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Center d’Immunologie de Marseille Luminy, Institut National de la Santé et de la Recherche Médicale/Centre National de la Recherche Scientifique/Université de la Méditérranée, Parc Scientifique de Luminy, Marseille, France
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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-producing cells. Ag and
cytokine availability therefore greatly limits the differentiation, but
not the initial proliferation, of CD4+ T cells into
IFN--producing cells. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Activation of naive CD4+ T cells and entry into the cell cycle was shown to require an exposure to Ag for a minimum of 12 h (4). The first division of activated CD4+ T cells is however not observed before 24 h following Ag stimulation. Past this lag time, progression through cell cycle is extremely rapid with a maximum generation time of 4–6 h (5, 6). Whether TCR re-engagement and/or cytokines are required to sustain cell division of activated CD4+ T cells during this intense proliferation phase is still unknown.
The differentiation of naive CD4+ T cells into
effector Th1 cells likewise requires a commitment phase during which
differentiating cells do not produce the prototype Th1 effector
cytokines, IFN- and TNF-
. Commitment to the Th1 lineage is
regulated by complex signals including ligand density, costimulatory
molecules, and cytokines (7, 8, 9, 10). Cytokines such as IL-12
play a preeminent role in driving Th1 differentiation (7).
Activation of STAT4 upon IL-12 binding to the IL-12R is essential in
inducing the commitment of naive CD4+ T cells to
the Th1 lineage; although, under some conditions, a STAT4-independent
Th1 differentiation may occur (11, 12, 13). Expression of the
transcription factor T-bet was also shown to be restricted to
differentiating Th1 cells and to specifically induce the expression of
IFN- while repressing IL-4 and IL-2
gene expression (14). Despite rapid induction of
these different transcription factors, commitment to the Th1 lineage
requires at least 48 h (15). Whether continuous TCR
engagement and/or cytokines signaling are required during this
commitment period is still a matter of debate. Indeed, in one study, a
24-h stimulation with plate-bound MHC-peptide complexes in the presence
of IL-12 was shown to be efficient at inducing Th1 differentiation
(16). In other studies however, a 72-h exposure to TCR and
IL-12 stimulation was necessary for full differentiation of effector
Th1 cells (5, 13). As both Ag and cytokines may be
limiting in the course of an immune response in vivo, it is essential
to evaluate to what extent these two parameters are regulating
CD4+ T cell responses.
In this report, we determined whether sustained TCR and cytokine signaling are required for clonal expansion and differentiation of effector Th1 cells. We found that transient exposure to Ag induces a program of proliferation that can develop for six to seven divisions, in vivo and in vitro, in the absence of subsequent contact with Ag. In contrast, Ag-driven differentiation of effector Th1 cells requires TCR re-engagement and cytokine signaling, both signals being necessary for the sustained expression of T-bet gene. As opposed to CD8+ T cells, for which initial Ag encounter induces a program of proliferation and differentiation that can proceed in the absence of Ag (17, 18), the differentiation, but not the initial burst of proliferation of CD4+ T cells appears therefore tightly regulated by Ag and cytokine availability.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The 3A9 mice are transgenic for a TCR specific for the hen egg lyzosyme (HEL)3 peptide 46–61 presented by I-Ak (19). The 3A9 mice were maintained on a CBA/J background or, for adoptive transfer, crossed with B10.BR Ly5.1+ congenic mice.
Adoptive transfer
(CBA/J x B10.BR)F1 recipient mice were injected i.v. with 2.5 x 106 naive or in vitro-activated CD4+ T cells from the 3A9-transgenic mice. Twenty-four hours before adoptive transfer, the mice were immunized by s.c. injection, in the backs of recipient mice, of 200 µg of native HEL protein (Sigma-Aldrich, St. Louis, MO) emulsified in CFA (Sigma-Aldrich) or CFA alone in a 200-µl volume, as previously described (20).
Abs and FACS staining
The Abs anti-CD4 (RM4-5), anti-IFN- (XMG1.2),
anti-IL-2 (JES6-5H4), anti-IL-4 (11B11), and anti-Ly5.1
(A20) were purchased from BD PharMingen (San Jose, CA). These Abs
were directly coupled to allophycocyanin, PE , FITC , or biotin, in
which case staining was revealed using streptavidin-allophycocyanin (BD
PharMingen). TO-PRO-3 (Molecular Probes, Eugene, OR) was used at a
final concentration of 0.5 nM. For cytokine staining, cells were
stimulated for 4 h with 50 ng/ml PMA and 500 ng/ml ionomycin
(Sigma-Aldrich), and 10 µg/ml brefeldin A (Sigma-Aldrich) was added
for the last 2 h or for the entire 4-h stimulation period when ex
vivo T cells were analyzed. Cells were harvested, stained for
appropriate surface markers, fixed with 2% paraformaldehyde, and
incubated in permeabilization buffer containing the anti-cytokine
Ab as previously described (1).
