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*
Unité de Biologie des Interactions Cellulaires, CNRS 1960, and
Unité de Biologie Moléculaire du Gène, INSERM 277, Institut Pasteur, Paris, France; and
Department of Microbiology and Immunology, University of Arkansas for Medical Sciences, Little Rock, AR 72205
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Chlamydia is an obligate intracellular Gram-negative bacterium whose proliferation in host epithelial cells is characterized by two distinct developmental stages (1). Infection is initiated by adherence to the host cell of the infectious but metabolically inert elementary bodies (EBs,3 0.3 µm in diameter). Although the mechanism of internalization remains controversial (2), the bacteria appear to survive within host epithelial cells through their ability to inhibit fusion between the entry vacuoles and host cell lysosomes (3, 4). Several hours after internalization, the EBs differentiate into 1.0-µm reticulate bodies (RBs), which are noninfectious and proliferate within vacuoles that give rise within 24 h to large "inclusion bodies" that may contain up to a thousand bacteria. After approximately 2 days of infection, the RBs differentiate back to EBs, the EBs are released from the infected cell, and a new cycle of infection can begin.
Consistent with the intracellular localization of Chlamydia, cell-mediated immune responses againt Chlamydia genital infections have been observed in humans, mice, and guinea pigs (2). Transfer of either CD4+ or CD8+ T lymphocytes into naive mice has been shown to protect the mice against challenge with C. trachomatis, and studies with mice deficient in MHC molecules confirm the importance of the T cell-dependent response. Nonetheless, despite the abundance of dendritic cells (DC) in the cervix and vagina and in other epithelial tissues (5, 6, 7, 8), the possible participation of DC in warding off Chlamydia infections has not been investigated.
DC are very efficient APCs and play a pivotal role in activating T
cell-dependent immune responses (9, 10). DC present in epithelial
tissues are thought to perform a "sentinel" function, due to their
ability to internalize pathogens and process their Ags before migration
to secondary lymphoid organs where they stimulate CD4+ and
CD8+ T cells (10). Accordingly, we have studied the
interaction of two Chlamydia species, C.
trachomatis and C. psittaci, with a fully functional DC
line. Key features of the interaction were confirmed with primary DC.
The bacteria were internalized through macropinocytosis in what
appeared to be a nonspecific manner. However, unlike the results
described for other microbes, the DC killed the chlamydiae, apparently
due to early fusion between Chlamydia vacuoles and host-cell
lysosomes, a fusion that is inhibited in epithelial cells. The
chlamydiae induced secretion of TNF-
and maturation of the DC, since
presentation of an exogenous Ag was inhibited after a 24-h incubation
with the bacteria. Most importantly, the mature DC presented
efficiently chlamydial Ags to Chlamydia-specific T cells.
Given the lack of success in the development of effective
Chlamydia vaccines (11) and the recent use of DC as
adjuvants for inducing antitumor immunity in vivo (12), the use of
Chlamydia-fed DC transferred in vivo should be considered as
part of future immunization strategies against Chlamydia
infections.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The two Chlamydia species used here, the guinea pig inclusion conjunctivitis (GPIC) serovar of C. psittaci and the C. trachomatis agent of mouse pneumonitis (MoPn), have been described elsewhere (2). The DC line, D2SC/1 (13), was generously provided by Dr. P. Ricciardi-Castagnoli (University of Milan, Italy). Primary dendritic cells were isolated from the bone marrow of BALB/c mice as described (14).
The following have been previously described (15): FITC-labeled
anti-Chlamydia mAb; the anti-Chlamydia
LPS mAb; the mAb against human LAMP-1, H4A3; the mAb against the human
transferrin receptor (TfR), 5E9; the polyclonal Ab against
dinitrophenol; the probe for acidic compartments,
3-(2,4-dinitroanilino)-3'-amino-N-methyldipropyl-amine
(DAMP); cytochalasin D (CCD); 1,4-diazalbicyclo[2.2.2]octane (DABCO);
rhodamine-coupled F(ab')2 fragment goat anti-mouse Igs;
and rhodamine-conjugated F(ab')2 goat anti-rabbit Igs.
