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Institut National de la Santé et de la Recherche Médicale Unite 454, Montpellier, France;
Laboratoire d’Immunologie des Infections Retrovirales, Institut de Biologie, Montpellier, France; and
Laboratoire de Virologie, Centre Hospitalier Universitaire St. Eloi, Montpellier, France
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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-chain-interacting cytokines as well as signals mediated via
noncognate interactions between activated dendritic cells and memory T
cells are involved in the up-regulation of CXCR4
expression. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Recently, it has been shown that CD45RO+ memory T cells can be divided into two subpopulations, based on the expression of the chemokine receptor CCR7, which controls homing to secondary lymphoid organs (9). Thus, central memory T cells express CCR7 and are capable to migrate to lymphnodes and to interact with dendritic cells. In contrast, another subpopulation of CD45RO+ T cells, called effector memory T cells, is CCR7-, and these cells migrate into and exert their effector function at sites of inflammation (9). Whereas CXCR4 is present on most naive CD45RO- T cells (10, 11), its expression is variable on central memory and effector memory T cells (9). Because IL-4 is mainly produced in response to inflammatory immune reactions, its presence alone cannot account for the constitutive expression of CXCR4 on memory T cells, and therefore it is likely that other, as yet to be determined, factors are involved in the regulation of CXCR4 on these cells as well.
It has been shown previously that IL-2 may also up-regulate CXCR4
expression (11), suggesting that signaling through the
common (c) chain that is shared by both
IL-2R and IL-4R (12) may be involved in the induction of
expression of this chemokine receptor. Thus, likely candidates are
IL-7, IL-9, and IL-15, the receptors that share the
c chain. Importantly, IL-7, like SDF-1, is
produced by stromal cells (13, 14) and acts
synergistically with this chemokine in certain immune functions
(2). In addition, IL-7 promotes HIV infection of T
lymphocytes by X4 HIV strains that use CXCR4 as a coreceptor for viral
entry (15, 16). IL-15 not only induces T cell chemotaxis
(17), but is also ubiquitously present throughout the
immune system.
Interestingly, transcripts for both IL-7 and IL-15, but not for IL-2, IL-4, and IL-9, have recently been shown to be expressed in in vitro-generated, activated, dendritic cells, the in vitro counterparts of Langerhans cells (18). Dendritic cells are not only potent APCs in secondary lymphoid organs, but are also a primary target and a reservoir for HIV infection, in particular for X4 HIV-1 strains (19, 20, 21, 22). Furthermore, activated dendritic cells express numerous cell-surface molecules, notably CD80, CD86, CD40, and CD134L, which following interaction with their respective ligands have been reported to be important in lymphocyte activation, differentiation, and effector function (23, 24, 25, 26) and might therefore be implicated in the regulation of CXCR4 expression as well.
In the present study, we have analyzed the role of cytokines that
signal though the c chain in the regulation of
CXCR4 expression using a population of CXCR4-
CCR7+ T cells generated in vitro from
CD4+ cord blood T lymphocytes that are
reminiscent of central memory T cells (9). In addition,
because of the important role that dendritic cells play in the
pathogenesis of HIV infection, we have investigated whether in
vitro-generated dendritic cells can modulate the expression of CXCR4 on
in vitro-generated central memory T cells, either indirectly, via the
production of cytokines, or directly via cell-cell interaction.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Peripheral or cord blood mononuclear cells were isolated from freshly collected, heparinized peripheral blood (Centre de Transfusion Sanguine, Montpellier, France) or cord blood (generously provided by the Department of Gynecology and Obstetrics, Centre Hospitalier Universitaire Arnaud de Villeneuve, Montpellier, France), respectively, by Ficoll-Hypaque density gradient centrifugation. CD4+ T cells were purified (purity >95%) by negative selection from mononuclear cell preparations, using mAb-coated magnetic beads (Stem Cell Technologies, Vancouver, Canada), according to the manufacturer’s instructions.
