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Ludwig Boltzmann Institute for Cell Biology and Immunobiology of the Skin, Department of Dermatology, University of Munster, Munster, Germany
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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,
LPS, or CD40 ligand (CD40L)) were analyzed morphologically,
phenotypically, and functionally and were tested for their ability to
promote prophylactic and/or therapeutic antitumor immunity. Each of the
culture conditions generated typical BmDC. Whereas cells cultured in
GM-CSF alone were functionally immature, cells incubated with CD40L or
LPS were mature BmDC, as evident by morphology, capacity to internalize
Ag, migration into regional lymph nodes, IL-12 secretion, and
alloantigen or peptide Ag presentation in vitro. The remaining cultures
exhibited intermediate dendritic cell maturation. The in vivo
Ag-presenting capacity of BmDC was compared with respect to induction
of both protective tumor immunity and immunotherapy of established
tumors, using the poorly immunogenic squamous cell carcinoma, KLN205.
In correspondence to their maturation stage, BmDC cultured in the
presence of CD40L exhibited the most potent immunostimulatory effects.
In general, although not entirely, the capacity of BmDC to induce an
antitumor immune response in vivo correlated to their degree of
maturation. The present data support the clinical use of mature, rather
than immature, tumor Ag-pulsed dendritic cells as cancer vaccines and
identifies CD40L as a potent stimulus to enhance their in vivo
Ag-presenting capacity. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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, TGF-ß, PGE2, ionomycin,
and others (1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21). In vitro, and possibly in vivo as well,
inflammatory stimuli (such as IL-1, IL-6, and TNF) (21, 22, 23) and contact
with T cells (via CD40/CD40L interaction) (24, 25) further activate DC,
resulting in mature DC with strong T cell stimulatory potential.
Although several different protocols exist for generation of BmDC,
resulting in cells with potentially different phenotypes and/or
functions, the in vitro regulation of BmDC maturation by relevant
molecules in a comparative way and the correlation with their function
in vivo remain to be elucidated. Due to their unique capacity to stimulate resting T cells, DC are the candidate cell type to use for immunization protocols, especially for induction of antiviral or antitumor immunity (26, 27). It has been demonstrated that murine BmDC are able to promote prophylactic and therapeutic antitumor immunity when pulsed with relevant tumor-associated T cell epitopes (28, 29, 30, 31, 32). The degree of DC differentiation (immature vs mature) may determine their subsequent function. Once pulsed with Ag in vitro, it may be argued that immature BmDC are the most appropriate DC to use for immunization protocols in vivo because of their capacity to internalize/process Ag and mature on their way to the regional LN. Alternatively, however, mature DC may be best suited for in vivo immunotherapy because of their capacity to efficiently present Ag to naive T cells, and because immature BmDC may induce tolerance rather than immunity. Since Ag processing (which is maximal in immature DC) and T cell sensitization (which is more effective in mature DC) are both essential for the development of antitumor immunity, it is crucial to understand the regulation of DC maturation to gain further insights into their in vivo function. Presently, no data are available that conclusively address whether terminally differentiated mature DC or incompletely differentiated immature DC are best to use for vaccinations in vivo.
Therefore, to understand the interplay of cytokines in the DC maturation process, BmDC generated under various culture conditions were directly compared with respect to morphology, phenotype, and in vitro function. These different BmDC populations were then analyzed in vivo by testing the ability of these cells to migrate into regional LN and to promote prophylactic and/or therapeutic antitumor immunity to define culture conditions that result in the generation of DC that are best suited for in vivo immunotherapy.
We report here that BmDC generated only in the presence of GM-CSF
display an immature phenotype. CD40L and LPS are strong inducers of
full DC maturation. DC cultured in the presence of GM-CSF and IL-4 with
or without the addition of Flt3L or TNF- exhibit an intermediate
maturation stage with respect to phenotype and in vitro Ag-presenting
capacity. The cytokine secretion profile of BmDC cultured in various
ways differs significantly. The capacity to induce an antitumor immune
response in vivo correlates to the degree of DC maturation. BmDC
generated in vitro migrate rather inefficiently into regional LN after
s.c. injection regardless of the culture conditions and maturation
stage. Of all culture conditions tested, BmDC generated in the presence
of CD40L mediate the most potent immune responses in vivo, including
the generation of protective and therapeutic tumor immunity.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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DBA2/N, BALB/c, and C57BL/6 mice (6–10 wk old) were obtained from Charles River (Sulzfeld, Germany) and housed according to government regulations.
Generation, culture, and flow cytometry of BmDC
BmDC were generated as described previously (6) with some modifications. Briefly, bone marrow was collected from tibias and femurs of female BALB/c mice, passed through a nylon mesh to remove small pieces of bone and debris, resuspended in complete medium (CM: RPMI 1640 containing 5% FCS, 50 µM 2-ME, 2 mM glutamine, 0.1 mM nonessential amino acids, and 20 µg/ml gentamicin; all from PAA, Linz, Austria), and cultured in tissue culture dishes (Becton Dickinson, Heidelberg, Germany) for 2 h. Nonadherent cells were collected, and aliquots of 1 x 106 cells were placed in 24-well plates (Becton Dickinson) containing 1 ml of CM with the cytokines listed below. Two-thirds of the medium was replaced on day 3. On days 5 and 7 of culture nonadherent cells were transferred into six-well plates in CM with cytokines (2.5 x 106 cells/2 ml/well) and maintained for 2 additional days in culture. On day 9 of culture, most of the nonadherent cells had acquired typical dendritic morphology, and these cells were used as the source of DC in subsequent experiments. At this time point supernatants from the different cultures were collected and stored at -20°C for cytokine determinations.
Six different combinations of cytokines were used for BmDC generation.
1) GM-CSF (150 U/ml; R&D Systems, Wiesbaden, Germany); 2) GM-CSF and
IL-4 (75 U/ml; PharMingen, Hamburg, Germany); and 3) GM-CSF, IL-4, and
Flt3L (250 ng/ml; Immunex, Seattle, WA) were used throughout the
culture period. Alternatively, cells were maintained in GM-CSF and IL-4
for 9 days with the addition of 4) TNF- (20 U/ml; Genzyme,
Cambridge, MA), 5) CD40L (1 µg/ml; Immunex), or 6) Escherichia
coli-LPS (0.1 µg/ml; Sigma, St. Louis, MO) for the final 2
days of cell culture.
Expression of surface molecules was quantitated by flow cytometry using the following Abs: I-Ab,d,q, I-Ed,k (M5/114), B7-1 (1G10), B7-2 (Gl-1), CD11b/Mac-1 (M1/70), and CD40 (all obtained from PharMingen, San Diego, CA); CD11c (N418, Endogen, Cambridge, MA); NLDC145 and ICAM-1 (YN1/1.7.4; from American Type Culture Collection, Manassas, VA; 10% culture supernatant); anti-rat or anti-hamster IgG-FITC (Boehringer Mannheim, Mannheim, Germany); and normal rat IgG2b (PharMingen) as isotype control (1 µg/ml, diluted in PBS/1% FCS(v/v)). For flow cytometry, aliquots of 1 x 105 BmDC were incubated with the mAbs for 60 min at 4°C. The cells were washed twice with PBS/0.1% FCS(v/v) and incubated with FITC-conjugated goat anti-rat or goat anti-hamster IgG (diluted 1/50 in PBS/1% FCS (v/v)) for 45 min on ice. At the end of this incubation, propidium iodide (100 nM; Sigma) was added to determine the percentage of dead cells, cells were washed twice and subsequently analyzed in a flow cytometer (EPICS XL, Coulter, Miami, FL). No gating was performed, except for elimination of dead cells.
