen), PE- or
FITC-conjugated rat anti-mouse CD8 (53-6.7; PharMingen),
FITC-conjugated rat anti-mouse IL-2Rß (TM-ß1; PharMingen),
PE-conjugated rat anti-mouse c (4G3; PharMingen), or
PE-conjugated hamster anti-mouse FasL (MFL3; PharMingen). To detect
intracellular Bcl-2 expression, cells were fixed in 1%
paraformaldehyde and permeabilized with 0.1% Triton X-100 before
staining with FITC-conjugated hamster anti-mouse Bcl-2 (3F11;
PharMingen). The appropriate conjugated, isotype control Abs were used
as negative controls. Stained cells were analyzed by single- or
dual-color flow cytometry on a Becton Dickinson FACScan.
Proliferation assay
CD8+1B2+ T lymphocytes isolated from primed 2C mice were cultured in triplicate at 37°C in a 5% CO2 incubator in complete RPMI medium (10% FCS, 2 mM glutamine, 100 U/ml penicillin, and 100 µg/ml streptomycin) in 96-well plates (2.5 x 105 cells/well) precoated with 1B2 hybridoma supernatant and hamster anti-mouse CD28 (37.51; PharMingen). mAbs to IL-2 and the IL-2R subunits were added at the beginning of the culture as indicated. Twenty-four hours later, the wells were pulsed with 0.5 µCi of [3H]TdR and harvested after 6 h onto fiberglass filter papers (PhD cell harvester; Cambridge Technology, Cambridge, MA). [3H]TdR uptake was measured in a scintillation counter (Beckman, Palo Alto, CA).
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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We established an AICD model driven by repeated stimulation of
TCR-tg mature CD8+ T cells (2C). 2C cells react
specifically to the Ld MHC class I Ag, and their
TCR is recognized by a clonotypic Ab (1B2) (25).
Splenocytes of either naive 2C mice or 2C mice primed with
Ld-expressing splenocytes were enriched for
CD8+1B2+ T cells and
cultured in complete medium in the presence of either plate-bound 1B2
Ab or isotype control mIgG. Apoptosis was measured by flow cytometry
24 h later. As shown in Fig. 1
A, a small proportion of
naive 2C cells underwent apoptosis when their TCR were cross-linked in
vitro with 1B2. This degree of apoptosis was most likely due to passive
cell death, because a similar proportion of naive 2C cells underwent
apoptosis when challenged with mIgG instead of 1B2 (Fig. 1A). In contrast, in vitro stimulation with 1B2 increased
the percentage of apoptotic cells when 2C cells were preactivated in
vivo by challenging 2C mice with Ld-expressing
splenocytes (Fig. 1A). Unlike 1B2, mIgG did not increase the
apoptosis of preactivated 2C cells, indicating that AICD in this model
is dependent on repeated stimulation of T lymphocytes via their
TCR.
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B). Excess
IL-2-neutralizing Ab, but not control rat IgG or IL-4-neutralizing Ab,
completely inhibited 1B2-induced apoptosis of preactivated 2C cells
(Fig. 1C). The data indicate that the AICD of
CD8+ T cells is dependent on IL-2.
To determine whether IL-4, IL-7, and IL-15 reduce the proportion of
apoptotic cells by preventing AICD or by increasing T cell
proliferation, we calculated the absolute number of apoptotic 2C cells
following the induction of AICD (Fig. 1D). We found that
IL-2 increases the number of apoptotic cells significantly, whereas
IL-4 and IL-7 reduced the number of apoptotic cells by a small, but
significant, amount (19–24% reduction). IL-15 did not have a
significant effect on AICD. These data suggest that IL-4 and IL-7 have
a modest antiapoptotic effect on activated CD8+ T
cells.
Counting apoptotic cells is subject to error, because dead cells can
degenerate in culture. We therefore quantitated the absolute number of
viable cells 24 h after in vitro stimulation of
CD8+ T lymphocytes obtained from either naive or
primed 2C mice (Fig. 1E). The number of viable 2C
lymphocytes obtained from naive 2C mice increased significantly
following their stimulation with 1B2 alone or with 1B2 plus IL-2, IL-4,
IL-7, or IL-15. Addition of anti-IL-2 inhibited 1B2-induced
expansion of these cells. In contrast, the number of viable 2C
lymphocytes obtained from primed 2C mice did not increase significantly
when stimulated with 1B2 alone or with 1B2 plus IL-2. The addition of
IL-4, IL-7, or IL-15 restored 1B2-induced expansion of these cells.