T cell clones were generated form purified CD4+ cord blood T cells, following stimulation with plate-bound anti-CD3 and anti-CD28 mAb (see under stimulation and culture conditions) and cloning by limiting dilution, as described previously (27).
Dendritic cells, reminiscent of Langerhans cells, were generated
according to the method described by Caux et al. (28).
Briefly, cord blood CD34+ hemopoietic progenitor
cells were purified by negative selection, using a mixture of
isotype-matched mAb, specific for B cells, CD4+
and CD8+ T cells, monocytes, NK cells,
erythrocytes, and IgG-coated magnetic beads (Stem Cell Technologies).
The isolated cells were 86–95% CD34+, as judged
by staining with the anti-CD34+ mAb B-B34
(Diaclone Research, Besançon, France).
CD34+ cells were cultured in the presence of stem
cell factor (25 ng/ml), GM-CSF (100 ng/ml), and TNF-
(2.5 ng/ml) at
a concentration of 2 x 104 cells/ml in a
75-cm2 flask. Cells were expanded at day 3 with
medium containing fresh GM-CSF and TNF- and maintained at 5 x
104 cells/ml until day 12 at which they were used
in the experiments. The resulting population of dendritic cells were
predominantly CD1a+,
CD14-, CD83+,
HLA-DR+.
Mouse fibroblasts, expressing human (h) CD154 (CD-40L), were a kind gift of Dr. Pierre Garonne, (Schering-Plough France, Dardilly, France). All cell cultures and experiments, with the exception of chemotaxis assays, were conducted in IMDM, supplemented with 10% FCS (Boehringer-Mannhein, Mannheim, Germany) and penicillin, streptomycin, referred to as culture medium.
Cytokines and mAbs
Human rIL-7, rTNF-, recombinant stem cell factor, and
rSDF-1ß were purchased from R&D Systems (Minneapolis, MN), whereas
rIL-9 and rIL-15 were obtained from (Peprotech, London, U.K.). rIL-2,
rIL-4, and rGM-CSF were generous gifts from Drs. Satish Menon and N.
Nagabushan (both at DNAX Research Institute, Palo Alto, CA) and
Gregorio Aversa (Novartis Research Institute, Vienna, Austria),
respectively.
The following neutralizing mAbs were used in this study: the anti-IL-7R mAb R34.34 and the anti-CD154 mAb LL48 (generously provided by Dr. Sem Saeland, Schering-Plough, France), a polyclonal anti-IL-15 Ab (Peprotech), the anti-CD11a mAb L15 and the anti-CD18 mAb L19 (a kind gift of Dr Carl Figdor, University of Nijmegen, The Netherlands), the anti-CD134 mAb 131 and CD134L mAb ik1 (kindly provided by Dr. Toshiyuki Hori, University of Kyoto, Japan), and the anti-86 and CD80 mAbs, B-T7 and 104 (kind gifts of Drs. John Wijdenes, Diaclone Research and André van Agthoven, Immunotech, Marseille, France, respectively).
Stimulation and culture conditions
One million purified cord blood-derived
CD4+ T lymphocytes/ml were stimulated with the
plate-bound anti-CD3 mAb SPV-T3b (Ref. 29 ; 24-well
tissue culture plates (Linbro, McLean, VA) were incubated with 10
µg/ml SPV-T3b in PBS for at least 24 h at 4°C, after which the
plates were washed three times with PBS) and the anti-CD28 mAb B-T3
(1 µg/ml; a generous gift of Dr. John Wijdenes). After 7 days of
culture in the presence of suboptimal concentrations (0.5 ng/ml) of
rIL-2, the cells were collected, washed, restimulated with anti-CD3
and anti-CD28 mAbs, and used in experiments 7 days later. This
procedure resulted in a down-regulation of CXCR4 on 90–99% of the
cells. These cells, which are CD45RO+
CCR7+ (Fig. 1
), are referred to as central memory
T cells (9). Cultures of central memory T cells
(106 cells/ml) with cytokines or cocultures with
autologous dendritic cells (105 cells/ml) were
conducted in the presence or absence of irradiated (50 Gy)
CD154-expressing mouse fibroblasts (25 x
103 cells/ml), in 24-well culture plates (Linbro,
McLean, VA). Alternatively, central memory T cells were incubated with
culture supernatants of dendritic cells that had been stimulated for 3
days with irradiated CD154+ L cell transfectants.