Detection of cytokines
IL-1ß, IL-10, IL-12, TNF-, and IFN- production by BmDC
was determined by ELISA (Endogen (Cambridge, MA) and Laboserv (Giessen,
Germany)). Additionally, IL-2 bioactivity in supernatants was assessed
in a bioassay based on the proliferation of the IL-2-dependent murine
cell line CTLL-2 (33). Briefly, CTLL-2 cells were washed extensively,
resuspended in CM, and seeded into 96-well, flat-bottom microtiter
plates (5 x 103 cells/well) in the presence of serial
dilutions of culture supernatants. In parallel, a standard curve was
generated using recombinant murine IL-2 (R&D Systems). After 48 h
of incubation, the proliferation of CTLL-2 cells was evaluated
photometrically using the Alamar-Blue reagent (BioSource, Camarillo,
CA).
Phagocytosis and endocytosis
Phagocytosis and endocytosis was assessed using FITC-conjugated E. coli particles and dextran, respectively (Molecular Probes, Leiden, The Netherlands). Briefly, 500 µl of E. coli particle suspension (0.625 mg/ml) or 20 µl of dextran (0.05 mM) was incubated with 5 x 105 BmDC, CTLL-2 (T cell line used as a negative control), and ML cells (murine macrophage cell line used as a positive control, provided by Dr. P. Ricciardi-Castagnoli) in RPMI 1640 medium at 37°C. After 2 h of incubation, cells were harvested and resuspended in medium with the addition of trypan blue, which quenches the fluorescence of extracellular particles. Next, BmDC were washed, resuspended in PBS, and analyzed in a flow cytometer.
Mixed lymphocyte reactions
BmDC were incubated in graded doses together with 2 x 105 allogeneic T cells in 96-well culture plates. The primary mixed lymphocyte reactions were performed in RPMI 1640 supplemented with 10% FCS, 100 U/ml penicillin, 100 mg/ml streptomycin, 0.1 mM nonessential amino acids, 2 mM L-glutamine, 1 mM sodium pyruvate, 0.01 M HEPES, 50 µM 2-ME, 20 µg/ml gentamicin, and 5 µg/ml indomethacin. T cells were obtained from spleen cells of C57BL/6 mice by nylon wool nonadherence and subsequent elimination of residual contaminating cells with Ab-coated T cell isolation columns (Cellect mouse T cell immunocolumns, Biotex, Alberta, Canada). The resulting cell preparation contained <0.1% IA+ cells. After 5 days, T cell proliferation was measured by adding [3H]thymidine to the cultures and subsequent liquid scintillation counting after an overnight incubation period.
Presentation of peptide Ag to Ag-specific T-T hybridomas in vitro
To assess the ability of BmDC to present soluble protein Ag or peptide to primed T cells, the OVA-specific T-T hybridoma D011.1 was used. This hybridoma recognizes the OVA323–339 peptide. For this assay, 1 x 104 BmDC/well or 1 x 105 A-20/well (B cell line, H-2d, used as positive control) were incubated in the presence of 0.08–5 µg of OVA323–339 peptide (recognized by D011.1 without requiring processing) (34). D011.1 cells (1 x 105) were added to each well, and the cultures were incubated in a total volume of 200 µl for 24 h at 37°C. One hundred microliters of culture supernatant was removed and assayed for IL-2 content using the IL-2-responsive cell line, CTLL-2, as described above.
In vivo migration
BmDC were labeled with the fluorescent dye, PKH2-2 (Sigma), according to the manufacturer’s protocol. Briefly, the cells were washed three times with PBS to remove FCS. Cells were resuspended in PKH2-2 staining solution for 5 min. CM containing 10% FCS was added to the cells, followed by removal of unbound PKH2-2 by extensive washing with PBS. Thereafter, 5 x 105 labeled cells were s.c. injected in 40 µl of PBS into hind footpads of mice. Forty-eight hours after injection, mice were killed, and regional LN (inguinal and popliteal LN) were removed. Single cell suspensions of LN cells were subjected to a density gradient (Nycoprep 1.077, Nycomed Pharma, Oslo, Norway), and a low density cell fraction, enriched in DC, was harvested after centrifugation at 900 x g for 30 min. Flow cytometric analysis was performed to detect fluorescent cells within the LN preparation.
Tumor cells and preparation of tumor Ag
The squamous cell carcinoma KLN205 (syngeneic to DBA2/N mice, H-2d) was obtained from American Type Culture Collection (Manassas, VA) and maintained in tissue culture at 37°C in RPMI 1640 containing 10% FCS, 100 U/ml penicillin, 100 mg/ml streptomycin, 0.1 mM nonessential amino acids, 2 mM L-glutamine, 1 mM sodium pyruvate, 0.01 M HEPES buffer, and 50 µM 2-ME. Upon injection of 2 x 105 tumor cells, the tumor grows progressively in syngeneic mice and can produce metastases at late tumor stages into regional LN. Previous experiments demonstrated that this tumor is very poorly immunogenic (K. Mahnke et al., manuscript in preparation). For preparation of TA, tumor fragments containing tumor-derived antigenic epitopes were prepared by disrupting 1 x 107 KLN205 cells/ml with four freeze-thaw cycles in liquid nitrogen. Lysates were centrifuged at 2000 x g for 10 min to remove insoluble cell fragments, and the supernatant was used as a source of tumor-associated Ag as previously described (35).
BmDC cultures were harvested and used for immunization experiments. To pulse BmDC with TA, 1 mg of TA from KLN205 cells was added to 1.5 x 106 BmDC in CM for 4 h at 37°C. Thereafter, cells were harvested and washed three times with PBS. Pulsed BmDC (2 x 104) were then injected s.c. into naive recipient mice on the lower abdomen. This immunization was repeated twice at weekly intervals. Two weeks before or 1 wk after the immunizations, mice were challenged with 2 x 105 live KLN205 cells s.c. on the lower lateral back, and tumor growth was assessed every 3 days by measurement with a spring-loaded caliper.
Statistical analysis
Student’s t test was applied to reveal significant differences in cytokine production by BmDC generated under different culture conditions. p < 0.05 was accepted as the level of significance.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The various culture conditions resulted in the generation of BmDC with typical morphology and fine membrane projections, although the number of dendritic processes was enhanced when CD40L or LPS were present in the culture medium and was decreased when cells were maintained in the presence of GM-CSF without addition of other factors. Homotypic BmDC aggregates were more abundant in cultures containing either CD40L or LPS, and cell yield was higher when GM-CSF or GM-CSF, IL-4, and Flt3L were present in the culture than with the remaining culture conditions.
