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*
Laboratory of Immunology, Istituto Dermopatico dell’Immacolata, Istituto di Ricovero e Cura a Carattere Scientifico, Rome, Italy; and
Department of Experimental and Diagnostic Medicine, Section of General Pathology, University of Ferrara, Ferrara, Italy
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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, IL-1
, TNF-, IL-6, and
IL-12, whereas IL-1 receptor antagonist and IL-10 production was not
affected. As a result, T cell lines generated from allogeneic naive
CD45RA+ T cells primed with DCs matured in the presence of
ATP produced lower amounts of IFN-
and higher levels of IL-4, IL-5,
and IL-10 compared with T cell lines obtained with LPS-stimulated DCs.
ATP inhibition of TNF- and IL-12 production by mature DCs was not
mediated by PGs or elevation of intracellular cAMP and did not require
ATP degradation. The inability of UTP and the similar potency of ADP to
reproduce ATP effects indicated that ATP could function through the P2X
receptor family. These results suggest that extracellular ATP may serve
as an important regulatory signal to dampen IL-12 production by DCs and
thus prevent exaggerated and harmful immune
responses. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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provided by T cells themselves (1, 2, 3, 4, 5). At the
initial stage of T cell-DC interaction, the cytokine microenvironment
plays a key role in Th cell differentiation toward Th1 or Th2 cell
types. In particular, mature DCs produce high levels of IL-12 and
preferentially promote the development of Th1 responses
(6). DC maturation is thus a fundamental checkpoint in the
initiation and shaping of immune responses, and it is tightly
regulated. In particular, IL-10, TGF-, corticosteroids, vitamin
D3, and PGE2 inhibit DC
maturation induced by different stimuli and can convert DCs from Th1-
to Th2-skewing APCs (2).
Extracellular nucleotides have emerged as important regulators of
inflammatory and immune responses. In particular, ATP can affect the
functions of B cells, T cells, macrophages, and eosinophils
(7, 8, 9) via activation of plasma membrane receptors known
as P2 purinoceptors (10, 11, 12). Two P2 receptors subfamilies
have been recognized: P2X and P2Y, the former identified as membrane
channels and the latter as seven membrane-spanning G protein-coupled
receptors (10, 13, 14). ATP can be released by regulated
exocytosis, traumatic cell lysis, or passive leakage from damaged
cells, and thus extracellular ATP concentration is likely raised during
tissue injury or inflammation (10). Recent evidence from
our and other laboratories showed that functional P2 receptors of the
P2X and P2Y subtypes are expressed on both mouse and human DCs
(15, 16, 17, 18). Most studies however have addressed the effects
of acute exposure and using relatively high doses of extracellular ATP.
In this study, we report on the ability of chronic (24 h) stimulation
with low doses of extracellular ATP to affect the maturation and
Ag-presenting functions of DCs. The major finding of this study was
that ATP at micromolar concentrations induced a distorted maturation of
DCs, with blocked LPS- and CD40L-dependent production of IL-1, TNF-,
IL-6, and IL-12 but unaffected release of IL-1 receptor antagonist
(IL-1ra), and IL-10. As a result, mature DCs exposed to ATP showed an
impaired ability to initiate Th1 responses.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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LPS (from Escherichia coli 055:B5), ADP, and
indomethacin were purchased from Sigma-Aldrich (Milan, Italy). ATP,
ATP--S, and UTP were obtained from Boehringer Mannheim (Mannheim,
Germany). Soluble CD40L (sCD40L) was obtained from Alexis (San Diego,
CA). Adenosine receptors antagonists 8-cyclopentyl-1,3-dipropylxanthine
(DPCPX), 8-(3-chlorostyryl)caffeine (CSC), MRS-1220, and the inhibitor
of cAMP-mediated activation of protein kinase A, Rp-adenosine
3',5'-cyclic monophosphothioate triethylamine (Rp-cAMPS), were
purchased from Research Biochemicals International (Natick, MA).