Neutralizing mAbs directed against IL-7R or IL-15, or against CD11a,
CD18, CD40, CD80, CD86, CD134, CD154, and CD134L, were added at a final
concentration of 10 µg/ml. Dendritic cells were cocultured with
CD154+ mouse fibroblasts in 24-well Linbro plates
in a final volume of 500 µl for 9 h, after which central memory
cells and mAbs were added in a volume of 500 µl. After 3 days of
culture with either recombinant cytokines, dendritic cells, or culture
supernatants from activated dendritic cells, in the presence or absence
of mAbs, central memory T cells were used in the various experimental
read-out systems.
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All immunofluoresence and flow cytometry procedures were conducted using the method of Lanier and Recktenwald (30). Cell-surface expression of CXCR4 was analyzed using the biotinylated anti-human CXCR4 mAb 12G5, a biotinylated isotype-matched control mAb (PharMingen, San Diego, CA), and PE-conjugated streptavidin (Becton Dickinson, San Jose, CA). CCR7 expression was measured using the rat mAb 3D12 (IgG2a), a generous gift of Dr. Martin Lipp (Max-Delbrück-Center for Molecular Medicine, Berlin, Germany) and a FITC-labeled goat anti-rat Ab (Caltag, Burlingame, CA). Expression of CD45RO, CD62L, and CD25 was analyzed using APC-conjugated, PE-conjugated, and FITC-conjugated mAbs, respectively (Becton Dickinson). Immunofluorescence was analyzed on a FACScalibur flow cytometer using CellQuest software (Becton Dickinson).
Chemotaxis
Migration of T cells in response to stimulation with SDF-1 was analyzed, using the method described by Bacon and Schall (31). Briefly, 29 µl of Yssel’s medium (Ref. 32 ; purchased from Diaclone Research), supplemented with 1% human AB+ serum, containing different concentrations of human rSDF-1ß, were added to the lower wells of a 96-well ChemoTx-96 disposable chamber with a filter sample site of 3.2 mm diameter and 5-µm pore size filters (Neuroprobe, Gaithersburg, MD). Forty thousand T cells, in a volume of 20 µl, were transferred directly in triplicate onto the filter sample sites. After 1 h of incubation in 5% CO2 at 37°C, cells that had migrated through the filter into the lower chamber were collected, resuspended in culture medium, and counted using a hemocytometer. Results are expressed as the ratio of (number of cells migrated in rSDF-1-containing medium - number of cells migrated in medium alone)/total number of cells used in the assay x 100%.
RNase protection assay
Total RNA was extracted from 5 x 106 central memory T cells that had been cultured for 6 h in the presence or absence of cytokines, using RNAzol B (Tel-Test, Friendswood, TX) and CXCR4 transcripts were detect by RNase protection assay using the multiprobe template set hCR-6, containing the DNA template for CXCR4 and GAPDH and L32 as house keeping genes, according to the manufacturer’s standard protocol (PharMingen). The ratio of expression between CXCR4 and those of GAPDH and L32, respectively, was determined by counting the radio activity (32P) of the relevant bands on the autoradiographs, blotted onto paper filters, by liquid ß-scintillation counting (Wallac, Turku, Finland).