These data are also portrayed as relative cell loss (percentage), which
reflects the difference in cell expansion between 2C lymphocytes
obtained from primed mice and those obtained from naive mice (Fig. 1F). These findings confirm the unique pro-apoptotic effect
of IL-2 on activated CD8+ T cells.
Blocking FasL or TNF- inhibits IL-2-dependent AICD of
CD8+ T cells
To test whether the IL-2-dependent AICD of
CD8+ T lymphocytes is mediated by the Fas and/or
the TNFR death pathways, we studied the effect of FasL- and
TNF--blocking Abs on AICD in our model. In vivo activated 2C cells
were stimulated in vitro with plate-bound 1B2 Ab, and the percentage of
apoptotic cells was determined 24 and 48 h later. We found that
the addition of excess TNF--neutralizing Ab reduced AICD at 24
h by about 20%, whereas blocking FasL-Fas interactions with
anti-FasL Ab inhibited AICD by about 60% (Fig. 2). In contrast, TNF- neutralization
resulted in more inhibition of AICD at 48 h than that achieved by
blocking FasL (
55 vs 10%). Combined blockade of FasL and TNF-
did not result in additive or synergistic inhibition of AICD at 24 or
48 h, suggesting that the roles of Fas and TNFR in
CD8+ T cell apoptosis are separated
temporally.
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IL-2 up-regulates TNFR expression on CD4+
and CD8+ T cells (29, 30),
suggesting that IL-2 sensitizes activated T cells to apoptosis via the
TNFR death pathway. IL-2 up-regulates the surface expression of FasL on
CD4+ T cells (12), but its effect on
FasL expression on CD8+ T cells is not known. To
address this issue, 2C mice were primed with
Ld-bearing splenocytes. Five days later, their
spleen cells were enriched for
CD8+1B2+ T cells and
restimulated in vitro with plate-bound 1B2 Ab or isotype control mIgG.
FasL expression was measured 24 h later by flow cytometry. As
shown in Fig. 3, activation of 2C cells
with plate-bound 1B2 up-regulated FasL expression. Adding rIL-2
enhanced FasL expression further, whereas IL-2-neutralizing Ab
inhibited FasL up-regulation (Fig. 3A). In contrast to IL-2,
cytokines that do not promote AICD (IL-4, IL-7, and IL-15) did not
alter FasL expression on activated 2C cells (Fig. 3B).
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c-blocking Abs enhance IL-2-dependent AICD of CD8+ T
cells and inhibit Bcl-2 induction
To test whether c regulates IL-2-dependent AICD of
CD8+ T cells, we examined the effect of
c-blocking Abs on lymphocyte apoptosis in our model. Abs that target
c (3E12 and 4G3), but do not interfere with the binding of IL-2 to
the and ß subunits of the IL-2R (28), enhanced AICD
in the 2C cell population (Fig. 4A). Enhanced AICD was
confirmed by determining the absolute number of apoptotic cells in this
experiment. A 34% increase in the absolute number of apoptotic cells
was observed following the addition of c-blocking Abs to
1B2-stimulated 2C cells (mean of three experiments). Dose-dependent
augmentation of AICD was observed when the concentrations of 3E12 and
4G3 were varied between 0.1–100 µg/ml (data not shown). If rIL-2 was
added to the medium along with 3E12 and 4G3, the proportion of
apoptotic cells increased further, confirming that IL-2 interacts with
its receptor and promotes AICD in the presence of c-blocking Abs
(Fig. 4A). In contrast, Abs that block the and ß
subunits of the IL-2R inhibited AICD completely (Fig. 4A).
Enhanced apoptosis induced by c-blocking Abs was not due to
increased passive cell death, because these Abs did not alter the
apoptosis of naive 2C cells cultured in the presence of plate-bound
mIgG (25.3, 25.8, 26.0, and 25.6% apoptosis in the presence of rat IgG
isotype control, anti-IL-2R, anti-IL-2Rß, and
anti-IL-2R, respectively) or plate-bound 1B2 (26.2, 27.2, 26.1,
and 29.8% apoptosis in the presence of rat IgG isotype control,
anti-IL-2R, anti-IL-2Rß, and anti-IL-2R,
respectively). Although anti-IL-2R, anti-IL-2Rß, and
anti-c Abs inhibited 1B2-induced proliferation of preactivated
2C cells (Fig. 4B), only anti-c Abs enhanced their
apoptosis (Fig. 4A). These observations indicate that c
is essential for both the proliferation and the survival of activated
CD8+ T cells. To further examine the
anti-apoptotic role of c, we tested the effect of c-blocking
Abs on Bcl-2 expression. We found that anti-c Abs, but not those
that block the - and ß-chains of the IL-2R, inhibit
activation-induced Bcl-2 expression in 2C cells (Fig. 4C).