The mAbs FITC-conjugated and pure anti-HLA-DR, anti-CD45RO and
anti-CD45RA, and FITC-conjugated anti-CD14, anti-CD3, and
anti-CD4 were purchased from Becton Dickinson (San Jose, CA).
FITC-conjugated anti-CD1a, anti-CD86, anti-CD40, and
anti-IFN-; pure anti-CD28; and PE-conjugated rat
anti-IL-4 were obtained from PharMingen (San Diego, CA).
FITC-conjugated anti-CD54, anti-CD80, and anti-CD83 and
pure anti-CD3 were purchased from Immunotech (Marseille, France).
Anti-MHC class I was obtained from Dako (Glostrup, Denmark). Control
mouse or rat Ig were purchased from Becton Dickinson or PharMingen. The
anti-human IL-10 mAb and human rIL-12 were provided by R&D Systems
(Minneapolis, MN). Anti-IL-10 mAb, sCD40L, and ATP had undetectable
endotoxin levels (<10 pg/mg) by the Limulus amebocyte
lysate assay (BioWhittaker, Walkersville, MD).
DC preparation and stimulation
DCs were prepared from purified peripheral blood monocytes of healthy individuals, as described previously (19). Briefly, >90% pure CD14+cells were cultured at 1 x 106 cells/ml in RPMI 1640 supplemented with 1 mM sodium pyruvate, 0.1 mM nonessential amino acids, 2 mM L-glutamine, 25 mM HEPES, 0.05 mM 2-ME, 100 U/ml penicillin, and 100 µg/ml streptomycin (all from Life Technologies, Chagrin Falls, OH; complete RPMI) supplemented with 10% FBS, 100 ng/ml human rGM-CSF, and 200 U/ml human rIL-4 (R&D Systems) at 37°C with 5% CO2. Medium was changed after 3 days, and, at day 6 of culture, cells were recovered and depleted of CD2+ and CD19+ cells by means of immunomagnetic beads coated with specific mAbs (Dynal, Oslo, Norway). This procedure gave >97% pure CD1a+ and CD14- DC preparations. DCs were left untreated or incubated for 24 h at 37°C with nucleotides or induced to mature with LPS (10 µg/ml) or sCD40L (1 µg/ml) in the presence or absence of nucleotides. Where indicated, DCs cultures were added with 25 µM DPCPX, 25 µM indomethacin, 50 µM MRS-1220, 1 µM CSC, or 300 µM Rp-cAMPS 30 min before the addition of ATP.
Flow cytometry analysis of DCs
DCs either untreated or stimulated for 24 h with LPS or sCD40L in the presence or absence of nucleotides were washed and then incubated with FITC-conjugated mAbs in PBS containing 2% FBS and 0.01% NaN3 for 40 min at 4°C. When pure mAbs were used, a second incubation with a FITC-coupled goat (Fab')2 anti-mouse IgG (Southern Biotechnology Associates, Birmingham, AL) was performed. Matched isotype mouse Ig were used in control samples. To detect DC apoptosis and necrosis, cells were stained with FITC-conjugated annexin V and propidium iodide using the annexin V-FITC apoptosis detection kit from Genzyme (Cambridge, MA). Cells were analyzed using a FACScan (Becton Dickinson).
Quantitation of endocytosis
DCs stimulated as reported above were washed, resuspended in complete medium, and then incubated with 1 mg/ml Texas Red-conjugated BSA (Molecular Probes, Eugene, OR) at 37°C or 4°C. At selected time points, uptake was stopped by adding cold PBS containing 2% FBS and 0.01% NaN3. Cells were then washed four times and analyzed in a FACScan. Surface binding values obtained by incubating cells at 4°C were subtracted from values measured at 37°C.
DC cytokine release
IL-10 was measured in DC supernatants by ELISA using a matched
pair of mAbs from PharMingen. IL-1, IL-1, IL-1ra, and IL-6
concentrations were determined using ELISA kits from R&D Systems. IL-12
(p70) and TNF- were measured using OptiEIA kits from PharMingen.