Infection with and detection of HIV-1
CD4+ central memory T cells, were cultured in medium in the absence or presence of either rIL-2, rIL-4, rIL-7, or rIL-15 or with resting or CD40-activated dendritic cells, respectively, in 24-well culture plates in a final volume of 1 ml. After 3 days, viral isolates of the HIV-1 T tropic strains HIV-1NDK (33) or HIV-1LAI (34) were added to the cultures. Thirty minutes before infection with HIV-1, the CXCR4-selective antagonist AMD3100 (Ref. 35 ; a kind gift of Dr. Dominique Schols, Rega Institute, Leuven, Belgium) was added at a final concentration of 10 µg/ml, where indicated. After 5 days of culture, cells were harvested, washed extensively with PBS, and fixed with paraformaldehyde. Frequencies of infected T cells were determined by analyzing the presence of the HIV-1 protein p24, detected by flow cytometry, using an intracellular staining technique as described (36) on a Coulter Epics flow cytometer (Coulter, Hialeah, FL).
Statistical analysis
The 
2 test was used to analyze the
significance of differences in inhibitory activity of mAbs.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Populations of CD4+
CD45RO- cord blood T cells that had been
stimulated twice in vitro with anti-CD3 and anti-CD28 mAbs were
analyzed 7 days after the last stimulation for the expression of
CD45RO, CXCR4, and CCR7 by flow cytometry. All cells were
CD45RO+ (Fig. 1
A) and furthermore expressed
CCR7 (Fig. 1B). As expected, following CD3-mediated
activation, the expression of CXCR4 was strongly down-regulated with
low expression remaining only on 1–10% of cells (Fig. 1C).
Furthermore, all CD45RO+ T cells expressed CD62L
(Fig. 1D). Based on the expression of CCR7 and CD62L, the in
vitro-generated anti-CD3- and anti-CD28 mAb-activated cord
blood cells seem to be reminiscent of the recently described
subpopulation of central memory T cells (9).
To determine whether a counterpart of the population of cultured
CD45RO+ CXCR4-
CCR7+ cells could be detected in vivo, freshly
isolated peripheral blood lymphocytes were analyzed for the expression
of these cell-surface molecules by immunostaining and three-color flow
cytometry. CCR7 was strongly expressed on naive cells, whereas its
expression varied on CD45RO+ cells (Fig. 1E), similar to the results reported by Sallusto et al.
(9). Although CCR7 was expressed on all
CXCR4+ CD45RO- T cells,
both CCR7+ and CCR7- cells
could be detected within the population of
CD45RO+ CXCR4- cells (Fig. 1, F and G). These results support the notion
that central memory T cells generated in vitro might be equivalent to
those detected in vivo (9) and therefore can be used in an
experimental model to study the regulation of CXCR4 expression on these
cells.
Modulation of CXCR4 surface expression on central memory T cells by
cytokines that bind to c chain-containing receptors
The capacity of cytokines that signal via the
c chain to induce CXCR4 expression was
determined by culturing central memory T cells in the presence of
rIL-2, rIL-4, rIL-7, rIL-9, or rIL-15 for different periods of time and
analyzing the cell-surface expression of CXCR4 by flow cytometry. As
shown in Fig. 2, rIL-2, rIL-4, rIL-7, and
rIL-15 induced CXCR4 expression to variable extent. Whereas rIL-4 and
rIL-7 induced strong and comparable levels of CXCR4 expression, rIL-2
and rIL-15 were slightly less efficient. Higher concentrations of any
of the cytokines did not further augment surface levels of CXCR4.
Maximal CXCR4 expression was detected between 48 and 72 h after
addition of the cytokines. In addition, all cytokines were able to
induce a proliferative response in central memory T cells (results not
shown). In contrast, rIL-9 was unable to induce the expression of CXCR4
(Fig. 2), even at concentrations as high as 100 ng/ml, nor did it
induce a proliferative response in these cells (results not shown).