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c expression on activated CD8+ T
cells in vitro and in vivo
Because c is critical for the survival and proliferation of
activated CD8+ T cells, we asked whether IL-2
induces sensitivity to AICD by down-regulating c expression on these
cells. 2C mice were primed with Ld-bearing
splenocytes. Five days later, their spleen and lymph node cells were
enriched for CD8+1B2+ T
cells and restimulated in vitro with plate-bound 1B2 Ab or isotype
control mIgG. As shown in Fig. 5A, cross-linking the 2C TCR
with 1B2 up-regulated c expression. However, c expression was
significantly higher in the presence of IL-2-neutralizing Ab and was
significantly lower when rIL-2 was added to the culture medium. IL-4,
IL-7, and IL-15, which do not promote AICD (Fig. 1B),
did not alter c expression. In contrast to its effects on
c, IL-2 enhanced the expression of the IL-2R ß-chain on activated
2C cells, whereas IL-4, IL-7, and IL-15 resulted in its down-regulation
(Fig. 5B).
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c and IL-2Rß
expression in vivo. 2C mice were primed with
Ld-bearing splenocytes on days 0 and 5, and their
CD8+ T cells were analyzed for c and IL-2Rß
expression by flow cytometry 24 h later. As shown in Fig. 6, IL-2-neutralizing Ab given to mice at
the time of secondary stimulation with allogeneic splenocytes resulted
in superinduction of c and inhibition of IL-2Rß expression.
Moreover, we found that c expression on CD8+ T
cells of adult IL-2 gene-knockout (IL-2-/-)
mice was higher than that observed in wild-type
(IL-2+/+) mice (Fig. 7). Comparable levels of c, on the
other hand, were detected on IL-2-/- and
IL-2+/+ CD4+ T cells (Fig. 7). IL-2Rß expression on both CD8+ and
CD4+ T cells was lower in the
IL-2-/- group (Fig. 7). These findings indicate
that down-regulation of c expression by endogenous IL-2 may be
restricted to the CD8+ T cell subpopulation.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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c, which is critical
for the survival and proliferation of CD8+ T
lymphocytes.
We found that both Fas and TNFR death pathways contribute to the
apoptosis of activated CD8+ T cells. The roles of
Fas and TNFR, however, were separated temporally. Fas contributed to
early apoptosis observed within 24 h of TCR engagement, whereas
TNFR prevailed as a mediator of T cell apoptosis after the first
24 h. Although Zheng et al. have shown that TNF--TNFR
interactions mediate the AICD of most CD8+ T
cells, they also observed the same temporal separation between the
actions of Fas and TNFR (13). Moreover, Kurts et al.
provided evidence that the peripheral deletion of autoreactive
CD8+ T cells induced by self-Ags involves
signaling through Fas (16). These data suggest that the
death pathway used by an activated T cell depends on the nature and
quantity of the antigenic stimulus (11, 16). We also
observed that FasL-blocking Abs, TNF--neutralizing Abs, or both fail
to suppress AICD completely in our model, suggesting that additional
members of the TNFR family, for example TRAMP and TRAIL receptors
(31, 32), contribute to the AICD of
CD8+ T cells. Importantly, we found that IL-2
up-regulates FasL expression on activated CD8+ T
cells within 24 h of TCR cross-linking, whereas IL-4, IL-7, and
IL-15, which do not promote AICD, fail to do so. Others have shown that
IL-2 up-regulates TNFR expression on CD8+ T cells
(30, 33). Taken together, these findings suggest that IL-2
sensitizes CD8+ T cells to AICD by up-regulating
the expression of cell surface molecules involved in triggering
apoptosis. Increased FasL expression alone, however, is not necessarily
sufficient for enhancing T cell apoptosis. Modulating the expression of
intracellular molecules that either inhibit or promote Fas-mediated
apoptosis is also required. These molecules include FLIP and the
product of the c-myc protooncogene (12, 34, 35). In addition to enhancing AICD, IL-2-induced up-regulation
of FasL on CD8+ T cells promotes their CTL
activity, because FasL-Fas is an important pathway by which CTL kill
target cells (36, 37).