Significant differences were observed in the morphology and expression
of cell surface molecules involved in APC/T cell interactions. Compared
with the other culture conditions, MHC class II, NLDC145, ICAM-1,
CD11c, CD40, B7-1, and B7-2 expression was consistently lower in cells
cultured in the presence of GM-CSF only or in cells cultured in GM-CSF,
IL-4, and Flt3L, whereas the highest surface expression of these
markers was observed in cells incubated with CD40L or LPS (Fig. 1
). To assess the percentage of
contaminating macrophages in the different culture conditions, the
expression of the macrophage-related surface molecules CD11b, F4/80,
ER-TR9, ER-HR3, and BM8 was determined. CD11b was broadly expressed on
BmDC, and all BmDC also expressed high amounts of ER-TR9, indicating
that these molecules are not macrophage specific. BmDC generated by
culture in GM-CSF alone also broadly expressed BM8, ER-HR3, and F4/80
(F4/80, 60%; ER-HR3, 35%; BM8, 80% positive cells), but the
expression of these markers could be down-regulated by subsequent
culture in LPS or CD40L, which is consistent with the immature
phenotype of these DC (data not shown). In all other BmDC culture
systems, the percentages of cells expressing significant levels of
ER-HR3, BM8, and F4/80 were <5%, indicating low numbers of
contaminating macrophages in the BmDC cultures.
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To investigate the modulation of cytokine production and secretion
profiles of BmDC emerging from the various culture conditions,
supernatants from BmDC cultures were collected and analyzed for
IL-1ß, IL-10, IL-12, TNF-, and IFN- contents using ELISA assays
(Table I).
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), whereas CD40L stimulation selectively induced the production
of IL-12 but not that of IL-1ß or TNF- by BmDC. Lower amounts of
IL-12, TNF-, and IL-1ß were measurable in supernatants of BmDC
generated under GM-CSF and IL-4 with or without the addition of TNF-
or Flt3L, and very low levels of these cytokines were detected in
supernatants from cells incubated with GM-CSF alone.
None of the tested stimuli induced endogenous production of IFN- by
BmDC, and only minute amounts of IL-10 were detected in supernatants
from LPS-cultured BmDC (mean ± SEM, 3.4 pg/ml ± 1.2), but
not in any of the other culture conditions.
Phagocytosis and endocytosis
The endocytic capacity of BmDC that emerged from the distinct
culture conditions is shown in Table II.
Incubation of BmDC with FITC-labeled E. coli provided
evidence of extensive phagocytosis in cells cultured in the presence of
GM-CSF, as judged by the appearance of many FITC-positive cells with
high mean fluorescence intensity. Incubation of cells with IL-4
resulted in a marked down-regulation of FITC-E. coli
uptake. Further addition of Flt3L, TNF-, CD40L, or LPS did not have
additional effects on phagocytosis. Similar results were obtained using
FITC-labeled latex beads instead of FITC-E. coli (data
not shown).
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and CD40L.
Significantly less efficient endocytosis was detectable in the rest of
the cultures. Thus, upon maturation, BmDC appear to shut off
phagocytosis but maintain a reduced, but still detectable, capacity for
endocytosis. Allostimulatory activity and presentation of OVA peptide
To further characterize BmDC generated under distinct culture
conditions, BmDC were compared for their capacities to stimulate
alloreactive T cells. Graded numbers of BmDC from each culture
condition were incubated with a fixed number of allogeneic T cells.
Fig. 2 shows that approximately 150 BmDC
could already trigger a substantial response in all the culture
conditions tested. However, at low stimulator cell concentrations, DC
cultured with either CD40L or LPS were at least twofold more effective
than the remaining cultures in stimulating naive allogeneic T cells. In
some experiments, as few as 30 BmDC/well from these cultures stimulated
allogeneic T cell responses of >100,000 cpm (data not shown). BmDC
generated from GM-CSF and IL-4 cultures with or without the addition of
Flt3L or TNF- exhibited intermediate ability to present alloantigen.
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, CD40L- and LPS-treated cells were the
most efficient APC also in this assay system, since they were able to
induce potent IL-2 production at low peptide concentrations. In
contrast, cells cultured in the presence of GM-CSF were less efficient
in OVA peptide presentation than BmDC grown under the other conditions.
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Syngeneic BmDC that had been labeled with a fluorescent dye were
s.c. injected into the hind footpad of mice. After 2 days, regional LN
were removed, and single cell suspensions were analyzed by flow
cytometry. In all experiments, dye-labeled BmDC were clearly detectable
in draining lymph nodes, but the vast majority of in vitro
differentiated BmDC injected s.c. into syngeneic mice remained at the
site of injection and failed to migrate into the draining LN regardless
of the culture conditions and degree of BmDC maturation (Table III). In general, approximately 3–10%
of the injected BmDC were detectable in regional LN, with a maximum
present 48 h after injection. The relatively poor migratory
capacity of s.c. injected BmDC was not due to interference of the
labeling procedure with BmDC viability or migration, since labeled and
unlabeled BmDC 1) did not differ in vitro with regard to survival or
allostimulatory capacity, 2) showed almost equal random migration
activity when placed into three-dimensional collagen gels and subjected
to time-lapse video microscopy, 3) were detectable in equally large
numbers (>75% of the injected cells) as viable (trypan
blue-excluding) cells in the skin at the injection site, and 4) were
detectable in the regional LN in equally low numbers when investigated
by direct fluorescence microscopy or after labeling by phagocytosis of
fluorescent E. coli instead of using the membrane dye
(data not shown). Thus, only a small percentage of BmDC migrates into
the regional LN after s.c. injection in mice.
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To assess which type of DC culture and which DC differentiation
stage are best suited for in vivo immunotherapy, we tested the ability
of these BmDC to induce protective and therapeutic immunity against the
poorly immunogenic squamous cell carcinoma, KLN205. For immunization
against the tumor, mice were injected twice at weekly intervals with
TA-pulsed BmDC as described in Materials and Methods.
Control groups received TA only (data not shown) or BmDC that had not
been pulsed with TA. One week after the last immunization, mice were
challenged by s.c. injection of viable KLN205 cells. Control groups
were not protected against tumor growth, suggesting that s.c. injected
DC do not induce tumor immunity by Ag-independent mechanisms, and that
immunization with TA alone does not induce a tumor-specific immune
response. In contrast to data obtained by others using different tumor
models, in the KLN205 tumor model immunization with TA-exposed BmDC
that were generated by culturing cells in GM-CSF and IL-4 did not lead
to consistent and profound protective tumor immunity, possibly due to
the low immunogenicity of this tumor cell line. However, immunization
of mice with TA-pulsed BmDC generated in vitro in the presence of CD40L
dramatically reduced tumor growth and incidence, whereas mice immunized
with TA-pulsed BmDC that were cultured with GM-CSF alone did not
exhibit any protective tumor immunity (Fig. 4).
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a), whereas LPS-stimulated
BmDC were not as potent as CD40L-treated cells when used in this
immunotherapy model (Fig. 5b).
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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CD40L was found to be a strong inducer of full DC maturation, as
evident by down-regulation of phagocytosis, low endocytic capacity, and
up-regulation of MHC class II, the adhesion molecules CD11c and ICAM-1,
and the costimulatory molecules B7-1 and B7-2. In addition, cells
incubated with CD40L secreted high levels of IL-12 and were very
efficient in presenting alloantigen or peptide Ag in vitro. In vivo,
these mature DC were excellent in the promotion of antitumor immunity.