The mAb against murine LAMP, anti-CD107a, was from PharMingen (San
Diego, CA). The hybridoma producing the anti-murine TfR, R17
217.1.3, was from American Type Culture Collection (ATCC), Rockville,
MD, and the biotinylated anti-MHC class II mAb recognizing
IAb,d was from PharMingen. Rhodamine B-labeled
dextran (neutral; 40,000 kDa) and
5-(N,N-dimethyl)amiloride were from
Molecular Probes (Leiden, The Netherlands). The Factor-Test-X mouse
TNF- ELISA kit was from Genzyme (Cambridge, MA).
Cell culture conditions and infection with Chlamydia
Chlamydiae were obtained from infected HeLa cells as described (16). A partially purified suspension was aliquoted and stored at -80°C until ready for use. For heat pretreatment experiments, the bacteria were incubated at 55°C for 30 min before incubating with HeLa or DC.
Typically, the DC line was prestimulated for 2 days with 100 U/ml
IFN-
, then incubated with bacteria in RPMI 1640 for the indicated
times at 37°C in 5% CO2. Unless indicated
otherwise, the Chlamydia concentration gave between 1 and 5
bacteria adhering to the DC after a 1-h incubation; this concentration
gave a multiplicity of infection (m.o.i.) of 1:3 in HeLa cells. HeLa
cells, L cells, and primary DC were similarly incubated with bacteria
at an m.o.i. of 1:3. The primary DC were used for experiments with
Chlamydia after 4 days of culture with 10 ng/ml recombinant
granulocyte/macrophage-CSF (PharMingen).
Confocal microscopy
Samples for confocal microscopy were fixed with paraformaldehyde, incubated with Abs, and mounted as previously described (17). After fixation, DC were first incubated with blocking Abs for Fc receptors (rabbit or human serum) in permeabilization buffer. Next, to determine the distribution of the TfR, LAMP, or MHC class II for both DC and L cells, the anti-murine TfR mAb RI72 17, the anti-murine LAMP mAb, or the anti-MHC class II mAb were used in the first Ab incubation with permeabilization buffer. These mAbs were revealed with 10 µg/ml rhodamine-coupled goat anti-mouse polyclonal Abs. The bacteria were detected by incubating the cells with FITC-coupled anti-Chlamydia mAbs.
Acidic compartments were detected with DAMP as previously described (15). Macropinosomes were visualized by incubating cells growing on coverslips with 1 mg/ml of rhodamine B-labeled dextran at 37°C during the last 15 min of the infection with bacteria, which were revealed with FITC-labeled anti-Chlamydia mAb.
The distribution of the TfR and LAMP-1 were studied in HeLa cells by incubating the fixed cells with anti-human TfR mAb or the anti-human LAMP-1 mAb as primary Ab. Chlamydiae were then detected with FITC-labeled anti-Chlamydia mAb, as described above for DC and L cells.
Fluorescently labeled samples were examined with a Leica confocal microscope attached to a Leitz diaplan microscope equipped with a double argon-krypton laser. Serial optical sections were typically recorded at 0.5-µm intervals with 63x and 100x lenses.
Measurement of bacteria internalization
To distinguish between intracellular bacteria and those still on the host cell surface, cells were infected with chlamydiae, and at indicated times the host cells and associated bacteria were fixed with paraformaldehyde. However, instead of permeabilizing the cells after fixation, they were first incubated with the unlabeled anti-Chlamydia mAb, followed by the second RITC-labeled mAb. After rinsing, the samples were then permeabilized and incubated with the FITC-labeled anti-Chlamydia mAb. Photographs were taken with both the RITC and FITC filters on the same photograph frame. Internalized bacteria appeared green, while bacteria on the surface appeared red or yellow. When host cells were fixed with the fixation buffer before incubating with bacteria, all bacteria appeared red or yellow.