Next, we investigated whether combinations of cytokines had additive or
synergistic effects on the induction of CXCR4 expression. Culture of
memory T cells in the presence of combinations of cytokines did not
modulate the induction of CXCR4 expression, induced by each cytokine
alone (data not shown). Similarly, rIL-9 did not affect the rIL-2-,
rIL-4-, rIL-7-, or rIL-15-mediated induction of CXCR4 expression. As a
control, the cells were cultured in medium alone or in the presence of
various concentrations of rGM-CSF, which does not signal via the
c chain. In neither culture condition was
CXCR4 expression induced (Fig. 2).
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To investigate whether cytokine-mediated induction of CXCR4
surface expression was due to a increase in CXCR4 transcription,
expression of CXCR4 mRNA was analyzed using a RNase protection assay. A
representative experiment is shown in Fig. 3A. Incubation of central
memory T cells with 10 ng/ml of rIL-7 or rIL-15 for 6 h resulted
in a 7.5- and 5.0-fold induction, respectively, of specific transcripts
for CXCR4, as compared with CXCR4 mRNA expression in cells cultured in
medium alone (Fig. 3B). rIL-4 induced the strongest increase
in CXCR4 mRNA (about 14-fold), whereas rIL-2, at similar
concentrations, had a significant but lower effect (3-fold) on CXCR4
transcription. The increase observed at 6 h was higher than that
observed at 24 h following activation for all cytokines except
rIL-9, which did not affect CXCR4 transcription (results not
shown).
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CXCR4 is involved in the migration of T cells via interaction with
its natural ligand, SDF-1. To determine whether cytokine-induced
modulation of CXCR4 expression was reflected in functional changes of
the responding cells, their in vitro chemotactic properties in response
to stimulation with SDF-1 were studied. Following a 3-day culture in
the presence of rIL-4, rIL-7, or rIL-15, central memory T cells
migrated in a dose-dependent fashion in response to SDF-1. For T cells
cultured in rIL-15, a net migration index of 50% was reached at a
concentration of 250 ng/ml, whereas 75% of cells cultured in IL-4 or
IL-7 migrated in response to the same concentrations of SDF-1 (Fig. 4). The functionality of cytokine-induced
CXCR4 expression was furthermore demonstrated by the increased
susceptibility of central memory T cells to infection with the X4
HIV strains HIV-1NDK and
HIV-1LAI (Fig. 5
and Table I). A 3-day culture of the
latter cells in the presence of either rIL-2, rIL-4, rIL-7, or rIL-15,
before infection with HIV-1NDK, resulted in
infection rates between 30 and 60% as demonstrated by the
presence of intracellular p24, whereas about 7% of cells cultured in
medium alone were infected with this virus (Fig. 5). The addition of
the CXCR4-specific antagonist AMD3100 to cultures of T cells that had
been precultured with rIL-2 or rIL-4 resulted in a total inhibition of
infection by HIV-1LAI, demonstrating the direct
involvement of CXCR4 in the cytokine-induced susceptibility of HIV-1
infection. The addition of AMD3100 inhibited
HIV-1NDK infection in rIL-2- and rIL-4-treated
cells by about 80 and 60%, respectively (Table I), indicating
that this strain might use chemokine receptors other than CXCR4 for
entry into T cells. Taken together, these results demonstrate that
cytokines induce the expression of a functional CXCR4 on central memory
T cells.
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In view of the effects of rIL-7 and rIL-15 in inducing CXCR4
expression, we analyzed whether CD34+ hemopoietic
progenitor cell-derived dendritic cells, were able to up-regulate CXCR4
expression on T cells in a similar manner. To this purpose, central
memory T cells were cocultured with autologous in vitro-differentiated
dendritic cells in the presence or absence of irradiated,
CD154-expressing, mouse fibroblasts, and CXCR4 expression was
determined by flow cytometry. We specifically monitored CXCR4
expression on T cells characterized by their forward and side scatter
properties, as well as their expression of CD3. Whereas nonactivated
dendritic cells induced low levels of CXCR4 expression on central
memory T cells, coculture with dendritic cells, stimulated with
CD154-expressing fibroblasts, resulted in a strong induction of CXCR4
(Fig. 6). The kinetics of induction of
CXCR4 expression were similar to that observed for exogenous cytokines
(data not shown).