Thus, the ability of the CD40L differentiated cells to promote
antitumor immunity correlates to their high efficiency of stimulating
resting T cells and to the high production of IL-12, which is in
accordance with in vitro data obtained by others (24, 25). At present,
we do not know yet whether these features are responsible for their
potent in vivo immunostimulatory capacity. We speculate, however, that
the IL-12 production by DC is critical for their in vivo function,
since in other systems, IL-12 was shown to generate a polarization of
the immune response toward the Th1 pathway in vivo. IL-12 is also a
potent inducer of IFN- and TNF- production by both NK cells and T
cells (37), and these cytokines are critically involved in the
development of cell-mediated immune responses (38), which is crucial
for the induction of antitumor immunity. In this direction, Zitvogel et
al. showed that the neutralization of both IFN- and TNF- or the
administration of anti-IL-12 Abs in mice totally abrogates the
DC-induced anti-MCA205 tumor response (30). At the same time,
however, LPS-stimulated DC were less capable of promoting tumor
immunity than CD40L-treated BmDC, although they secreted similar
amounts of IL-12 in vitro and had an in vitro maturation stage
comparable to that of CD40L-cultured BmDC. Besides the functional
characteristics previously described for the CD40L-generated cells, the
cells cultured in the presence of LPS produced, in addition to IL-12,
high levels of TNF- and IL-1ß. Although a role for these cytokines
in tumor resistance has also been suggested (11, 39), the different
cytokine secretion profiles of LPS- vs CD40L-stimulated BmDC may
contribute to their different in vivo effectiveness.
In contrast to other laboratories (8), in our hands BmDC differentiated in the presence of only GM-CSF consistently generated immature DC as evident by morphological, phenotypical, and functional analyses in vitro as well as in vivo. Most likely, this difference is due to the use of different protocols for DC generation. Clearly, our data also show that IL-4 is a potent enhancer of mouse DC maturation, which is in agreement with data obtained by others (21, 29, 30, 32). Compared with DC differentiated in the presence of GM-CSF alone, supplementation of IL-4 significantly enhanced DC differentiation, leading to an intermediate degree of maturation. Expression of MHC class II Ags and the costimulatory molecules B7-1 and B7-2 was higher on DC cultured with GM-CSF and IL-4 than on those cultured with GM-CSF alone. Furthermore, DC grown in GM-CSF plus IL-4 were more potent stimulators of mixed lymphocyte reactions as well as more efficient in Ag presentation than cells grown in medium containing GM-CSF alone. Thus, and in agreement with other publications investigating human DC (9, 24, 25), this combination of cytokines provided good conditions for the generation of cells with the characteristic phenotype and functional properties of DC in the murine system. However, these in vitro generated BmDC retarded tumor growth only to some extent when DC were injected before tumor challenge and were poorly able to induce the rejection of preexisting tumors. These data contrast with the results of studies performed by Mayordomo et al. (32), which indicate that BmDC grown in medium containing GM-CSF and IL-4 are capable of generating a complete protective antitumor immune response. It is unlikely that this discrepancy is due to the inadequate quality of the BmDC generated in our study, given their immunophenotype and functional properties in vitro. Instead, it may be attributed to the use of different tumor models. The KLN205 is a very poorly immunogenic tumor, as determined by the fact that it generates no concomitant immunity and that vaccination with dead tumor cells alone or together with CFA, with irradiated cells, or with soluble TA did not affect tumor growth after a subsequent challenge with viable KLN205 cells (data not shown).
Surprisingly, the addition of TNF- or Flt3L to the GM-CSF plus IL-4
cultures did not substantially affect the morphology or the in vitro
functions of these cells. However, in vivo tumor experiments showed the
ability of these cells to prevent tumor growth in some of the injected
animals when tumor cells were injected after DC vaccination, whereas
GM-CSF and IL-4 differentiated cells were able to retard tumor growth
but not to prevent it. Interestingly, however, BmDC generated in the
presence of Flt3L decreased tumor growth in a similar way as that
observed by the CD40L differentiated BmDC, when tumor challenge
preceded BmDC vaccination. Despite the functional similarities found in
these cultures in vitro (GM-CSF and IL-4; GM-CSF, IL-4, and Flt3L; and
GM-CSF, IL-4, and TNF-), different antitumor responses were
determined in vivo. The mechanisms underlying these differences remain
to be elucidated.
The capacity of DC to migrate into T cell areas of LN is a key event in initiating immunity, and it may be critical for sensitization against tumor Ags. Thus, we were surprised by our consistent finding that BmDC appear to migrate very inefficiently into regional LN after s.c. injection, at least in the murine system. However, these data are in agreement with those of other groups (40, 41, 42), who also reported that the majority of DC that were s.c. injected remained at the site of injection and failed to migrate to the LN. In contrast, in vitro generated DC injected into the s.c. tissue of chimpanzees were reported to migrate rapidly and apparently completely to the regional LN (43). This different migratory behavior of BmDC may be due to different properties of chimpanzee and murine BmDC, respectively, or may reflect the fact that large numbers of DC (4 x 106) were injected directly adjacent to the inguinal LN in that study. We believe that the poor migratory capacity of s.c. injected BmDC was not a technical artifact, since labeled and unlabeled BmDC exhibited equal in vitro survival, random migration in collagen gels, and allostimulatory capacity, and identical results were obtained when using an entirely different labeling technique. Thus, s.c. injection may not be an optimal cell delivery system for in vitro generated BmDC, at least in the mouse. Nevertheless, although only a small number of labeled BmDC was detected in the LN, a potent in vivo immunostimulatory capacity was observed in most cases, resulting in the generation of protective and therapeutic tumor immunity. In our hands, incubation of BmDC with CD40L is currently the most potent stimulus to generate efficient in vivo immunostimulatory activity of murine BmDC. Thus, it might be crucial to optimize the maturation stage and migratory capacities of DC for designing future strategies using TA-pulsed DC for tumor immunotherapy.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. S. Grabbe, Department of Dermatology, University of Munster, Von-Esmarch Str. 56, D-48149 Munster, Germany. E-mail address:
3 Abbreviations used in this paper: DC, dendritic cell(s); Flt3L, Flt3 ligand; GM-CSF, granulocyte-macrophage colony-stimulating factor; CD40L, CD40 ligand; BmDC, bone marrow-derived dendritic cells; LN, lymph node; CM, complete medium; TA, tumor antigen.
Received for publication April 20, 1998. Accepted for publication September 10, 1998.