The effects of 10 µg/ml CCD or 10 µM dimethyl amiloride were measured by incubating the host cells with inhibitor for 30 min at 37°C before adding the chlamydiae. The inhibitors were then maintained at the same concentration throughout the internalization experiment.
Inhibition of adherence and early internalization steps of Chlamydia by heparin was measured with either HeLa or DC that had been incubated with bacteria and the indicated final concentrations of heparin (Sigma, St. Louis, MO). The bacteria were incubated in the heparin solutions in PBS for 1 h at 4°C before adding them to an equivalent volume of HeLa cells or DC in culture medium. After an hour, unbound bacteria were removed by washing once with PBS and replacing the supernatant with culture medium. After an additional 3 h, the cells were fixed, permeabilized, and incubated with FITC-labeled anti-Chlamydia mAb. Bacteria were then counted with a 63x lens using a Zeiss Axiophot fluorescence microscope.
Electron microscopy
DC or HeLa cells infected with chlamydiae for 0, 4, or 24 h were fixed with 2.5% glutaraldehyde for at least 2 h at room temperature. The fixed cells were then prepared for electron microscopy as previously described (18). Thin sections were poststained with uranyl acetate and lead citrate for examination on a Zeiss electron microscope at an accelerating voltage of 50 kV.
Measurement of TNF- secretion
Secretion of TNF- from DC was measured using the Genzyme
mouse TNF- ELISA kit following the manufacturer’s instructions. DC
were stimulated in 96-well plates with 100 IFN- for 2 days, and then
incubated with chlamydiae (at an m.o.i. of 1:3 for GPIC and 1:30 for
MoPn) for the indicated times. The supernatant was collected,
centrifuged in Eppendorf tubes to remove cellular debris, and kept
frozen at -80°C until ready for use. Supernatants were diluted in
the ELISA kit wash buffer, and the absolute concentrations of TNF-
were obtained by calibrating the ELISA kit with a known concentration
of TNF- provided by the manufacturer.
HEL Ag presentation assays
DC were plated at a concentration of 3 x
105 cells/ml and stimulated with 100 U/ml murine
rIFN- (Genzyme) and 2 ng/ml murine rGM/CSF (Genzyme) for 2 days. The
cells were washed with PBS and trypsinized for 20 to 30 min at 37°C.
After washing twice with complete medium, infected or uninfected cells
were ready for use in Ag presentation assays.
DC were fed GPIC (m.o.i. = 1:30) or MoPn (1:300) for 1 day. The DC and the HEL-specific T cell hybridoma B9.1 (19) were plated into 96-well tissue culture plates at a concentration of 105 cells/well of each cell type in 200 µl of complete medium. The DC were used to present HEL (10 mg/ml) or the HEL-derived 103–117 peptide (1 mg/ml) to stimulate IL-2 production of the B9.1 hybridoma. Cells were incubated for 24 h at 37°C, and plated in triplicate in 96-flat-bottom well microtiter plates (Costar, Cambridge, MA). IL-2 secretion after 24 h of culture was measured by transferring the supernatants to microculture wells to stimulate proliferation of the IL-2-dependent cell line CTLL-2. Proliferation was assessed by the addition of 1 µCi of [3H]thymidine per well during the last 12 h of a 36-h culture period. Cells were harvested onto paper with an automatic cell harvester (Skatron, Sterling, VA), and radioactive counts were measured in a beta plate counter (Beckman, Fullerton, CA).
Purification of Chlamydia-specific CD4+cells and proliferation assay
The ILN were removed from five mice 13 days after vaginal
inoculation with the MoPn and pooled (20). Single-cell suspensions were
incubated for 30 min on ice with TIB146 (anti-B220) and HB58
(anti-
light chain) from ATCC and anti-CD11b (clone M1/70)
and anti-CD8 (53-6.7) from PharMingen. The cells were then
incubated with microbeads conjugated to goat anti-rat Ig (Miltenyi
Biotec, Auburn, CA) for 30 min on ice. After washing, the cells were
applied to MiniMACS columns (Miltenyi Biotec). The resulting population
was 96% pure for CD4 cells. The DC line was seeded at
106 cells/ml with rIFN- (100 U/ml) and recombinant
granulocyte/macrophage-CSF (2 ng/ml) for 2 days at 37°C. The DC were
trypsinized and treated with 100 µg/ml of mitomycin C for 20 min at
37°C. ILN CD4 cells (20,000) were incubated for 5 days in 96-well
plates with 10,000 DC and 5 µg of UV-inactivated MoPn. Proliferation
was measured by [3H]thymidine incorporation
(20).