Cell preparation and in vitro activation
CD4+ T cells were purified from the lymph
nodes of 3A9-transgenic mice by negative selection using a mixture of
Ab composed of an anti-I-Ek (M5/114.15.2), an
anti-B220 (RA36B2), an anti-CD8 (H-59-101-2), an
anti-FcRII/III (2.4.G2), and an anti-CD11b (M1/70.15.11.5HL)
followed by an incubation with sheep anti-rat IgG magnetic beads
(Dynal Biotech, Oslo, Norway). On average, the population recovered was
composed of >90% CD4+ T cells, of which <3%
were CD44+. The cells were labeled with CFSE
(Molecular Probes) as previously described (21). T
cell-depleted splenic APCs were prepared by Ab-mediated complement
lysis using an anti-Thy1.2 Ab (J1J) and were gamma irradiated at 24
Gy. Naive T cells (5 x 105) were stimulated
in 24-well plates with APCs (1 x 106) and
0.6 µg/ml HEL 46–61 peptide. Forty-eight hours later, dividing
CFSE+CD4+ T cells were FACS
sorted by excluding cells that were in the most brightest peak (0
division). FACS-sorted populations were >98% pure. Sorted
CD4+ T cells (7.5 x
104) were seeded either in the upper or lower
compartment of a 12-well Transwell plate with a 0.4-µm membrane pore
size (Costar, Cambridge, MA) or in a standard 24-well-plate. When
indicated, 5 x 105 APCs were added along
with 0.6 µg/ml HEL 46–61 peptide. For anti-CD3 stimulation,
plates were coated overnight with 10 µg/ml anti-CD3 (145.2C11)
and 1 µg/ml soluble anti-CD28 Ab (37.51) was added during the
culture period. When indicated, neutralizing anti-IL-2 (2 µg/ml,
JES6-1A12; BD PharMingen), anti-IL-12 (5 µg/ml,C17-6, a generous
gift from G. Trinchieri, Schering-Plough Research Institute, Dardilly,
France), anti-IL-4 (5 µg/ml, 11B11; BD PharMingen), or
anti-IFN- (5 µg/ml, XMG1.2; BD PharMingen) Ab were added.
Recombinant murine cytokines were IL-2 (10 or 50 U/ml; PeproTech,
Princeton, NJ), IL-12 (3.5 ng/ml; PeproTech), or IFN-
(50U/ml; Genzyme, Cambridge, MA).
RNA preparation and RT-PCR
Total RNA was extracted using the High Pure RNA Isolation kit
(Roche Diagnostic, Mannheim, Germany) according to the manufacturer’s
instructions, treated with DNase I (Roche Diagnostic) and reverse
transcribed using random primers and Superscript II RT (Life
Technologies, Grand Island, NY). Real-time PCR was performed on cDNA
samples using the Taqman Sybr Green system (PE Biosystems, Warrington,
U.K.). Primers used were hypoxanthine phosphoribosyltransferase
(HPRT) sense 5'-AGCCCTCTGTGTGCTAAGG-3', HPRT antisense
5'-CTGATAAAATCTACAGTCATAGGAATGGA-3'; T-bet sense
5'-CAACAACCCCTTTGCCAAAG-3', and T-bet antisense
5'-TCCCCCAAGCAGTTGACAGT-3'. Cycling conditions were 1 cycle at
50°C for 2 min, 1 cycle at 95°C for 10 min, and 40 cycles each
corresponding to 15 s at 95°C and 1 min at 60°C. Analysis used
the sequence detection software supplied with the instrument. The
relative quantitation value is expressed as
2-
CT and percentage of maximal
2-CT, where CT is
the difference between the mean CT value of
triplicates of the sample and of the endogenous HPRT control.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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We analyzed the response of CD4+ T cells
isolated from the 3A9 TCR-transgenic mice that are specific for the HEL
peptide presented by the class II MHC molecule
I-Ak (referred to hereafter as 3A9
CD4+ T cells (19)). To follow T cell
division, we used the vital dye CFSE that is equally partitioned to
daughter cells at each division (21). To analyze the role
of TCR re-engagement or cytokine signaling in further driving the
division and differentiation of cycling CD4+ T
cells, we stimulated the 3A9 CD4+ T cells for
48 h and FACS sorted the 3A9 CD4+ T cells
that had accomplished at least one division. At this time point, on
average, 90% of the CD4+ T cells had achieved
one to three divisions (data not shown) and did not produce significant