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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]
. J. Exp. Med. 184:695. and is inhibited by TGF-ß. Eur. J. Immunol. 24:793.[Medline]
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U. Sriram, C. Biswas, E. M. Behrens, J.-A. Dinnall, D. K. Shivers, M. Monestier, Y. Argon, and S. Gallucci IL-4 Suppresses Dendritic Cell Response to Type I Interferons J. Immunol., November 15, 2007; 179(10): 6446 - 6455. [Abstract] [Full Text] [PDF]
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P. Reichardt, B. Dornbach, S. Rong, S. Beissert, F. Gueler, K. Loser, and M. Gunzer Naive B cells generate regulatory T cells in the presence of a mature immunologic synapse Blood, September 1, 2007; 110(5): 1519 - 1529. [Abstract] [Full Text] [PDF]
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 A. P. Phadke, G. Akangire, S. J. Park, S. A. Lira, and B. Mehrad The Role of CC Chemokine Receptor 6 in Host Defense in a Model of Invasive Pulmonary Aspergillosis Am. J. Respir. Crit. Care Med., June 1, 2007; 175(11): 1165 - 1172. [Abstract] [Full Text] [PDF]
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G. Varga, S. Balkow, M. K. Wild, A. Stadtbaeumer, M. Krummen, T. Rothoeft, T. Higuchi, S. Beissert, K. Wethmar, K. Scharffetter-Kochanek, et al. Active MAC-1 (CD11b/CD18) on DCs inhibits full T-cell activation Blood, January 15, 2007; 109(2): 661 - 669. [Abstract] [Full Text] [PDF]
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M. Udagawa, C. Kudo-Saito, G. Hasegawa, K. Yano, A. Yamamoto, M. Yaguchi, M. Toda, I. Azuma, T. Iwai, and Y. Kawakami Enhancement of Immunologic Tumor Regression by Intratumoral Administration of Dendritic Cells in Combination with Cryoablative Tumor Pretreatment and Bacillus Calmette-Guerin Cell Wall Skeleton Stimulation Clin. Cancer Res., December 15, 2006; 12(24): 7465 - 7475. [Abstract] [Full Text] [PDF]
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M.-G. de Goer de Herve, D. Durali, T.-A. Tran, G. Maigne, F. Simonetta, P. Leclerc, J.-F. Delfraissy, and Y. Taoufik Differential effect of agonistic anti-CD40 on human mature and immature dendritic cells: the Janus face of anti-CD40 Blood, October 15, 2005; 106(8): 2806 - 2814. [Abstract] [Full Text] [PDF]
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H. K. Kang, H.-Y. Lee, M.-K. Kim, K. S. Park, Y. M. Park, J.-Y. Kwak, and Y.-S. Bae The Synthetic Peptide Trp-Lys-Tyr-Met-Val-D-Met Inhibits Human Monocyte-Derived Dendritic Cell Maturation via Formyl Peptide Receptor and Formyl Peptide Receptor-Like 2 J. Immunol., July 15, 2005; 175(2): 685 - 692. [Abstract] [Full Text] [PDF]
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R. Yamanaka, J. Homma, N. Yajima, N. Tsuchiya, M. Sano, T. Kobayashi, S. Yoshida, T. Abe, M. Narita, M. Takahashi, et al. Clinical Evaluation of Dendritic Cell Vaccination for Patients with Recurrent Glioma: Results of a Clinical Phase I/II Trial Clin. Cancer Res., June 1, 2005; 11(11): 4160 - 4167. [Abstract] [Full Text] [PDF]
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T. Ito, C. Nishiyama, M. Nishiyama, H. Matsuda, K. Maeda, Y. Akizawa, R. Tsuboi, K. Okumura, and H. Ogawa Mast Cells Acquire Monocyte-Specific Gene Expression and Monocyte-Like Morphology by Overproduction of PU.1 J. Immunol., January 1, 2005; 174(1): 376 - 383. [Abstract] [Full Text] [PDF]
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G. J. D. van Mierlo, Z. F. H. M. Boonman, H. M. H. Dumortier, A. Th. den Boer, M. F. Fransen, J. Nouta, E. I. H. van der Voort, R. Offringa, R. E. M. Toes, and C. J. M. Melief Activation of Dendritic Cells That Cross-Present Tumor-Derived Antigen Licenses CD8+ CTL to Cause Tumor Eradication J. Immunol., December 1, 2004; 173(11): 6753 - 6759. [Abstract] [Full Text] [PDF]
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P. A. Stumbles, R. Himbeck, J. A. Frelinger, E. J. Collins, R. A. Lake, and B. W. S. Robinson Cutting Edge: Tumor-Specific CTL Are Constitutively Cross-Armed in Draining Lymph Nodes and Transiently Disseminate to Mediate Tumor Regression following Systemic CD40 Activation J. Immunol., November 15, 2004; 173(10): 5923 - 5928. [Abstract] [Full Text] [PDF]
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M. Gunzer, C. Weishaupt, A. Hillmer, Y. Basoglu, P. Friedl, K. E. Dittmar, W. Kolanus, G. Varga, and S. Grabbe A spectrum of biophysical interaction modes between T cells and different antigen-presenting cells during priming in 3-D collagen and in vivo Blood, November 1, 2004; 104(9): 2801 - 2809. [Abstract] [Full Text] [PDF]
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S.-i. Fujii, K. Liu, C. Smith, A. J. Bonito, and R. M. Steinman The Linkage of Innate to Adaptive Immunity via Maturing Dendritic Cells In Vivo Requires CD40 Ligation in Addition to Antigen Presentation and CD80/86 Costimulation J. Exp. Med., June 21, 2004; 199(12): 1607 - 1618. [Abstract] [Full Text] [PDF]
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R. M. Cisco, Z. Abdel-Wahab, J. Dannull, S. Nair, D. S. Tyler, E. Gilboa, J. Vieweg, Y. Daaka, and S. K. Pruitt Induction of Human Dendritic Cell Maturation Using Transfection with RNA Encoding a Dominant Positive Toll-Like Receptor 4 J. Immunol., June 1, 2004; 172(11): 7162 - 7168. [Abstract] [Full Text] [PDF]
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M. Terme, E. Tomasello, K. Maruyama, F. Crepineau, N. Chaput, C. Flament, J.-P. Marolleau, E. Angevin, E. F. Wagner, B. Salomon, et al. IL-4 Confers NK Stimulatory Capacity to Murine Dendritic Cells: A Signaling Pathway Involving KARAP/DAP12-Triggering Receptor Expressed on Myeloid Cell 2 Molecules J. Immunol., May 15, 2004; 172(10): 5957 - 5966. [Abstract] [Full Text] [PDF]
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J. R. Ramirez-Pineda, A. Frohlich, C. Berberich, and H. Moll Dendritic Cells (DC) Activated by CpG DNA Ex Vivo Are Potent Inducers of Host Resistance to an Intracellular Pathogen That Is Independent of IL-12 Derived from the Immunizing DC J. Immunol., May 15, 2004; 172(10): 6281 - 6289. [Abstract] [Full Text] [PDF]
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S. Nair, C. McLaughlin, A. Weizer, Z. Su, D. Boczkowski, J. Dannull, J. Vieweg, and E. Gilboa Injection of Immature Dendritic Cells into Adjuvant-Treated Skin Obviates the Need for Ex Vivo Maturation J. Immunol., December 1, 2003; 171(11): 6275 - 6282. [Abstract] [Full Text] [PDF]