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The internalization rate of chlamydiae in epithelial cells has not
been rigorously characterized before, and it was not known whether
chlamydiae could infect DC. The bacteria were therefore incubated with
HeLa cells or DC for different times, the unbound bacteria were
removed, and the host cells with associated bacteria were fixed.
Bacteria that had not entered the host cells were then revealed by
incubating the cells with an anti-Chlamydia mAb,
followed by incubation with a RITC-labeled second Ab. Internalized
bacteria were then revealed by permeabilizing the cells and incubating
them with a FITC-labeled anti-Chlamydia mAb, which would
bind to both internalized bacteria and external bacteria. Thus,
internalized bacteria appeared green, while external bacteria were red
or yellow. Figure 1
A shows the
kinetics for bacterial entry into HeLa cells and DC. The epithelial
cells, representing the preferential physiologic target for
Chlamydia (1), were infected inefficiently, with only 20 to
30% of the associated bacteria having entered within 6 h. The DC
not only internalized the bacteria faster, but to a greater extent,
with >60% of the bacteria entering within 6 h.
Cytoskeleton-dependent phagocytosis or macropinocytosis (21) appeared
to be involved in the entry in both cell types, however, since CCD
inhibited entry by 76 ± 12% in HeLa cells and 87 ± 13% in
DC (not shown). Macropinocytosis was further implicated by the
observation that dimethyl amiloride, which inhibits macropinocytosis in
primary DC and bone marrow macrophages (22, 23), decreased chlamydial
internalization by the DC by 75 ± 7% (not shown).
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B).
On the other hand, heparin did not have a significant effect on
internalization by DC lines (Fig. 1B) or primary DC
(not shown). In addition, heat inactivation, which inhibits C.
psittaci attachment to HeLa cells (16), had no effect on the
kinetics of internalization by DC (not shown). These data suggest that
different receptors and/or entry mechanisms may be used by
Chlamydia in DC. Differential intracellular fate of Chlamydia in epithelial cells and dendritic cells
To investigate the possibility that the Chlamydia
vacuoles may be routed differently in epithelial cells and DC, host
cells were incubated with bacteria for 4 or 24 h, and the
interaction between the entry vacuoles and lysosomes was analyzed by
confocal microscopy using LAMP as a marker for lysosomes (26). Most
Chlamydia vacuoles in epithelial cells avoided fusion with
lysosomes at early stages of entry (Fig. 2A), and within 24
h the vacuoles had developed into the expected inclusions, having
diameters of approximately 10 µm, which also excluded the lysosomal
marker (Fig. 2B). However, the Chlamydia
vacuoles displayed a very different behavior in DC, as essentially all
vacuoles fused with lysosomes within a few hours. Figure 2C
shows one vacuole that had fused with lysosomes in the DC line; the
same results were obtained with a 20-fold increase in the bacterial
concentration. Fusion between Chlamydia vacuoles and
lysosomes was also observed in primary DC (not shown). No inclusions
were ever detected at 24 h. To exclude the possibility that the
intracellular fate of Chlamydia in DC may be due to
prestimulation of the DC line with IFN-, the same experiments were
repeated in the absence of IFN- prestimulation. No differences were
observed in vacuole fusion with lysosomes or survival of bacteria in
either stimulated or unstimulated DC (not shown).