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Induction of CXCR4 on central memory T cells by activated dendritic cells involves CD40/CD154 and CD134/CD134L interactions
Although cocultures of central memory T cells with activated
dendritic cells resulted in induction of CXCR4 expression, culture
supernatants from the latter cells were ineffective (data not shown).
In addition, neutralizing anti-IL-15 or IL-7R mAbs had no effect,
indicating that dendritic cell-mediated induction of CXCR4 expression
is unlikely to be due to endogenously produced cytokines. Therefore,
the nature of the signal mediated by the interaction of activated
dendritic cells and central memory T cells was investigated, using
neutralizing mAbs directed against various costimulatory and adhesion
molecules known to be expressed at the cell surface of either activated
dendritic cells or T cells. For these experiments, dendritic cells were
preactivated with CD154-expressing fibroblasts for 9 h, and
subsequently added to central memory T cells, in the presence or
absence of neutralizing mAbs. The addition of neutralizing mAbs
directed at CD40 or CD134, as well as their ligands CD154 and CD134L,
respectively, inhibited the induction of CXCR4 by activated dendritic
cells by about 50% (Fig. 7 and Table II). Moreover, the addition of both
anti-CD40 and anti-CD134 or anti-CD154 and anti-CD134L
mAbs resulted in an additive effect with a nearly complete inhibition
of the induction of CXCR4 expression by activated dendritic cells. In
contrast, neutralizing mAbs specific for CD80 and CD86, the ligands for
CD28, did not, or only marginally, inhibited the induction of CXCR4,
whereas mAbs specific for CD11a and CD18 had no effect on dendritic
cell-induced expression of CXCR4 (Table II).
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As shown previously (8), the expression of CXCR4 is
down-regulated following Ag-specific activation of T cells, and its
expression is mutually exclusive with that of CD25, a marker of T cell
activation. Therefore, it was of interest to determine whether
activated dendritic cells were able to simultaneously activate central
memory T cells and to induce CXCR4 expression, following a noncognate
interaction. Interestingly, about 30% of the T cells that had
up-regulated CXCR4, following interaction with CD40-activated dendritic
cells, coexpressed CD25 (Fig. 8C), whereas the addition of
exogenous rIL-4 or rIL-7 further enhanced CXCR4 expression (not shown).
In contrast, however, the addition of an anti-CD3 mAb to T
cell/activated dendritic cell cultures, irrespective of the presence of
exogenous cytokines resulted in a rapid loss of expression of CXCR4 and
a concomitant strong enhancement of CD25 expression (Fig. 8D). Taken together, these results suggest that
CD40-stimulated dendritic cells are able to deliver an "activation"
signal(s) to T cells, resulting in the up-regulation of CXCR4, as well
as in the induction of CD25 expression, via a pathway that is
functionally distinct from that induced by triggering of the TCR/CD3
complex.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The CXCR4 expression-inducing cytokines IL-2, IL-4, IL-7, and IL-15 all
bind to the c chain, suggesting that the
JAK/STAT signaling pathway activated through their receptors is
important for CXCR4 induction. Of note, rIL-9, the receptor of which
also contains the c chain, did not induce
CXCR4 expression; however, as it also failed to induce T cell
proliferation, it appears that IL-9 does not act on T lymphocytes (Ref.
38, 39 and this report). Cytokine-mediated induction of
CXCR4 expression on memory T cells involves c
chain-mediated signaling, because its induction was inhibited, although
to variable extent, in the presence of the inhibitor AG-490, which
blocks signaling via JAK-3, a tyrosine kinase that binds specifically
to the c chain (P. Jourdan and H. Yssel,
unpublished results).