levels of cytokines such as IL-2, IL-4, or IFN- without
restimulation (Fig. 1
). Most of these
activated 3A9 CD4+ T cells are undifferentiated
since they produce IL-2 but little IFN- or IL-4 following
restimulation with PMA and ionomycin or upon TCR re-engagement (Fig. 1, b and d). We first determined whether TCR
re-engagement and cytokine signaling were necessary to sustain cycling
of activated T cells. To mimic as closely as possible the Ag and
cytokine environment that may occur during an in vivo response, we used
a Transwell system with a membrane pore size of 0.4 µm that allows
the passage of soluble factors such as cytokines but not cells. Sorted
CD4+ T cells were seeded in the upper and lower
chambers of the Transwell while APCs and Ag were only added to the
upper chamber. The upper well thus contained CD4+
T cells that had re-engaged their TCR and, as a consequence of this TCR
stimulation, may produce cytokines available for themselves and for the
CD4+ T cells present in the lower well that did
not re-engage their TCR. As shown in Fig. 2a, CD4+
T cells present in the upper and lower chambers of the Transwell
divided equally within the 48 h of reculture. The continuation of
cell division in the lower chamber did not result from carryover of
Ag-loaded APC since the CD4+ T cells were FACS
sorted to >98% purity. Furthermore, similar results were obtained
when the upper and lower wells contained CD4+ T
cells recognizing distinct MHC-peptide complexes (data not shown).
Finally, naive 3A9 CD4+ T cells when added to the
lower chamber were not activated nor did they divide (data not shown).
These results indicate that although entry into the cell cycle requires
TCR engagement, further progression through cell division, during the
48 h of analysis, does not require TCR re-engagement.
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d). Anti-IL-2 blocking Ab was therefore added to the
Transwell system. Cycling of CD4+ T cells in the
upper chamber of the Transwell was similarly sustained whether IL-2 was
present or not during the reincubation period, indicating that cycling
of cells induced by TCR re-engagement may be IL-2 independent (Fig. 2a). Although continuous IL-2 signaling was not necessary to
sustain the proliferation of activated 3A9 CD4+ T
cells, IL-2 greatly contributed to the survival of these activated T
cells in vitro (Fig. 2a). Indeed, 73% of the cycling
CD4+ T cells died when restimulated in the
absence of IL-2 as evidenced by TO-PRO-3 uptake, a dye that
specifically stains dead cells. 3A9 CD4+ T cells
present in the lower well also progressed through further division
whether IL-2 was present or not (Fig. 2a). Indeed, in the
absence of IL-2, the 48-h activated CD4+ T cells
made an additional three divisions within 24 h (Fig. 2a). Past this 24-h time period, when deprived of IL-2, the
CD4+ T cells present in the lower well did not
further divide and most of them died (Fig. 2a). We further
tested that no other cytokines, such as IL-4, produced by activated T
cells or IL-7 or -15 produced by the APC present in the upper well, may
contribute to cycling of the CD4+T cells present
in the lower chamber. We reincubated sorted cycling
CD4+ T cells in medium alone, a condition where
none of these cytokines are produced or, as control, in combination
with syngeneic APCs with or without IL-2 or HEL peptide. Activated
CD4+ T cells further divided in all four culture
conditions (Fig. 2b). As observed in the Transwell system,
activated 3A9 CD4+ T cells made three to four
additional divisions within 24 h regardless whether cytokines or
Ag were added or not during the restimulation period. Past this 24-h
period, 3A9 CD4+ T cells maintained in medium
alone stopped cycling and most of them died, in contrast to cells
reincubated with IL-2 or Ag-loaded APCs that continued to divide (Fig. 2b).