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Extracellular material that is phagocytosed and sorted to
phagolysosomes is normally destined for degradation, although microbes
that survive in phagolysosomes have also been identified (30, 31). The
Chlamydia epitope recognized by the mAb LPS was sometimes
detected in DC 24 h after infection, but as LPS from both
Shigella and Chlamydia can detach from the
bacteria and may be present even when the bacteria are degraded (18, 32), we evaluated the state of the chlamydiae in HeLa cells and DC by
electron microscopy. Chlamydia EBs in tight-fitting vacuoles
were observed in HeLa cells within 4 h of infection (Fig. 3A), and at 24 h
most of the bacteria had differentiated into RBs and were proliferating
within large inclusions (Fig. 3B). In DC, many EBs
were observed in large, transparent vacuoles at 4 h (Fig. 3C), and many bacteria that appeared to be severely
damaged were also observed (not shown), although in most cases it was
difficult to distinguish them from other cellular debris. No intact
bacteria were ever found after a 24-h infection.
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Pathogen-induced maturation of DC implies that, among other
changes, the DC lose the capacity to present new exogenous Ag (9, 10).
We therefore determined whether a 24-h incubation with chlamydiae had
any effect on the ability to subsequently present HEL to a HEL-specific
CD4+ T cell hybridoma (19). DC were able to present
HEL efficiently to the hybridoma, but the ability to stimulate the
hybridoma decreased dramatically following incubation with either
C. psittaci (Fig. 4) or
C. trachomatis (not shown). The inhibition of HEL
presentation was not due to loss of surface expression of MHC class II,
since the DC incubated with either of the two Chlamydia
species could still present an HEL-derived peptide added exogenously,
which can presumably bind to surface MHC class II without being
internalized (33).
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Given the ability of chlamydiae to inhibit presentation of
exogenous HEL, we examined the interaction between Chlamydia
vacuoles and compartments involved in Ag presentation by MHC class II.
The DC line displayed numerous macropinosomes with diameters of 2 µm
to over 10 µm, as determined by incubating DC with rhodamine-labeled
dextran. At early stages of Chlamydia entry, bacteria also
colocalized with the dextran (Fig. 5A), suggesting that
many if not all bacteria are internalized through macropinocytosis.
Since entry of bacteria into DC is slow, whereas the fluorescence
signal from macropinosomes loaded with dextran is intense and saturates
within 30 min, the cells were incubated with chlamydiae for 2 h
before incubating with dextran for an additional 15 min. Thus, not all
internalized bacteria colocalized with dextran, probably because some
of the bacteria entered before the incubation with dextran. No dextran
was accumulated in HeLa cells under the same conditions (not shown).
The entry vacuoles in DC were also acidic (not shown), consistent with
their expression of lysosomal markers, and expressed MHC class II
molecules (Fig. 5B), suggesting that chlamydiae could
be processed for Ag presentation in these compartments.
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Chlamydia-induced TNF- secretion by dendritic cells
In addition to bacterial products, TNF- was shown to induce
differentiation of primary DC into a phenotype with few macropinosomes
and low ability to present exogenous Ags (22, 34). To ascertain whether
chlamydiae might induce differerentiation of the DC line independently
of bacterial internalization and degradation, we measured whether
incubation with Chlamydia would cause secretion of TNF-
by the DC. As seen in Figure 6, both
C. trachomatis and C. psittaci induced secretion
of high levels of TNF-. Significant levels of TNF- in the
supernatant were already detected within 2 h, at which time few
bacteria have been internalized, suggesting that simple contact between
bacterial surface epitopes and the DC may contribute to TNF-
secretion.