It is of note that activation of this cytokine-mediated signal transduction pathway does not seem to be sufficient for the induction of CXCR4 surface expression. Specifically, rIL-2 and rIL-15 failed to induce CXCR4 expression in cord blood-derived T cell clones, reminiscent of the CD45RO+ CCR7- effector memory T cells, described by Sallusto et al. (9), whereas they induced strong proliferative responses in these cells (data not shown). The observed proliferation indicates that, although the JAK/STAT pathway is involved in this process, the activation of cytoplasmic substrates following cytokine stimulation in these cells was not adequate for CXCR4 up-regulation. This was not due to a particular property of cord blood-derived T lymphocytes, because similar results were obtained using T cell clones generated from peripheral blood or human skin. The identity of other factors required for cytokine-induced CXCR4 up-regulation is currently being assessed.
Dendritic cells, derived from CD34+ hemopoietic progenitor cells and activated via CD40, have been shown to express transcripts for a large variety of cytokines, including IL-7 and IL-15, but not IL-2, IL-4, or IL-9 (16). However, it is not clear whether these cells secrete IL-7 and IL-15 protein. Our observation that culture supernatants of activated dendritic cells were unable to induce either CXCR4 expression or proliferative responses by effector memory T cells indicates that this particular subpopulation of dendritic cells does not secrete IL-7 and IL-15 at levels that are sufficient to induce biological activity. It has been suggested that the membrane-bound form of IL-15, rather than soluble IL-15, is biologically active (40). However, the lack of inhibitory activity of a neutralizing anti-IL-15 mAb added to cocultures of effector memory T cells and activated dendritic cells (data not shown) suggests that membrane-bound IL-15 does not play an important role in our culture system.
The importance of the interaction of dendritic cells and T cells in the transmission of HIV has been reported extensively in the literature (19, 21, 22, 41, 42, 43). In addition, the ability of dendritic cells to augment infection of T cells by HIV T tropic strains was found to be mediated, at least in part, by the interaction of CD40 and its ligand (44). However, the effect of CD40-mediated signaling on the expression of coreceptors for HIV has not been directly addressed in these studies. Here, we demonstrate that the increased susceptibility of T cells to infection with T tropic strains of HIV, induced by autologous activated dendritic cells, is associated with an up-regulation of CXCR4 expression. This is due, at least in part, to the interaction of CD40 and CD134 with their respective ligands. In addition, we find that CD40 and CD154 play a dual role in this process because the interaction of these molecules is involved in the activation of the dendritic cells, as well as in the propagation of a CXCR4 expression-inducing signal.
The required activation of dendritic cells in vivo involves activated T cells, which express high levels of CD154. CD134 is also expressed on activated T cells (25), whereas CD134L is expressed on dendritic cells following CD40-mediated activation (26). It is possible that central memory T cells express CD154 and CD134 on their surface, which, albeit undetectable by flow cytometry (data not shown), might be functionally active. Interaction of CD154 with CD40, constitutively expressed on dendritic cells, would then result in the up-regulation of other costimulatory molecules and their ligands. This reciprocal dialogue between activated dendritic cells and central memory T cells will lead to the induction of CXCR4 expression on the T cells. Because both central memory T cells and activated Langerhans cells are able to specially migrate to secondary lymph nodes, it is likely that this type of noncognate interactions takes place in these sites.
In conclusion, regulation of CXCR4 expression on T cells involves a
wide range of signals, including soluble factors such as IL-2, IL-4,
IL-7, and IL-15, the receptors of which share the
c chain, cell-surface molecules like CD134 and
CD154, as wells as other unrelated stimuli such as those mediated by
glucocorticoids (45). Further studies will be necessary to
determine the precise role, under physiological conditions, of each of
these CXCR4 regulatory stimuli on T cell migration and HIV
infection.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Hans Yssel, Institut National de la Santé et de la Recherche Médicale Unite 454, Centre Hospitalier Universitaire Arnaud de Villeneuve, 371 Avenue Doyen Gaston Giraud, 34295 Montpellier, Cedex 5, France.