Samples were assayed in triplicate for each condition.
RNase protection assay
Total RNA was extracted from purified DCs after a 24-h incubation with the indicated stimuli using TRIzol (Life Technologies) according to the manufacturer’s instructions. Two multiprobe template sets, hCK2 and hCK3 (RiboQuant; PharMingen), were used for in vitro transcription reactions in the presence of a GACU pool and a T7 RNA-polymerase to synthesize [32P]UTP-labeled antisense probes. RNase protection analysis of 3 µg of total RNA was performed after overnight hybridization at 60°C with 2.5 x 106 cpm of hCK2 or hCK3, followed by digestion with RNase A and T1 according to standard protocols. Protected fragments were treated with proteinase K, extracted with phenol-chloroform plus isoamyl alcohol (50:1), and finally precipitated in ethanol in the presence of ammonium acetate. The samples were electrophoresed on 5% denaturing sequencing gels and then exposed to Kodak films (Kodak, Rochester, NY).
DC-T cell cocultures
For the primary MLR assay, T lymphocytes were purified (>95%
CD3+) from the heavy density fraction (50–60%)
of Percoll gradients followed by immunomagnetic depletion using a
mixture of anti-HLA-DR and anti-CD19 mAb-conjugated beads
(Dynal). DCs were washed and then cultured in 96-well microculture
plates in serial dilutions along with 1.5 x
105/well allogeneic T lymphocytes in complete
RPMI complemented with 5% human serum. Cocultures were pulsed at day 3
with 1 µCi/well [3H]thymidine for about
16 h at 37°C and then harvested onto fiber-coated 96-well
plates. Radioactivity was measured in a beta counter. Results are given
as mean cpm ± SD of triplicate cultures. Where indicated,
allogeneic naive T cells were purified (>95%
CD45RA+) by incubation of T cells with
anti-CD45RO mAb followed by a goat anti-mouse Ig coupled to
immunomagnetic beads and then cocultured (106
cells/well) with DCs (5 x 104 cells/well)
in a 24-well plate in complete RPMI plus 5% human serum for 6 days.
Thereafter, T cells were restimulated or not with plate-coated
anti-CD3 and soluble anti-CD28 mAbs (both at 1 µg/ml), and
examined for intracellular IFN- and IL-4 after 6 h. For
two-color intracellular staining, monensin (10 µM; Sigma, St. Louis,
MO) and brefeldin A (10 µg/ml; Sigma) were added to the cultures
before the staining to prevent cytokine secretion. T cells were then
fixed with 2% paraformaldehyde, permeabilized with 0.5% saponin,
stained with FITC-conjugated mouse anti-IFN- and PE-conjugated
rat anti-IL-4, and finally analyzed with a FACScan. In control
samples, staining was performed using isotype-matched control Ab.
Cytokines were also measured in the T cell supernatants 48 h after
activation by ELISA using matched pairs of mAbs (for IL-4, IL-10, and
IFN-) and OptEIA kit (for IL-5) from PharMingen. T cells that were
not restimulated at day 6 did not show any lymphokine production (data
not shown).
Statistical analysis
The Mann-Whitney U test was used to compare
differences in cytokine release, endocytosis, and T cell proliferation.
The p values 
0.05 were considered significant.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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DCs maturation is associated with higher expression of MHC and
costimulatory molecules, as well as de novo expression of CD83.
Immature DCs incubated for 24 h with 250 µM ATP displayed
enhanced levels of CD83, CD86, CD80, and CD54 (Fig. 1
), as previously reported by Berchtold
et al. (16). In addition, ATP further increased the
expression of CD83, CD86, and CD54, but not CD80, induced by either LPS
or sCD40L, whereas the expression of MHC molecules was not at all
affected. The effects of ATP on CD83, CD86, CD80, and CD54 membrane
levels were dose dependent (Fig. 2). As a
consequence of the increased expression of CD86, but not CD80, the
CD86:CD80 ratio was 30–40% higher in ATP/LPS- or
ATP/sCD40L-stimulated DCs compared with DC stimulated solely with LPS
or sCD40L. A reduced ability to capture exogenous Ags is an early event
during DC maturation and has been demonstrated for fluid phase tracers,
receptor-mediated endocytosis, as well as for the uptake of
particulates and bacteria (1, 19). In keeping with these
observations, ATP induced a significant decrease in the uptake of BSA
in immature DCs (Fig. 3A).