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To determine whether the DC were capable of presenting Ag to
sensitized T cells, mice were infected intravaginally with C.
trachomatis, and ILN were removed 13 days later. CD4+
T cells were obtained and incubated with C. trachomatis and
DC. When the proliferative response was assessed, DC or
CD4+ T cells incubated separately with Ag displayed little
if any proliferation, while a large increase in proliferation was
observed when CD4+ T cells were incubated in the presence
of DC and Chlamydia Ag (Fig. 7). These data demonstrate that the DC
not only express Chlamydia Ag-MHC complexes on the cell
surface, but that these complexes are capable of providing a
stimulatory signal for sensitized CD4+ T cells.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The general outlines of the development and migration of DC have been previously described (9, 10). Bone marrow-derived DC precursors translocate to nonlymphoid tissue, including epithelia of the genital tract, where they develop into "immature DC" capable of internalizing and processing Ag and expressing high levels of MHC molecules. Pathogens and inflammatory cytokines can induce their differentiation into "mature DC" with a decreased capacity for Ag processing but stable presentation of previously acquired Ags by the MHC molecules. The mature DC migrate into secondary lymphoid tissues, where they initiate T cell-dependent immune responses.
We have characterized the interaction between two species of Chlamydia, C. trachomatis and C. psittaci, and DC using a fully functional DC line (13). In parallel, key features of internalization, namely the effects of heparin and fusion of Chlamydia vacuoles with lysosomes, were confirmed with primary DC. These results were compared with the intracellular behavior of the chlamydiae in an epithelial cell line, HeLa, representing the preferred physiologic target cell of the bacteria. The most striking difference between the epithelial cells and the DC is that the entry vacuoles avoid fusion with lysosomes in the epithelial cells, evolving within a day into large inclusions, while in DC there is rapid fusion between the Chlamydia vacuoles and host cell lysosomes. By electron microscopy, chlamydiae, mostly at the metabolically active RB developmental stage, were found proliferating in large numbers in the epithelial cells after 1 day. In DC, on the other hand, no intact bacteria were detected in DC after a 1-day incubation, suggesting that the bacteria were killed by the DC.
At early time points, chlamydiae were detected in large vacuoles in the DC, consistent with macropinosomes, and the internalization was inhibited by dimethyl amiloride. This internalization appears to be nonspecific or at least different from that used in epithelial cells, since heparin, which inhibits invasion of epithelial cells (16, 24, 25), has no effect on DC internalization. Thus, it is conceivable that DC could also internalize other bacteria through macropinocytosis.
Interaction of the DC with Chlamydia leads to maturation of the DC, as defined by down-regulation of macropinocytosis and loss of the ability to present new exogenous Ag. The decrease in macropinocytosis was observed by fluorescence microscopy, which revealed a large drop in the number of acidic vesicles expressing LAMP and capable of accumulating dextran. The intracellular distribution of MHC class II in the DC line suggests that the DC line behaves as immature primary DC (14), while the disappearance of intracellular class II molecules after incubation with Chlamydia is consistent with the phenotype of mature primary DC (14). In addition, after a 1-day incubation with Chlamydia, the DC were no longer able to present exogenous HEL to HEL-specific T cells, but they still presented chlamydial Ags efficiently to T cells, in line with the role of mature DC to present Ags from the pathogen that induced the maturation.
As both infectious and inactivated chlamydiae were used in our
experiments, it is likely that some of the stimulatory activity of
C. trachomatis and C. psittaci may be due to
TNF- secretion induced by simple contact with Chlamydia
surface epitopes. Assuming that surface epitopes (e.g., LPS) from other
bacteria may also induce differentiation of DC and that other bacteria
may also be internalized through macropinocytosis, the only requirement
for subsequent presentation of bacterial Ags by the DC is that the
macropinosomes fuse with lysosomes (39), where the bacteria could be
degraded for subsequent presentation by MHC class II. Although this
appears to be the case for Chlamydia, this argument cannot
be generalized to all microbes, as seen in the example of L.
monocytogenes, which escapes from DC phagolysosomes (40).