Collectively, these results indicate that once into cycle,
CD4+ T cells proceed through several rounds of
divisions in the absence of subsequent TCR and cytokine signaling. We
further determined whether in vivo cycling of activated
CD4+ T cells also proceeded in an Ag-independent
fashion. For these experiments, CFSE-labeled 3A9
CD4+ T cells expressing the Ly5.1 allotype were
activated for 48 h in vitro, purified to remove any Ag-loaded
APCs, and then transferred into syngenic mice expressing the
Ly5.2 allele. Twenty-four hours before adoptive transfer,
the recipient mice were injected s.c. with CFA with or without HEL, or
left unimmunized. Activated T cells further divided when transferred
into unimmunized animals (Fig. 3a). Under those conditions,
they performed four additional divisions within 48 h and some of
them further divided. As observed in vitro, most activated 3A9
CD4+ T cells stopped dividing after 48-h
posttransfer into unimmunized hosts, although under those in vivo
conditions the CD4+ T cells did not die since the
number of 3A9 CD4+ T cells recovered at 48 and
72 h were comparable (Fig. 3b). Cell cycle arrest in
this case is therefore not resulting from starvation or death of the
CD4+ T cells but instead reflects the
extent of the programming ofproliferation induced by initial
Ag encounter. To ensure that the proliferation of activated 3A9
CD4+ T cells when transferred into unimmunized
hosts occurred in the absence of Ag, we cotransferred naive 3A9
CD4+ T cells along with purified activated 3A9
CD4+T cells. In this case, the 3A9
CD4+ T cells expressing the host Ly5.2 molecule
were stimulated in vitro and cotransferred with CFSE-labeled naive 3A9
CD4+ T cells expressing the Ly5.1
allele. The proliferation of naive 3A9 CD4+ T
cells that served as sensors for the presence of residual Ag was then
evaluated. As shown in Fig. 3c, no proliferation of the
sensor cells was detected over the 3-day period of analysis, indicating
that indeed, proliferation of the 3A9 CD4+ T
cells transferred into unimmunized hosts proceeded in the absence
of Ag.
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Repeated Ag exposure is necessary for the differentiation of Th1 cells
Having shown that TCR re-engagement was not required to sustain
the initial phase of proliferation of activated T cells, we wished to
determine whether the same applied for the differentiation of activated
T cells into IFN--producing Th1 cells. We adoptively transferred
48-h activated 3A9 CD4+ T cells into mice
injected 24 h previously with CFA with or without HEL, as
described above. The ability of the 3A9 CD4+ T
cells to produce IFN- following transfer was determined by
intracellular FACS staining upon 4-h in vitro stimulation with PMA and
ionomycin. Importantly, the 48-h activated 3A9
CD4+ T cells used for the transfer are mainly
undifferentiated, most of them producing no IFN- (Fig. 1). The
3A9+CD4+ T cells that were
transferred into mice previously immunized with HEL further
differentiated into IFN--producing Th1 cells (Fig. 3d).
In contrast, 3A9 CD4+ T cells transferred in the
absence of Ag did not differentiate into IFN--producing cells (Fig. 3d).
Altogether, these results indicate that, in contrast to the early burst of proliferation, the full differentiation of effector Th1 cells requires repeated contacts with Ag-loaded APCs.
Sustained TCR and cytokine signaling are necessary for optimal differentiation of effector Th1 cells
The above experiment indicated that the differentiation of
CD4+ T cells into Th1 effector cells required
repeated contact with Ag-loaded APCs. Since both TCR engagement and
IL-12 signaling may result from this interaction, it was essential to
determine which of those signals was critical for the further
differentiation of activated CD4+ T cells. To
address this question, we used the Transwell system described above.