Since ATP affected DC phenotype and endocytic activity, we next studied
the effect of ATP on the Ag-presenting function of DCs. Fig. 3B shows that ATP stimulation increased the capacity of
immature DCs to induce proliferation of allogeneic naive
CD45RA+ T lymphocytes. In contrast, ATP did not
increase the alloantigen-presenting functions of LPS-stimulated
DCs.
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A major attribute of mature DCs is the synthesis and release of
cytokines with important modulatory function on inflammation and T cell
differentiation. ATP did not induce cytokine release by immature DCs,
with the exception of IL-1ra, whereas strongly inhibited LPS or sCD40L
induced production of TNF-, IL-1, IL-1, IL-6, and IL-12. In
contrast, IL-10 and IL-1ra synthesis and release by mature DCs were
only slightly or not affected (Table I
and Fig. 4). The changes in cytokine
production were not due to a toxic effect of ATP since neither the
trypan blue exclusion test nor lactate dehydrogenase release assay
showed increased cell death after 250 µM ATP stimulation for 24
h (data not shown). In addition, 250 µM ATP did not induce DC
apoptosis (see below). The cytokine profile of DCs matured in the
presence of extracellular ATP was thus characterized by abundant IL-10
and IL-1ra, but absent production of IL-12 and proinflammatory
cytokines. Because autocrine IL-10 inhibits IL-12 and TNF-
production by DCs,5 we
stimulated DCs with LPS along with ATP in the presence of
anti-IL-10-neutralizing mAb or isotype control mAb. Neutralization
of autocrine IL-10 only partially restored IL-12 and TNF-
production, indicating that ATP suppression of IL-12 and TNF-
production by mature DCs was largely independent from autocrine IL-10
(data not shown).
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IL-12 released by mature DCs is the most important factor that
drives the differentiation of naive T cells toward the
IFN--producing Th1 phenotype (6). Given the inhibitory
effect of ATP on IL-12 production by DCs, we next examined the quality
of primary T cell response induced by DCs matured in the presence of
extracellular ATP. Naive allogeneic T cells primed with mature DCs
differentiated into Th1 lymphocytes as they produced large amounts of
IFN- and low levels of IL-4, IL-5, and IL-10 (Fig. 5). In contrast, T lymphocytes primed
with DCs matured in the presence of ATP displayed a reduced production
of IFN- and a higher release of IL-4, IL-5, and IL-10. Addition of
rIL-12 at the beginning of the T cell culture restored IFN-
production and inhibited IL-4, IL-5, and IL-10 release, suggesting that
most of the effects of ATP on the T cell-differentiating properties of
DCs were consequent to inhibition of IL-12 production.