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S. Xu, G. K. Koski, M. Faries, I. Bedrosian, R. Mick, M. Maeurer, M. A. Cheever, P. A. Cohen, and B. J. Czerniecki Rapid High Efficiency Sensitization of CD8+ T Cells to Tumor Antigens by Dendritic Cells Leads to Enhanced Functional Avidity and Direct Tumor Recognition Through an IL-12-Dependent Mechanism J. Immunol., September 1, 2003; 171(5): 2251 - 2261. [Abstract] [Full Text] [PDF]
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C. L. Doxsee, T. R. Riter, M. J. Reiter, S. J. Gibson, J. P. Vasilakos, and R. M. Kedl The Immune Response Modifier and Toll-Like Receptor 7 Agonist S-27609 Selectively Induces IL-12 and TNF-{alpha} Production in CD11c+CD11b+CD8- Dendritic Cells J. Immunol., August 1, 2003; 171(3): 1156 - 1163. [Abstract] [Full Text] [PDF]
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N. Setterblad, C. Roucard, C. Bocaccio, J.-P. Abastado, D. Charron, and N. Mooney Composition of MHC class II-enriched lipid microdomains is modified during maturation of primary dendritic cells J. Leukoc. Biol., July 1, 2003; 74(1): 40 - 48. [Abstract] [Full Text] [PDF]
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M. Ghosh, C. Pal, M. Ray, S. Maitra, L. Mandal, and S. Bandyopadhyay Dendritic Cell-Based Immunotherapy Combined with Antimony-Based Chemotherapy Cures Established Murine Visceral Leishmaniasis J. Immunol., June 1, 2003; 170(11): 5625 - 5629. [Abstract] [Full Text] [PDF]
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S. Flynn and B. Stockinger Tumor and CD4 T-cell interactions: tumor escape as result of reciprocal inactivation Blood, June 1, 2003; 101(11): 4472 - 4478. [Abstract] [Full Text] [PDF]
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A. S. Yang and E. C. Lattime Tumor-induced Interleukin 10 Suppresses the Ability of Splenic Dendritic Cells to Stimulate CD4 and CD8 T-Cell Responses Cancer Res., May 1, 2003; 63(9): 2150 - 2157. [Abstract] [Full Text] [PDF]
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J. J. Kobie, R. S. Wu, R. A. Kurt, S. Lou, M. K. Adelman, L. J. Whitesell, L. V. Ramanathapuram, C. L. Arteaga, and E. T. Akporiaye Transforming Growth Factor {beta} Inhibits the Antigen-Presenting Functions and Antitumor Activity of Dendritic Cell Vaccines Cancer Res., April 15, 2003; 63(8): 1860 - 1864. [Abstract] [Full Text] [PDF]
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A. Soruri, Z. Kiafard, C. Dettmer, J. Riggert, J. Kohl, and J. Zwirner IL-4 Down-Regulates Anaphylatoxin Receptors in Monocytes and Dendritic Cells and Impairs Anaphylatoxin-Induced Migration In Vivo J. Immunol., March 15, 2003; 170(6): 3306 - 3314. [Abstract] [Full Text] [PDF]
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W. Hou, Y. Wu, S. Sun, M. Shi, Y. Sun, C. Yang, G. Pei, Y. Gu, C. Zhong, and B. Sun Pertussis Toxin Enhances Th1 Responses by Stimulation of Dendritic Cells J. Immunol., February 15, 2003; 170(4): 1728 - 1736. [Abstract] [Full Text] [PDF]
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K. Abe, F. O. Yarovinsky, T. Murakami, A. N. Shakhov, A. V. Tumanov, D. Ito, L. N. Drutskaya, K. Pfeffer, D. V. Kuprash, K. L. Komschlies, et al. Distinct contributions of TNF and LT cytokines to the development of dendritic cells in vitro and their recruitment in vivo Blood, February 15, 2003; 101(4): 1477 - 1483. [Abstract] [Full Text] [PDF]
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B. J. Weigel, N. Nath, P. A. Taylor, A. Panoskaltsis-Mortari, W. Chen, A. M. Krieg, K. Brasel, and B. R. Blazar Comparative analysis of murine marrow-derived dendritic cells generated by Flt3L or GM-CSF/IL-4 and matured with immune stimulatory agents on the in vivo induction of antileukemia responses Blood, December 1, 2002; 100(12): 4169 - 4176. [Abstract] [Full Text] [PDF]
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H.-R. Jiang, E. Muckersie, M. Robertson, H. Xu, J. Liversidge, and J. V. Forrester Secretion of interleukin-10 or interleukin-12 by LPS-activated dendritic cells is critically dependent on time of stimulus relative to initiation of purified DC culture J. Leukoc. Biol., November 1, 2002; 72(5): 978 - 985. [Abstract] [Full Text] [PDF]
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T. D. de Gruijl, S. A. Luykx-de Bakker, B. W. Tillman, A. J. M. van den Eertwegh, J. Buter, S. M. Lougheed, G. J. van der Bij, A. M. Safer, H. J. Haisma, D. T. Curiel, et al. Prolonged Maturation and Enhanced Transduction of Dendritic Cells Migrated from Human Skin Explants After In Situ Delivery of CD40-Targeted Adenoviral Vectors J. Immunol., November 1, 2002; 169(9): 5322 - 5331. [Abstract] [Full Text] [PDF]
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J. M. Lee, A. Mahtabifard, R. Yamada, R. G. Crystal, and R. J. Korst Adenovirus Vector-mediated Overexpression of a Truncated Form of the p65 Nuclear Factor {kappa}B cDNA in Dendritic Cells Enhances Their Function Resulting in Immune-mediated Suppression of Preexisting Murine Tumors Clin. Cancer Res., November 1, 2002; 8(11): 3561 - 3569. [Abstract] [Full Text] [PDF]
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J. R. Arron, Y. Pewzner-Jung, M. C. Walsh, T. Kobayashi, and Y. Choi Regulation of the Subcellular Localization of Tumor Necrosis Factor Receptor-associated Factor (TRAF)2 by TRAF1 Reveals Mechanisms of TRAF2 Signaling J. Exp. Med., October 7, 2002; 196(7): 923 - 934. [Abstract] [Full Text] [PDF]
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M. B. Lutz, M. Schnare, M. Menges, S. Rossner, M. Rollinghoff, G. Schuler, and A. Gessner Differential Functions of IL-4 Receptor Types I and II for Dendritic Cell Maturation and IL-12 Production and Their Dependency on GM-CSF J. Immunol., October 1, 2002; 169(7): 3574 - 3580. [Abstract] [Full Text] [PDF]
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G. Miller, V. G. Pillarisetty, A. B. Shah, S. Lahrs, Z. Xing, and R. P. DeMatteo Endogenous Granulocyte-Macrophage Colony-Stimulating Factor Overexpression In Vivo Results in the Long-Term Recruitment of a Distinct Dendritic Cell Population with Enhanced Immunostimulatory Function J. Immunol., September 15, 2002; 169(6): 2875 - 2885. [Abstract] [Full Text] [PDF]
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B. J. O'Sullivan and R. Thomas CD40 Ligation Conditions Dendritic Cell Antigen-Presenting Function Through Sustained Activation of NF-{kappa}B J. Immunol., June 1, 2002; 168(11): 5491 - 5498. [Abstract] [Full Text] [PDF]
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R. J. Steptoe, J. M. Ritchie, and L. C. Harrison Increased Generation of Dendritic Cells from Myeloid Progenitors in Autoimmune-Prone Nonobese Diabetic Mice J. Immunol., May 15, 2002; 168(10): 5032 - 5041. [Abstract] [Full Text] [PDF]