Forty-eight-hour activated CD4+ T cells were
sorted and reincubated in the upper and lower chambers of the Transwell
system. Forty-eight hours later, the fraction of IFN-- and
IL-2-producing cells was determined after a 4-h stimulation with PMA
and ionomycin. Since the transcription of the IL-2 locus is
progressively lost as CD4+ T cells differentiate
into IFN--producing Th1 cells (22), we could then more
precisely evaluate the differentiation of the distinct
CD4+ T cells populations. Importantly, cell
division was identical under all culture conditions (Fig. 2a
and data not shown). As observed in vivo, efficient Th1 differentiation
was only observed for cells that re-engaged their TCR. Indeed, 47
± 3.5% (n = 3) of the 3A9 CD4+
T cells in the upper well further differentiated into effector Th1
cells producing IFN- but not IL-2 (Fig. 4Ab). In contrast, only
16 ± 4.7% (n = 3) of the 3A9
CD4+ T cells in the lower well were fully mature
Th1 cells producing IFN- but not IL-2 and 14 ± 4.2%
(n = 3) remained undifferentiated producing IL-2 and
IFN- (Fig. 4Ac). Similar results were obtained with
higher doses of HEL peptide, indicating that the poor differentiation
of the 3A9 CD4+ T cells in the lower well did not
result from suboptimal stimulation of the naive
CD4+ T cells (data not shown). In addition, the
limited differentiation of 3A9 CD4+ T cells
present in the lower well into mature Th1 cells was not due to low
levels of IL-12 or IFN-, as similar profiles where obtained when
cycling CD4+ T cells were reincubated with large
amounts of these cytokines (Fig. 4Cb). IL-12 and IFN-
were nonetheless essential for the limited maturation of these cells
since their differentiation was almost completely abolished when
anti-IL-12 and anti-IFN- Ab were added during the 48-h
reincubation period (Fig. 4A, compare a,
c, and e). The differentiation of the 3A9
CD4+ T cells in the upper compartment was also
affected by blocking IL-12 and IFN- (Fig. 4Ad). Indeed,
under those conditions, 25 ± 9.1% (n = 3) of the
3A9 CD4+ T cells differentiated into effector Th1
cells as compared with 47 ± 3.5% when IL-12 and IFN- were
present (Fig. 4A, compare b and d).
Importantly, however, under those conditions, very few cells had an
immature IL-2+IFN-+
phenotype. To ensure that under those conditions IL-12 signaling did
not occur, we restimulated the 48-h activated 3A9
CD4+ T cells with plate-bound anti-CD3, in
the absence of APCs that may produce IL-12 and further added
anti-IL-12 and anti-IFN- blocking Ab. Under those
conditions, differentiation of the 3A9 CD4+ T
cells into mature Th1 cells also occurred, although less efficiently
than when cytokines were also present (Fig. 4C, c
and d). As expected, in the total absence of IL-12 and
IFN-, that is when anti-IL12 and anti-IFN- blocking Ab
were added in both the primary and secondary stimulation periods, no
IFN--producing cells developed (Fig. 4B).
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and TCR
signaling have complementary and synergistic roles in driving the full
differentiation of effector Th1 cells. Signaling through the TCR seems
however more efficient in driving full differentiation of effector Th1
cells that produce IFN- but not IL-2. Interestingly, it was recently
shown that the transcription factor T-bet was essential for Th1 cell
differentiation and could induce IFN- gene expression and
repress IL-2 gene expression (14). We therefore
analyzed T-bet expression in the different populations showing distinct
cytokine profile. Sustained high level of T-bet gene expression correlates with Th1 differentiation
T-bet gene expression was measured by quantitative
RT-PCR in the different culture conditions described above. The level
of expression of T-bet mRNA was very low in naive 3A9
CD4+ T cells and was up-regulated by a 48-h
stimulation in the presence but not in the absence of IL-12 and
IFN-, suggesting that induction of T-bet gene expression
in naive CD4+ T cells requires both TCR and
cytokine signaling (Fig. 5). Sustained
high levels of T-bet mRNA expression correlated with full
differentiation of Th1 effector cells (Fig. 5). Indeed, cycling
CD4+ T cells restimulated with anti-CD3 in
the presence of IL-12 and IFN- that did efficiently differentiate
into effector Th1 cells express a high level of T-bet mRNA. Likewise,
cells restimulated with anti-CD3 in the absence of cytokines, of
which 33% produce IFN- also express high levels of T-bet mRNA. In
contrast, 3A9+ T cells that remained
undifferentiated, i.e., when maintained for an additional 48 h in
medium containing or not polarizing cytokines, had reduced levels of
T-bet mRNA expression. In addition, this study shows that both TCR and
cytokine signaling are necessary for the induction of T-bet expression
in naive T cells. In committed CD4+ T cells
however, expression of T-bet can be maintained by TCR signaling
only.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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-chain-dependent
cytokines that have been involved in the proliferation and survival of
naive or memory T cells (23, 24, 25, 26), is not necessary to
sustain the proliferation of these activated CD4+
T cells (Figs. 1 and 2). This observation is in agreement with the
-chain independence of the clonal expansion of naive T cells in vivo
(27). The continuous presence of IL-2 in vitro is however
essential for survival of the activated 3A9 CD4+
T cells. This may suggest that in vivo, other not yet defined
receptor(s) may deliver the necessary survival signals to activated
CD4+ T cells. The programmed proliferation
induced by initial Ag stimulation is however not endless and will cease
after a total of six to seven divisions when proliferation can be
reinduced by re-exposure to Ag. This early programmed proliferation may
however ensure a significant clonal expansion (64- to 128-fold
increase) of rare Ag-specific T cells even when Ag is limiting, as
during the early phase of an infection.