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release by ATP is not mediated by PGs
or elevation in intracellular cAMP and does not require extracellular
ATP degradation
PGE2 and various cAMP-elevating agents such
as cholera toxin, rolipram, and cicaprost have been reported to inhibit
TNF- and IL-12, but not IL-10 production by monocytes and DCs
(20, 21, 22). Fig. 6 shows that
blocking PG synthesis by indomethacin or the activation of
cAMP-dependent protein kinase A by Rp-cAMPS could not restore TNF-
and IL-12 production. Interestingly, indomethacin markedly increased
IL-12 and TNF- release by mature DCs, confirming previously
published data (23). Extracellular nucleotides are
subjected to rapid degradation by plasma membrane ectoenzymes, with ATP
and ADP degraded to AMP by apyrase, and AMP metabolized to adenosine by
5'-nucleotidase (24). DCs express functional apyrase and
5'-nucleotidase (16, 25), and adenosine and its metabolite
inosine have been reported to inhibit TNF- and IL-12 production by
monocytes (26). Incubation of DCs with the adenosine
receptors antagonist DPCPX did not prevent the inhibitory effect of ATP
on TNF- and IL-12 production by maturing DCs (Fig. 6). Selective
inhibitors of adenosine A1 (8CSC) and
A2 (MRS-1220) receptors also failed to counteract
ATP effects (data not shown). To further asses whether ATP could act
through its metabolites, we tested the nonhydrolyzable ATP analogue
ATP--S, which dose-dependently inhibited TNF- and IL-12 release
by LPS-stimulated DCs with an efficiency 100-fold higher than ATP (Fig. 7). However, ATP--S at concentrations
10 µM reduced also IL-10 release, most likely as a result of a
direct toxic effect. Indeed, ATP--S at these concentrations
increased significantly the number of apoptotic and necrotic DCs, a
property not shared by ATP, UTP, or ADP (Fig. 8 and data not shown). Fig. 7 also shows
that UTP, which activates P2Y but not P2X receptors, was unable to
change the cytokine profile of mature DCs. In contrast, ADP inhibited
TNF- and IL-12 production by mature DCs, with a potency similar
to ATP.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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, but higher amounts of IL-4, IL-5, and IL-10.
Addition of exogenous IL-12 at the time of T cell priming restored a
potent Th1 polarization, suggesting that the immune deviation induced
by ATP is mainly due to the suppression of IL-12 production in maturing
DCs. Moreover, DCs matured in the presence of ATP expressed a higher
CD86:CD80 ratio, and this can be an additional factor that may favor
the induction of Th2 response, in line with the selective role of CD86
in Th2 polarization (27, 28).
DC maturation is an important control point in the development of
immune responses, as it converts Ag-capturing DCs into potent
immunostimulatory APCs and can also determine whether peripheral
tolerance or immunity is induced (29). Maturation of DCs
is stimulated by factors signaling tissue danger such as
microorganisms, dying cells, and proinflammatory cytokines. Recently, a
variety of factors has emerged that can limit DC maturation. For
example, intracellular cAMP-elevating agents such as
PGE2, cholera toxin, and
2-adrenoreceptor agonists inhibit IL-12 and
TNF- and enhance IL-10 expression by LPS-stimulated DCs (21, 23, 30). In contrast, IL-10, glucocorticoids, and vitamin
D3 interfere with DC maturation as a whole by
blocking the up-regulation of presenting and costimulatory molecules,
the release of IL-12 and TNF- but not IL-10, and maintain DCs in an
Ag-capturing state (31, 32, 33). Extracellular ATP acted
independently of the above-cited mechanisms since its activity was not
affected by indomethacin, inhibition of cAMP-dependent protein kinase
A, or neutralization of endogenous IL-10. We also excluded that ATP
worked through products of its extracellular metabolism. DCs express on
their surface both ecto-apyrase and ecto-5'-nucleotidase which rapidly
hydrolyze ATP to AMP and adenosine (16), and adenosine
receptors activation is reported to selectively inhibit IL-12 and
TNF- production by human monocytes (26, 34, 35). The
nonhydrolyzable ATP analogue, ATP--S, was more efficient than ATP at
inhibiting IL-12 and TNF- production, and antagonists of adenosine
receptors could not prevent ATP activity. Thus, extracellular ATP and
P2 purinoceptors appear to play a direct role in the modulation of DCs
response to both noncognate and cognate maturation stimuli. Given the
number of P2X and P2Y receptors subtypes expressed by DCs and the lack
of specific agonists or inhibitors, it is difficult to link receptors
subtypes to a given cellular response, with the exception of the
P2X7 receptor which was shown previously to
mediate membrane permeabilization, cell death, and IL-1 release at
concentrations much higher than those used in this study
(18). However, the inability of UTP and the similar
potency of ADP to reproduce ATP effects indicated that ATP could
function through the P2X receptor family.