Alternatively, IL-2 could prepare CD8+ T cells
for AICD by down-regulating the expression of surface receptors that
promote lymphocyte survival and proliferation. A greatly diminished
amount of CD8+ T cells in neonatal and adult c
gene-knockout mice suggests that c is required for the development,
survival, and proliferation of these cells (20, 23, 24).
We found in this study that c-blocking Abs increase the proportion
of apoptotic cells following the induction of AICD in a
CD8+ TCR-tg cell population. In contrast, Abs
that block the and ß subunits of the IL-2R inhibited AICD
completely. Anti-c Abs increased the proportion of apoptotic cells
in our model by blocking both mitogenic and survival signals because
these Abs inhibited the proliferation of activated
CD8+ T cells and prevented the induction of
Bcl-2, an anti-apoptotic molecule. The latter finding is consistent
with the markedly reduced intracellular levels of Bcl-2 in c
gene-knockout T cells (23, 38, 39). Studies demonstrating
increased apoptosis of activated T cells in Bcl-2 gene-knockout mice
(40), and those demonstrating a correlation between low
intracellular Bcl-2 concentrations and enhanced susceptibility of CTLs
to Ag-mediated apoptosis (41) provide further evidence
that Bcl-2 is an important regulator of AICD.
Because c is critical for the survival and proliferation of
activated CD8+ T cells, we asked in this study
whether c expression is regulated by IL-2. We found that exogenous
IL-2 down-regulates c expression in vitro and that endogenous IL-2
limits c expression on activated CD8+ T cells
in vitro and in vivo. Cytokines that do not promote AICD (IL-4, IL-7,
and IL-15) did not influence c expression. These data suggest that
IL-2 prepares CD8+ T cells for AICD by limiting
the surface expression of c. IL-2 could regulate both
transcriptional and post-transcriptional events responsible for c
production. Ohbo et al. (42) showed that IL-2 decreases
the expression of a reporter gene positioned downstream of the c
promoter region. Alternatively, Noguchi et al. (43) found
that c is cleaved by calpain following activation of murine
thymocytes, suggesting that IL-2 may use proteolytic pathways to reduce
c expression on T cells.
IL-2-/- mice develop severe lymphoproliferation
characterized by the accumulation of activated
CD4+ and CD8+ T cells,
which are resistant to AICD (5, 7, 44, 45). We observed in
this study that CD8+ T cells from adult
IL-2-/- mice display exaggerated c
expression. c expression on IL-2-/-
CD4+ cells, however, was not greater than that on
IL-2+/+ lymphocytes. This observation raises the
possibility that IL-2-dependent down-regulation of c is a feedback
mechanism by which activated CD8+ T cells, but
not activated CD4+ T cells, are sensitized to
apoptosis. In fact, c-dependent signals may play a crucial role in
the peripheral deletion of mature CD4+ T cells,
because in c gene-knockout mice these cells accumulate over time,
display activation markers, have reduced FasL expression, and are
resistant to superantigen-induced elimination (20, 23, 46). In contrast, mature CD8+ T cells are
severely diminished in neonatal c gene-knockout mice and do not
accumulate over time (20, 24).
In summary, IL-2 ensures that the expansion of activated
CD8+ T cells is limited by at least two
mechanisms: 1) by up-regulating pro-apoptotic molecules, such as FasL,
and 2) by down-regulating c, which provides essential mitogenic and
survival signals to CD8+ T lymphocytes. These
homeostatic mechanisms may prevent nonspecific tissue injury following
persistent viral infections and may play a role in the induction of
immunologic tolerance to transplanted organs.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Fadi G. Lakkis, Veterans Administration Medical Center and Emory University, Research 151N, 1670 Clairmont Road, Atlanta, GA 30033. E-mail address:
3 Abbreviations used in this paper: c, common cytokine receptor -chain; AICD, activation-induced cell death; FasL, Fas ligand; IL-2+/+, wild-type mice; IL-2-/-, IL-2 gene-knockout mice; tg, transgenic; FLIP, IL-1ß-converting enzyme-like protease-like inhibitory protein.
Received for publication May 14, 1999. Accepted for publication July 7, 1999.