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S. K. Basak, A. Harui, M. Stolina, S. Sharma, K. Mitani, S. M. Dubinett, and M. D. Roth Increased dendritic cell number and function following continuous in vivo infusion of granulocyte macrophage-colony-stimulating factor and interleukin-4 Blood, April 15, 2002; 99(8): 2869 - 2879. [Abstract] [Full Text] [PDF]
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G. G. Pendl, C. Robert, M. Steinert, R. Thanos, R. Eytner, E. Borges, M. K. Wild, J. B. Lowe, R. C. Fuhlbrigge, T. S. Kupper, et al. Immature mouse dendritic cells enter inflamed tissue, a process that requires E- and P-selectin, but not P-selectin glycoprotein ligand 1 Blood, February 1, 2002; 99(3): 946 - 956. [Abstract] [Full Text] [PDF]
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T. Schuler, T. Kammertoens, S. Preiss, P. Debs, N. Noben-Trauth, and T. Blankenstein Generation of Tumor-associated Cytotoxic T Lymphocytes Requires Interleukin 4 from CD8+ T Cells J. Exp. Med., December 17, 2001; 194(12): 1767 - 1775. [Abstract] [Full Text] [PDF]
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K. Kato, Y. Takaue, and H. Wakasugi T-cell-conditioned medium efficiently induces the maturation and function of human dendritic cells J. Leukoc. Biol., December 1, 2001; 70(6): 941 - 949. [Abstract] [Full Text] [PDF]
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C. J. Kirk, D. Hartigan-O'Connor, and J. J. Mule The Dynamics of the T-Cell Antitumor Response: Chemokine-secreting Dendritic Cells Can Prime Tumor-reactive T Cells Extranodally Cancer Res., December 1, 2001; 61(24): 8794 - 8802. [Abstract] [Full Text] [PDF]
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J. Li, B. Schuler-Thurner, G. Schuler, C. Huber, and B. Seliger Bipartite regulation of different components of the MHC class I antigen-processing machinery during dendritic cell maturation Int. Immunol., December 1, 2001; 13(12): 1515 - 1523. [Abstract] [Full Text] [PDF]
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N. Okada, T. Saito, Y. Masunaga, Y. Tsukada, S. Nakagawa, H. Mizuguchi, K. Mori, Y. Okada, T. Fujita, T. Hayakawa, et al. Efficient Antigen Gene Transduction Using Arg-Gly-Asp Fiber-Mutant Adenovirus Vectors Can Potentiate Antitumor Vaccine Efficacy and Maturation of Murine Dendritic Cells Cancer Res., November 1, 2001; 61(21): 7913 - 7919. [Abstract] [Full Text] [PDF]
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Y. Kotera, K. Shimizu, and J. J. Mule Comparative Analysis of Necrotic and Apoptotic Tumor Cells As a Source of Antigen(s) in Dendritic Cell-based Immunization Cancer Res., November 1, 2001; 61(22): 8105 - 8109. [Abstract] [Full Text] [PDF]
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A. Mehling, K. Loser, G. Varga, D. Metze, T. A. Luger, T. Schwarz, S. Grabbe, and S. Beissert Overexpression of Cd40 Ligand in Murine Epidermis Results in Chronic Skin Inflammation and Systemic Autoimmunity J. Exp. Med., September 3, 2001; 194(5): 615 - 628. [Abstract] [Full Text] [PDF]
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D. Stober, R. Schirmbeck, and J. Reimann IL-12/IL-18-Dependent IFN-{{gamma}} Release by Murine Dendritic Cells J. Immunol., July 15, 2001; 167(2): 957 - 965. [Abstract] [Full Text] [PDF]
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J. D. Ahlers, I. M. Belyakov, S. Matsui, and J. A. Berzofsky Mechanisms of cytokine synergy essential for vaccine protection against viral challenge Int. Immunol., July 1, 2001; 13(7): 897 - 908. [Abstract] [Full Text] [PDF]
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S. Auffermann-Gretzinger, E. B. Keeffe, and S. Levy Impaired dendritic cell maturation in patients with chronic, but not resolved, hepatitis C virus infection Blood, May 15, 2001; 97(10): 3171 - 3176. [Abstract] [Full Text] [PDF]
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S. Liu, Y. Yu, M. Zhang, W. Wang, and X. Cao The Involvement of TNF-{{alpha}}-Related Apoptosis-Inducing Ligand in the Enhanced Cytotoxicity of IFN-{{beta}}-Stimulated Human Dendritic Cells to Tumor Cells J. Immunol., May 1, 2001; 166(9): 5407 - 5415. [Abstract] [Full Text] [PDF]
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K. A. Candido, K. Shimizu, J. C. McLaughlin, R. Kunkel, J. A. Fuller, B. G. Redman, E. K. Thomas, B. J. Nickoloff, and J. J. Mulé Local Administration of Dendritic Cells Inhibits Established Breast Tumor Growth: Implications for Apoptosis-inducing Agents Cancer Res., January 1, 2001; 61(1): 228 - 236. [Abstract] [Full Text]
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L. A. Lambert, G. R. Gibson, M. Maloney, B. Durell, R. J. Noelle, and R. J. Barth Jr. Intranodal Immunization with Tumor Lysate-pulsed Dendritic Cells Enhances Protective Antitumor Immunity Cancer Res., January 1, 2001; 61(2): 641 - 646. [Abstract] [Full Text]
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C. Brunner, J. Seiderer, A. Schlamp, M. Bidlingmaier, A. Eigler, W. Haimerl, H.-A. Lehr, A. M. Krieg, G. Hartmann, and S. Endres Enhanced Dendritic Cell Maturation by TNF-{alpha} or Cytidine-Phosphate-Guanosine DNA Drives T Cell Activation In Vitro and Therapeutic Anti-Tumor Immune Responses In Vivo J. Immunol., December 1, 2000; 165(11): 6278 - 6286. [Abstract] [Full Text] [PDF]
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S. Koido, M. Kashiwaba, D. Chen, S. Gendler, D. Kufe, and J. Gong Induction of Antitumor Immunity by Vaccination of Dendritic Cells Transfected with MUC1 RNA J. Immunol., November 15, 2000; 165(10): 5713 - 5719. [Abstract] [Full Text] [PDF]
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K. Brasel, T. De Smedt, J. L. Smith, and C. R. Maliszewski Generation of murine dendritic cells from flt3-ligand-supplemented bone marrow cultures Blood, November 1, 2000; 96(9): 3029 - 3039. [Abstract] [Full Text] [PDF]
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D. Haddad, J. Ramprakash, M. Sedegah, Y. Charoenvit, R. Baumgartner, S. Kumar, S. L. Hoffman, and W. R. Weiss Plasmid Vaccine Expressing Granulocyte-Macrophage Colony-Stimulating Factor Attracts Infiltrates Including Immature Dendritic Cells into Injected Muscles J. Immunol., October 1, 2000; 165(7): 3772 - 3781. [Abstract] [Full Text] [PDF]
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I. E. A. Flesch, D. Stober, R. Schirmbeck, and J. Reimann Monocyte inflammatory protein-1{alpha} facilitates priming of CD8+ T cell responses to exogenous viral antigen Int. Immunol., September 1, 2000; 12(9): 1365 - 1370. [Abstract] [Full Text] [PDF]