Exposure of DCs to necrotic cells induces full maturation of DCs, including high membrane expression of costimulatory and MHC-presenting molecules and acquisition of potent capacity to stimulate effector T cell responses (4, 5), but the nature of the activating stimuli provided by necrotic cells are not known. We have shown here that ATP increases DC expression of CD83, CD86, and CD54, but not MHC molecules, and inhibits selectively the production of proinflammatory cytokines and IL-12. The effects of ATP on cytokine production are rather reminiscent of those observed in macrophages that have ingested apoptotic cells (36, 37). The intracellular concentration of ATP ranges from 5 to 10 mM, and any agent that causes plasma membrane damage or rapid cell death can promote the release of intracellular ATP (14). Hence, micromolar concentrations of ATP in the extracellular milieu are likely to be present at sites of inflammation associated with tissue damage. In this context, extracellular ATP may act as an important signal which can limit the initiation of Th1 response and favor Th2 immune deviation and prevent detrimental proinflammatory cytokine release by DCs. Moreover, the unchanged IL-10 production by maturing DCs exposed to ATP may allow the emergence of immune regulatory cells, such as T regulatory cells 1 (38, 39), an hypothesis currently tested in our laboratory. In conclusion, extracellular ATP may serve as an important signal for an alternative DC maturation (40) and thus for the prevention of exaggerated and harmful immune responses. This phenomenon may be especially relevant in the context of chronic inflammatory disorders during which ATP can be easily released by damaged cells and DCs can differentiate from monocyte precursors (41).
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Footnotes |
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2 A.L.S. and D.F. contributed equally to this work.
3 Address correspondence and reprint requests to Dr. Andrea la Sala, Laboratory of Immunology, Istituto Dermopatico dell’Immacolata, Via dei Monti di Creta, 104, 00167 Rome, Italy.
4 Abbreviations used in this paper: DC, dendritic cell; sCD40L, soluble CD40 ligand; IL-1ra, IL-1 receptor antagonist; DPCPX, 8-cyclopentyl-1–3-dipropylxanthine; CSC, 8-(3-chlorostyryl)caffeine; LDH, lactate dehydrogenase; Rp-cAMPS, Rp-adenosine 3',5'-cyclic monophosphothioate triethylamine.
5 S. Corinti, C. Albanesi, A. la Sala, S. Pastore, and G. Girolomoni. Regulatory activity of autocrine interleukin-10 on dendritic cell functions. Submitted for publication.
Received for publication September 25, 2000. Accepted for publication November 7, 2000.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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release by activating the purinergic P2Z receptor of human macrophages. J. Immunol. 159:1451.[Abstract]
1 and 2 chain expression. J. Exp. Med. 189:541.2-agonists prevent Th1 development by selective inhibition of IL-12. J. Clin. Invest. 100:1513.[Medline]
,25-Dihydroxyvitamin D3 inhibits differentiation, maturation, activation, and survival of dendritic cells leading to impaired alloreactive T cell activation. J. Immunol. 164:2405. expression by adenosine. J. Immunol. 156:3435.[Abstract]
, PGE2 and PAF. J. Clin. Invest. 101:890.[Medline]
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F. Marteau, D. Communi, J.-M. Boeynaems, and N. Suarez Gonzalez Involvement of multiple P2Y receptors and signaling pathways in the action of adenine nucleotides diphosphates on human monocyte-derived dendritic cells J. Leukoc. Biol., October 1, 2004; 76(4): 796 - 803. [Abstract] [Full Text] [PDF]
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M. Galgani, V. De Rosa, S. De Simone, A. Leonardi, U. D'Oro, G. Napolitani, A. M. Masci, S. Zappacosta, and L. Racioppi Cyclic AMP Modulates the Functional Plasticity of Immature Dendritic Cells by Inhibiting Src-like Kinases through Protein Kinase A-mediated Signaling J. Biol. Chem., July 30, 2004; 279(31): 32507 - 32514. [Abstract] [Full Text] [PDF]