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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J. Kan-Mitchell, B. Bisikirska, F. Wong-Staal, K. L. Schaubert, M. Bajcz, and M. Bereta The HIV-1 HLA-A2-SLYNTVATL Is a Help-Independent CTL Epitope J. Immunol., May 1, 2004; 172(9): 5249 - 5261. [Abstract] [Full Text] [PDF]
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B. H. Nelson IL-2, Regulatory T Cells, and Tolerance J. Immunol., April 1, 2004; 172(7): 3983 - 3988. [Abstract] [Full Text] [PDF]
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 F. G. Lakkis Transplantation tolerance: a journey from ignorance to memory Nephrol. Dial. Transplant., October 1, 2003; 18(10): 1979 - 1982. [Full Text] [PDF]
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 S. Oh, J. A. Berzofsky, D. S. Burke, T. A. Waldmann, and L. P. Perera Coadministration of HIV vaccine vectors with vaccinia viruses expressing IL-15 but not IL-2 induces long-lasting cellular immunity PNAS, March 18, 2003; 100(6): 3392 - 3397. [Abstract] [Full Text] [PDF]
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 Y. M. Mueller, V. Makar, P. M. Bojczuk, J. Witek, and P. D. Katsikis IL-15 enhances the function and inhibits CD95/Fas-induced apoptosis of human CD4+ and CD8+ effector-memory T cells Int. Immunol., January 1, 2003; 15(1): 49 - 58. [Abstract] [Full Text] [PDF]
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M. Takahashi, E. Osono, Y. Nakagawa, J. Wang, J. A. Berzofsky, D. H. Margulies, and H. Takahashi Rapid Induction of Apoptosis in CD8+ HIV-1 Envelope-Specific Murine CTLs by Short Exposure to Antigenic Peptide J. Immunol., December 1, 2002; 169(11): 6588 - 6593. [Abstract] [Full Text] [PDF]
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M. P. Rubinstein, A. N. Kadima, M. L. Salem, C. L. Nguyen, W. E. Gillanders, and D. J. Cole Systemic Administration of IL-15 Augments the Antigen-Specific Primary CD8+ T Cell Response Following Vaccination with Peptide-Pulsed Dendritic Cells J. Immunol., November 1, 2002; 169(9): 4928 - 4935. [Abstract] [Full Text] [PDF]
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L. E. Cheng and P. D. Greenberg Selective Delivery of Augmented IL-2 Receptor Signals to Responding CD8+ T Cells Increases the Size of the Acute Antiviral Response and of the Resulting Memory T Cell Pool J. Immunol., November 1, 2002; 169(9): 4990 - 4997. [Abstract] [Full Text] [PDF]
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B. Pantenburg, F. Heinzel, L. Das, P. S. Heeger, and A. Valujskikh T Cells Primed by Leishmania major Infection Cross-React with Alloantigens and Alter the Course of Allograft Rejection J. Immunol., October 1, 2002; 169(7): 3686 - 3693. [Abstract] [Full Text] [PDF]
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P. Shrikant and M. F. Mescher Opposing Effects of IL-2 in Tumor Immunotherapy: Promoting CD8 T Cell Growth and Inducing Apoptosis J. Immunol., August 15, 2002; 169(4): 1753 - 1759. [Abstract] [Full Text] [PDF]
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, no
cytokine added. C, Endogenous IL-2 is required for AICD.
2C cells were isolated from primed 2C mice, stimulated in vitro with
either mIgG (control) or 1B2, and analyzed for apoptosis as described
in A. Isotype control (rat IgG) or cytokine-neutralizing
Abs (10 µg/ml each) were added at the beginning of the culture. The
mean ± SD of three experiments are shown. D,
Effect of T cell mitogenic cytokines on AICD. The AICD of 2C cells was
studied as described in B and C. The
absolute number of apoptotic 2C cells was calculated by multiplying the
percentage of TUNEL+ cells by the total number of cells
present in each well at the end of the experiment. The mean ± SD
of three experiments are shown. E, Effect of T cell
mitogenic cytokines on the number of viable naive (open bars) or primed
(solid bars) 2C cells 24 h following in vitro stimulation with
mIgG or 1B2. Cell viability was determined by trypan blue exclusion.
The mean ± SD of three experiments are shown. F,
Effect of T cell mitogenic cytokines on the relative loss of primed 2C
cells following in vitro stimulation with mIgG or 1B2. Relative cell
loss (percentage) in each experimental condition was calculated based
on cell numbers shown in E according to the following
formula: (open bar - solid bar/open bar) x 100.