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A. Mehling, S. Grabbe, M. Voskort, T. Schwarz, T. A. Luger, and S. Beissert Mycophenolate Mofetil Impairs the Maturation and Function of Murine Dendritic Cells J. Immunol., September 1, 2000; 165(5): 2374 - 2381. [Abstract] [Full Text] [PDF]
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M. Bellone, D. Cantarella, P. Castiglioni, M. C. Crosti, A. Ronchetti, M. Moro, M. P. Garancini, G. Casorati, and P. Dellabona Relevance of the Tumor Antigen in the Validation of Three Vaccination Strategies for Melanoma J. Immunol., September 1, 2000; 165(5): 2651 - 2656. [Abstract] [Full Text] [PDF]
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 J. W. Hodge, A. N. Rad, D. W. Grosenbach, H. Sabzevari, A. G. Yafal, L. Gritz, and J. Schlom Enhanced Activation of T Cells by Dendritic Cells Engineered to Hyperexpress a Triad of Costimulatory Molecules J Natl Cancer Inst, August 2, 2000; 92(15): 1228 - 1239. [Abstract] [Full Text] [PDF]
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S. Vendetti, J.-G. Chai, J. Dyson, E. Simpson, G. Lombardi, and R. Lechler Anergic T Cells Inhibit the Antigen-Presenting Function of Dendritic Cells J. Immunol., August 1, 2000; 165(3): 1175 - 1181. [Abstract] [Full Text] [PDF]
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D. H. Schuurhuis, S. Laban, R. E.M. Toes, P. Ricciardi-Castagnoli, M. J. Kleijmeer, E. I.H. van der Voort, D. Rea, R. Offringa, H. J. Geuze, C. J.M. Melief, et al. Immature Dendritic Cells Acquire Cd8+Cytotoxic T Lymphocyte Priming Capacity upon Activation by T Helper Cell-Independent or-Dependent Stimuli J. Exp. Med., July 3, 2000; 192(1): 145 - 150. [Abstract] [Full Text] [PDF]
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T. K. Hoffmann, N. Meidenbauer, G. Dworacki, H. Kanaya, and T. L. Whiteside Generation of Tumor-specific T Lymphocytes by Cross-Priming with Human Dendritic Cells Ingesting Apoptotic Tumor Cells Cancer Res., July 1, 2000; 60(13): 3542 - 3549. [Abstract] [Full Text]
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T. Kikuchi, M. A. S. Moore, and R. G. Crystal Dendritic cells modified to express CD40 ligand elicit therapeutic immunity against preexisting murine tumors Blood, July 1, 2000; 96(1): 91 - 99. [Abstract] [Full Text] [PDF]
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C. Klein, H. Bueler, and R. C. Mulligan Comparative Analysis of Genetically Modified Dendritic Cells and Tumor Cells as Therapeutic Cancer Vaccines J. Exp. Med., May 15, 2000; 191(10): 1699 - 1708. [Abstract] [Full Text] [PDF]
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Y. Tada, A. Asahina, K. Nakamura, M. Tomura, H. Fujiwara, and K. Tamaki Granulocyte/Macrophage Colony-Stimulating Factor Inhibits IL-12 production of Mouse Langerhans Cells J. Immunol., May 15, 2000; 164(10): 5113 - 5119. [Abstract] [Full Text] [PDF]
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A. I. Chapoval, K. Tamada, and L. Chen In vitro growth inhibition of a broad spectrum of tumor cell lines by activated human dendritic cells Blood, April 1, 2000; 95(7): 2346 - 2351. [Abstract] [Full Text] [PDF]
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M. K. Song, S. W. Lee, Y. S. Suh, K. J. Lee, and Y. C. Sung Enhancement of Immunoglobulin G2a and Cytotoxic T-Lymphocyte Responses by a Booster Immunization with Recombinant Hepatitis C Virus E2 Protein in E2 DNA-Primed Mice J. Virol., March 15, 2000; 74(6): 2920 - 2925. [Abstract] [Full Text]
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S. M. Barratt-Boyes, M. I. Zimmer, L. A. Harshyne, E. M. Meyer, S. C. Watkins, S. Capuano III, M. Murphey-Corb, L. D. Falo Jr., and A. D. Donnenberg Maturation and Trafficking of Monocyte-Derived Dendritic Cells in Monkeys: Implications for Dendritic Cell-Based Vaccines J. Immunol., March 1, 2000; 164(5): 2487 - 2495. [Abstract] [Full Text] [PDF]
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S. Schnell, J. W. Young, A. N. Houghton, and M. Sadelain Retrovirally Transduced Mouse Dendritic Cells Require CD4+ T Cell Help to Elicit Antitumor Immunity: Implications for the Clinical Use of Dendritic Cells J. Immunol., February 1, 2000; 164(3): 1243 - 1250. [Abstract] [Full Text] [PDF]
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T. Fukao, S. Matsuda, and S. Koyasu Synergistic Effects of IL-4 and IL-18 on IL-12-Dependent IFN-{gamma} Production by Dendritic Cells J. Immunol., January 1, 2000; 164(1): 64 - 71. [Abstract] [Full Text] [PDF]
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S. Corinti, D. Medaglini, A. Cavani, M. Rescigno, G. Pozzi, P. Ricciardi-Castagnoli, and G. Girolomoni Human Dendritic Cells Very Efficiently Present a Heterologous Antigen Expressed on the Surface of Recombinant Gram-Positive Bacteria to CD4+ T Lymphocytes J. Immunol., September 15, 1999; 163(6): 3029 - 3036. [Abstract] [Full Text] [PDF]
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N. Kohrgruber, N. Halanek, M. Groger, D. Winter, K. Rappersberger, M. Schmitt-Egenolf, G. Stingl, and D. Maurer Survival, Maturation, and Function of CD11c- and CD11c+ Peripheral Blood Dendritic Cells Are Differentially Regulated by Cytokines J. Immunol., September 15, 1999; 163(6): 3250 - 3259. [Abstract] [Full Text] [PDF]
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C. Curiel-Lewandrowski, K. Mahnke, M. Labeur, B. Roters, W. Schmidt, R. D. Granstein, T. A. Luger, T. Schwarz, and S. Grabbe Transfection of Immature Murine Bone Marrow-Derived Dendritic Cells with the Granulocyte-Macrophage Colony-Stimulating Factor Gene Potently Enhances Their In Vivo Antigen-Presenting Capacity J. Immunol., July 1, 1999; 163(1): 174 - 183. [Abstract] [Full Text] [PDF]
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A. Melcher, S. Todryk, A. Bateman, H. Chong, N. R Lemoine, and R. G Vile Adoptive Transfer of Immature Dendritic Cells with Autologous or Allogeneic Tumor Cells Generates Systemic Antitumor Immunity Cancer Res., June 1, 1999; 59(12): 2802 - 2805. [Abstract] [Full Text] [PDF]
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B. W. Tillman, T. D. d. Gruijl, S. A. L.-d. Bakker, R. J. Scheper, H. M. Pinedo, T. J. Curiel, W. R. Gerritsen, and D. T. Curiel Maturation of Dendritic Cells Accompanies High-Efficiency Gene Transfer by a CD40-Targeted Adenoviral Vector J. Immunol., June 1, 1999; 162(11): 6378 - 6383. [Abstract] [Full Text] [PDF]
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