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A. Mazzoni and D. M. Segal Controlling the Toll road to dendritic cell polarization J. Leukoc. Biol., May 1, 2004; 75(5): 721 - 730. [Abstract] [Full Text] [PDF]
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M. Schnurr, T. Toy, P. Stoitzner, P. Cameron, A. Shin, T. Beecroft, I. D. Davis, J. Cebon, and E. Maraskovsky ATP gradients inhibit the migratory capacity of specific human dendritic cell types: implications for P2Y11 receptor signaling Blood, July 15, 2003; 102(2): 613 - 620. [Abstract] [Full Text] [PDF]
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E. Panther, S. Corinti, M. Idzko, Y. Herouy, M. Napp, A. la Sala, G. Girolomoni, and J. Norgauer Adenosine affects expression of membrane molecules, cytokine and chemokine release, and the T-cell stimulatory capacity of human dendritic cells Blood, May 15, 2003; 101(10): 3985 - 3990. [Abstract] [Full Text] [PDF]
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K. Sak, J.-M. Boeynaems, and H. Everaus Involvement of P2Y receptors in the differentiation of haematopoietic cells J. Leukoc. Biol., April 1, 2003; 73(4): 442 - 447. [Abstract] [Full Text] [PDF]
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A. la Sala, D. Ferrari, F. Di Virgilio, M. Idzko, J. Norgauer, and G. Girolomoni Alerting and tuning the immune response by extracellular nucleotides J. Leukoc. Biol., March 1, 2003; 73(3): 339 - 343. [Abstract] [Full Text] [PDF]
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M. Idzko, S. Dichmann, D. Ferrari, F. Di Virgilio, A. la Sala, G. Girolomoni, E. Panther, and J. Norgauer Nucleotides induce chemotaxis and actin polymerization in immature but not mature human dendritic cells via activation of pertussis toxin-sensitive P2y receptors Blood, July 18, 2002; 100(3): 925 - 932. [Abstract] [Full Text] [PDF]
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A. R. Companjen, L. I. v. d. Wel, L. Boon, E. P. Prens, and J. D. Laman CD40 ligation-induced cytokine production in human skin explants is partly mediated via IL-1 Int. Immunol., June 1, 2002; 14(6): 669 - 676. [Abstract] [Full Text] [PDF]
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H. Ni, J. Capodici, G. Cannon, D. Communi, J.-M. Boeynaems, K. Kariko, and D. Weissman Extracellular mRNA Induces Dendritic Cell Activation by Stimulating Tumor Necrosis Factor-alpha Secretion and Signaling through a Nucleotide Receptor J. Biol. Chem., April 5, 2002; 277(15): 12689 - 12696. [Abstract] [Full Text] [PDF]
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A. la Sala, S. Sebastiani, D. Ferrari, F. Di Virgilio, M. Idzko, J. Norgauer, and G. Girolomoni Dendritic cells exposed to extracellular adenosine triphosphate acquire the migratory properties of mature cells and show a reduced capacity to attract type 1 T lymphocytes Blood, March 1, 2002; 99(5): 1715 - 1722. [Abstract] [Full Text] [PDF]
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G. Caron, Y. Delneste, E. Roelandts, C. Duez, J.-Y. Bonnefoy, J. Pestel, and P. Jeannin Histamine Polarizes Human Dendritic Cells into Th2 Cell-Promoting Effector Dendritic Cells J. Immunol., October 1, 2001; 167(7): 3682 - 3686. [Abstract] [Full Text] [PDF]
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), ATP and 10 µg/ml LPS (
), or ATP plus 1 µg/ml sCD40L
(
). DCs were then stained with mAbs against the indicated surface
markers. ATP induced higher expression of CD83, CD86, and CD54 in both
immature and LPS- or sCD40L-stimulated DCs, whereas CD80 was
up-regulated only in immature DCs. Similar results were observed in
three distinct experiments.
), 10 µg/ml LPS
(•), or LPS plus ATP (
), the
nonhydrolyzable ATP analogue ATP-