3 Abbreviations used in this paper: SDF-1, stromal-derived factor-1; c, common ; h, human.
Received for publication October 1, 1999. Accepted for publication April 26, 2000.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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cooperate in the generation of dendritic Langerhans cells. Nature 360:258.[Medline]
-upregulated interleukin-15. Blood 93:3531.This article has been cited by other articles:
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  E. N. M. Nolte-'t Hoen, E. P. J. Boot, J. P. A. Wagenaar-Hilbers, J. H. M. van Bilsen, G. J. A. Arkesteijn, G. Storm, L. A. Everse, W. van Eden, and M. H. M. Wauben Identification and monitoring of effector and regulatory T cells during experimental arthritis based on differential expression of CD25 and CD134 J. Leukoc. Biol., January 1, 2008; 83(1): 112 - 121. [Abstract] [Full Text] [PDF]
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 A. Audige, E. Schlaepfer, H. Joller, and R. F. Speck Uncoupled Anti-HIV and Immune-Enhancing Effects when Combining IFN-{alpha} and IL-7 J. Immunol., September 15, 2005; 175(6): 3724 - 3736. [Abstract] [Full Text] [PDF]
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K. Beider, A. Nagler, O. Wald, S. Franitza, M. Dagan-Berger, H. Wald, H. Giladi, S. Brocke, J. Hanna, O. Mandelboim, et al. Involvement of CXCR4 and IL-2 in the homing and retention of human NK and NK T cells to the bone marrow and spleen of NOD/SCID mice Blood, September 15, 2003; 102(6): 1951 - 1958. [Abstract] [Full Text] [PDF]
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L. A. Napolitano, C. A. Stoddart, M. B. Hanley, E. Wieder, and J. M. McCune Effects of IL-7 on Early Human Thymocyte Progenitor Cells In Vitro and in SCID-hu Thy/Liv Mice J. Immunol., July 15, 2003; 171(2): 645 - 654. [Abstract] [Full Text] [PDF]
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J. D. Estes, B. F. Keele, K. Tenner-Racz, P. Racz, M. A. Redd, T. C. Thacker, Y. Jiang, M. J. Lloyd, S. Gartner, and G. F. Burton Follicular Dendritic Cell-Mediated Up-Regulation of CXCR4 Expression on CD4 T Cells and HIV Pathogenesis J. Immunol., September 1, 2002; 169(5): 2313 - 2322. [Abstract] [Full Text] [PDF]
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T. J. Fry and C. L. Mackall Interleukin-7: from bench to clinic Blood, May 13, 2002; 99(11): 3892 - 3904. [Full Text] [PDF]
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A. Llano, J. Barretina, A. Gutierrez, J. Blanco, C. Cabrera, B. Clotet, and J. A. Este Interleukin-7 in Plasma Correlates with CD4 T-Cell Depletion and May Be Associated with Emergence of Syncytium-Inducing Variants in Human Immunodeficiency Virus Type 1-Positive Individuals J. Virol., November 1, 2001; 75(21): 10319 - 10325. [Abstract] [Full Text] [PDF]
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D. Misse, P.-O. Esteve, B. Renneboog, M. Vidal, M. Cerutti, Y. St Pierre, H. Yssel, M. Parmentier, and F. Veas HIV-1 glycoprotein 120 induces the MMP-9 cytopathogenic factor production that is abolished by inhibition of the p38 mitogen-activated protein kinase signaling pathway Blood, August 1, 2001; 98(3): 541 - 547. [Abstract] [Full Text] [PDF]
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P. Hjelmström Lymphoid neogenesis: de novo formation of lymphoid tissue in chronic inflammation through expression of homing chemokines J. Leukoc. Biol., March 1, 2001; 69(3): 331 - 339. [Abstract] [Full Text]
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) or L32 mRNA (
), respectively
(B). The results represent the mean ± SD of two
independent experiments.