In sharp contrast, the differentiation of naive
CD4+ T cells into effector Th1 cells requires
repeated contact with Ag-loaded APCs. Indeed, we found both in vitro
and in vivo that when Ag and cytokines are removed 48 h after
primary stimulation, activated T cells did not further differentiate
and produced mainly IL-2 but limited IFN-. Clearly, both cytokine-
and TCR-derived signals were essential at this stage to induce optimal
differentiation. Nonetheless, differentiation, although less efficient,
was also induced by TCR re-engagement or cytokine signaling alone. In
lymphoid organs, the most likely source of IL-12 is dendritic cells
(DC). Mature DC do not constitutively produce IL-12 but are induced to
do so by innate and T cell-derived signals including MHC class II and
CD40 ligation (28, 29, 30, 31). Under these conditions,
CD4+ T cells would be exposed to IL-12 mainly
upon direct interaction with DC. Differentiation of
CD4+ T cells in vivo may be therefore mainly
driven by Ag availability. In agreement, 48-h activated T cells did not
differentiate into IFN--producing Th1 cells when transferred into
hosts preinjected with CFA, that due to microbial constituents ought to
induce IL-12 production (32).
The observed difference in the duration of TCR stimulation required to
induce proliferation or differentiation of naive
CD4+ T cells does not simply reflect difference
in the kinetics of the two processes. Indeed, naive 3A9
CD4+ T cells did not differentiate efficiently
into effector Th1 cells when Ag is retrieved at 48 h, but IL-12
and IFN- signaling is maintained for an additional 48–72 h.
Recurrent TCR engagement is thus more likely required to sustain
expression of specific transcription factors and to induce the
epigenetic modifications that are critical for Th1 effector cells
differentiation. In agreement, our study shows that sustained
expression of T-bet correlates with full differentiation of effector
Th1 cells. Furthermore, our results suggest that the critical role for
TCR re-engagement in inducing effector Th1 differentiation is in
maintaining a high level of T-bet gene expression.
The regulation of Th1 differentiation greatly diverges from that of
CD8+ T cells. Indeed, in
CD8+ T cells, initial Ag encounter will induce a
program of proliferation and differentiation into IFN--producing Tc1
cells that can proceed in the absence of Ag (17, 18). This
programmed differentiation of CD8+ T cells may
reflect the rapid induction of IFN- gene expression,
reaching maximal levels by 48 h of activation, as well as its
IL-12 and STAT-4 independence (33). In addition, transient
exposure to Ag-loaded APCs suffices to induce optimal differentiation
of effector Th2 cells when IL-4 signaling is maintained (Ref.
6 and M. Bajénoff and S. Guerder, unpublished
observation). It is intriguing that CD4+ T cells
have developed specific regulatory circuits to control selectively Th1
effector cell development. Although the biological significance of such
tight regulation, by Ag and cytokine, of Th1 effector cell development
is unknown, it is tempting to speculate that it may simply reflect the
function of effector Th1 cells as compared with undifferentiated or
committed CD4+ T cells in memory responses. We
found that 48-h activated T cells are committed to the Th1 lineage
since they rapidly differentiate into IFN--producing effector cells
upon restimulation. As opposed to effector cells, they are reversible
(M. Bajénoff and S. Guerder, unpublished observation). They may
therefore provide a pool of memory cells able to rapidly respond to
novel Ag challenge with flexible effector function. This population may
correspond to the recently described lymphoid memory cells that were
shown to produce IL-2 only but not IFN- upon 2-h stimulation
(34, 35). In contrast, cells that were stimulated several
times with Ag-loaded APCs may preferentially home to nonlymphoid tissue
where, as effector memory cells, they may produce rapidly IFN- upon
restimulation, serving as an immediate barrier to novel infection
(34, 35). Clearly, unraveling the contribution of these
different memory populations to the control of recurrent infection may
help understanding why Th1 cell differentiation is tightly regulated by
Ag and cytokine availability.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Sylvie Guerder, Center d’Immunologie de Marseille-Luminy, Institut National de la Santé et de la Recherche Médicale/Centre National de la Recherche Scientifique/Université de la Méditerranée, Parc Scientifique de Luminy, Case 906, 13288 Marseille, Cedex 09, France. E-mail address: guerder{at}ciml.univ-mrs.fr
3 Abbreviations used in this paper: HEL, hen egg lysozyme; HPRT, hypoxanthine phosphoribosyltransferase; DC, dendritic cell.
Received for publication September 26, 2001. Accepted for publication December 12, 2001.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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-transgenic model. J. Exp. Med. 182:1579. production. J. Exp. Med. 179:299. Chain required for naive CD4+ T cell survival but not for antigen proliferation. Nat. Immunol. 1:54.[Medline]
production from CD4+ versus CD8+ T cells. J. Exp. Med. 189:1355.This article has been cited by other articles:
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E. L. Wissinger, W. W. Stevens, S. M. Varga, and T. J. Braciale Proliferative Expansion and Acquisition of Effector Activity by Memory CD4+ T Cells in the Lungs following Pulmonary Virus Infection J. Immunol., March 1, 2008; 180(5): 2957 - 2966. [Abstract] [Full Text] [PDF]
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N. J. Singh, M. Cox, and R. H. Schwartz TLR Ligands Differentially Modulate T Cell Responses to Acute and Chronic Antigen Presentation J. Immunol., December 15, 2007; 179(12): 7999 - 8008. [Abstract] [Full Text] [PDF]
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 D. A. Blair and L. Lefrancois Increased competition for antigen during priming negatively impacts the generation of memory CD4 T cells PNAS, September 18, 2007; 104(38): 15045 - 15050. [Abstract] [Full Text] [PDF]
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K. L. Mueller, M. S. Thomas, B. J. Burbach, E. J. Peterson, and Y. Shimizu Adhesion and Degranulation-Promoting Adapter Protein (ADAP) Positively Regulates T Cell Sensitivity to Antigen and T Cell Survival J. Immunol., September 15, 2007; 179(6): 3559 - 3569. [Abstract] [Full Text] [PDF]
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V. R. Moulton, N. D. Bushar, D. B. Leeser, D. S. Patke, and D. L. Farber Divergent Generation of Heterogeneous Memory CD4 T Cells J. Immunol., July 15, 2006; 177(2): 869 - 876. [Abstract] [Full Text] [PDF]
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 C. A. Lazarski, F. A. Chaves, and A. J. Sant The impact of DM on MHC class II-restricted antigen presentation can be altered by manipulation of MHC-peptide kinetic stability J. Exp. Med., May 15, 2006; 203(5): 1319 - 1328. [Abstract] [Full Text] [PDF]
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G. Verdeil, D. Puthier, C. Nguyen, A.-M. Schmitt-Verhulst, and N. Auphan-Anezin STAT5-mediated signals sustain a TCR-initiated gene expression program toward differentiation of CD8 T cell effectors. J. Immunol., April 15, 2006; 176(8): 4834 - 4842. [Abstract] [Full Text] [PDF]
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S. Colombetti, V. Basso, D. L. Mueller, and A. Mondino Prolonged TCR/CD28 Engagement Drives IL-2-Independent T Cell Clonal Expansion through Signaling Mediated by the Mammalian Target of Rapamycin. J. Immunol., March 1, 2006; 176(5): 2730 - 2738. [Abstract] [Full Text] [PDF]
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J. K. Whitmire, N. Benning, and J. L. Whitton Precursor Frequency, Nonlinear Proliferation, and Functional Maturation of Virus-Specific CD4+ T Cells. J. Immunol., March 1, 2006; 176(5): 3028 - 3036. [Abstract] [Full Text] [PDF]
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K. E. Foulds and H. Shen Clonal Competition Inhibits the Proliferation and Differentiation of Adoptively Transferred TCR Transgenic CD4 T Cells in Response to Infection. J. Immunol., March 1, 2006; 176(5): 3037 - 3043. [Abstract] [Full Text] [PDF]
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J. Cassataro, S. M. Estein, K. A. Pasquevich, C. A. Velikovsky, S. de la Barrera, R. Bowden, C. A. Fossati, and G. H. Giambartolomei Vaccination with the Recombinant Brucella Outer Membrane Protein 31 or a Derived 27-Amino-Acid Synthetic Peptide Elicits a CD4+ T Helper 1 Response That Protects against Brucella melitensis Infection Infect. Immun., December 1, 2005; 73(12): 8079 - 8088. [Abstract] [Full Text] [PDF]
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S. Celli, Z. Garcia, and P. Bousso CD4 T cells integrate signals delivered during successive DC encounters in vivo J. Exp. Med., November 7, 2005; 202(9): 1271 - 1278. [Abstract] [Full Text] [PDF]
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H.-J. Ko, S.-Y. Ko, Y.-J. Kim, E.-G. Lee, S.-N. Cho, and C.-Y. Kang Optimization of Codon Usage Enhances the Immunogenicity of a DNA Vaccine Encoding Mycobacterial Antigen Ag85B Infect. Immun., September 1, 2005; 73(9): 5666 - 5674. [Abstract] [Full Text] [PDF]
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R. Obst, H.-M. van Santen, D. Mathis, and C. Benoist Antigen persistence is required throughout the expansion phase of a CD4+ T cell response J. Exp. Med., May 16, 2005; 201(10): 1555 - 1565. [Abstract] [Full Text] [PDF]
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M. Inobe and R. H. Schwartz CTLA-4 Engagement Acts as a Brake on CD4+ T Cell Proliferation and Cytokine Production but Is Not Required for Tuning T Cell Reactivity in Adaptive Tolerance J. Immunol., December 15, 2004; 173(12): 7239 - 7248. [Abstract] [Full Text] [PDF]
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M. A. Williams and M. J. Bevan Shortening the Infectious Period Does Not Alter Expansion of CD8 T Cells but Diminishes Their Capacity to Differentiate into Memory Cells J. Immunol., December 1, 2004; 173(11): 6694 - 6702. [Abstract] [Full Text] [PDF]
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M. S. Zand, B. J. Briggs, A. Bose, and T. Vo Discrete Event Modeling of CD4+ Memory T Cell Generation J. Immunol., September 15, 2004; 173(6): 3763 - 3772. [Abstract] [Full Text] [PDF]
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 K Matsuoka, N Inoue, T Sato, S Okamoto, T Hisamatsu, Y Kishi, A Sakuraba, O Hitotsumatsu, H Ogata, K Koganei, et al. T-bet upregulation and subsequent interleukin 12 stimulation are essential for induction of Th1 mediated immunopathology in Crohn's disease Gut, September 1, 2004; 53(9): 1303 - 1308. [Abstract] [Full Text] [PDF]
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A. Boissonnas, C. Combadiere, E. Lavergne, M. Maho, C. Blanc, P. Debre, and B. Combadiere Antigen Distribution Drives Programmed Antitumor CD8 Cell Migration and Determines Its Efficiency J. Immunol., July 1, 2004; 173(1): 222 - 229. [Abstract] [Full Text] [PDF]
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 O. Wurtz, M. Bajenoff, and S. Guerder IL-4-mediated inhibition of IFN-{gamma} production by CD4+ T cells proceeds by several developmentally regulated mechanisms Int. Immunol., March 1, 2004; 16(3): 501 - 508. [Abstract] [Full Text] [PDF]
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M. Bajenoff and S. Guerder Homing to Nonlymphoid Tissues Is Not Necessary for Effector Th1 Cell Differentiation J. Immunol., December 15, 2003; 171(12): 6355 - 6362. [Abstract] [Full Text] [PDF]
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R. J. De Boer, D. Homann, and A. S. Perelson Different Dynamics of CD4+ and CD8+ T Cell Responses During and After Acute Lymphocytic Choriomeningitis Virus Infection J. Immunol., October 15, 2003; 171(8): 3928 - 3935. [Abstract] [Full Text] [PDF]
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M. Bajenoff, S. Granjeaud, and S. Guerder The Strategy of T Cell Antigen-presenting Cell Encounter in Antigen-draining Lymph Nodes Revealed by Imaging of Initial T Cell Activation J. Exp. Med., September 2, 2003; 198(5): 715 - 724. [Abstract] [Full Text] [PDF]
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N. Auphan-Anezin, G. Verdeil, and A.-M. Schmitt-Verhulst Distinct Thresholds for CD8 T Cell Activation Lead to Functional Heterogeneity: CD8 T Cell Priming Can Occur Independently of Cell Division J. Immunol., March 1, 2003; 170(5): 2442 - 2448. [Abstract] [Full Text] [PDF]
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C. Rush, T. Mitchell, and P. Garside Efficient Priming of CD4+ and CD8+ T Cells by DNA Vaccination Depends on Appropriate Targeting of Sufficient Levels of Immunologically Relevant Antigen to Appropriate Processing Pathways J. Immunol., November 1, 2002; 169(9): 4951 - 4960. [Abstract] [Full Text] [PDF]
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