The observation that infection with Chlamydia gives rise to short-lived CD4+-dependent immunity against subsequent infections (2) implies that the frequency of T cells recognizing Chlamydia Ags is low. The frequency of active, Chlamydia-specific T cells may thus be augmented by transferring Chlamydia-fed DC into animals, as has recently been done with DC to induce protective immunity to lethal challenge by tumors (12). This strategy could be extended to microbes that nominally survive within DC by incubating DC with previously inactivated microbes.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. David Ojcius, Institut Pasteur, Unité de Biologie des Interactions Cellulaires, 25 rue du Dr. Roux, 75724 Paris Cedex 15, France. E-mail address:
3 Abbreviations used in this paper: EB, elementary body; CCD, cytochalasin D; DC, dendritic cell; HEL, hen egg lysozyme; RB, reticulate body; GPIC, guinea pig inclusion conjunctivitis; MoPn, mouse pneumonitis; RITC, rhodamine isothiocyanate; ILN, iliac lymph nodes; LAMP, lysosome associated membrane protein; DAMP, 3-(2,4-dinitroanilino)-3'-amino-N-methyldipropylamine; TfR, transferrin receptor; m.o.i., multiplicity of infection.
Received for publication July 23, 1997. Accepted for publication October 14, 1997.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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S. L. Newman and A. Holly Candida albicans Is Phagocytosed, Killed, and Processed for Antigen Presentation by Human Dendritic Cells Infect. Immun., November 1, 2001; 69(11): 6813 - 6822. [Abstract] [Full Text] [PDF]
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A. Rupper, K. Lee, D. Knecht, and J. Cardelli Sequential Activities of Phosphoinositide 3-Kinase, PKB/Akt, and Rab7 during Macropinosome Formation in Dictyostelium Mol. Biol. Cell, September 1, 2001; 12(9): 2813 - 2824. [Abstract] [Full Text] [PDF]
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C. W. Cutler, R. Jotwani, and B. Pulendran Dendritic Cells: Immune Saviors or Achilles' Heel? Infect. Immun., August 1, 2001; 69(8): 4703 - 4708. [Full Text] [PDF]
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L. A. Gildea, R. E. Morris, and S. L. Newman Histoplasma capsulatum Yeasts Are Phagocytosed Via Very Late Antigen-5, Killed, and Processed for Antigen Presentation by Human Dendritic Cells J. Immunol., January 15, 2001; 166(2): 1049 - 1056. [Abstract] [Full Text] [PDF]
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 R. Coutinho-Silva, J.-L. Perfettini, P. M. Persechini, A. Dautry-Varsat, and D. M. Ojcius Modulation of P2Z/P2X7 receptor activity in macrophages infected with Chlamydia psittaci Am J Physiol Cell Physiol, January 1, 2001; 280(1): C81 - C89. [Abstract] [Full Text] [PDF]
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D. Askew, R. S. Chu, A. M. Krieg, and C. V. Harding CpG DNA Induces Maturation of Dendritic Cells with Distinct Effects on Nascent and Recycling MHC-II Antigen-Processing Mechanisms J. Immunol., December 15, 2000; 165(12): 6889 - 6895. [Abstract] [Full Text] [PDF]
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M. Svensson, C. Johansson, and M. J. Wick Salmonella enterica Serovar Typhimurium-Induced Maturation of Bone Marrow-Derived Dendritic Cells Infect. Immun., November 1, 2000; 68(11): 6311 - 6320. [Abstract] [Full Text] [PDF]
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 P. M. Bavoil, R.-c. Hsia, and D. M. Ojcius Closing in on Chlamydia and its intracellular bag of tricks Microbiology, November 1, 2000; 146(11): 2723 - 2731. [Full Text]
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N. Mohagheghpour, A. van Vollenhoven, J. Goodman, and L. E. Bermudez Interaction of Mycobacterium avium with Human Monocyte-Derived Dendritic Cells Infect. Immun., October 1, 2000; 68(10): 5824 - 5829. [Abstract] [Full Text] [PDF]
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M. Amyere, B. Payrastre, U. Krause, P. V. D. Smissen, A. Veithen, and P. J. Courtoy Constitutive Macropinocytosis in Oncogene-transformed Fibroblasts Depends on Sequential Permanent Activation of Phosphoinositide 3-Kinase and Phospholipase C Mol. Biol. Cell, October 1, 2000; 11(10): 3453 - 3467. [Abstract] [Full Text]
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C. S. Subauste and M. Wessendarp Human Dendritic Cells Discriminate Between Viable and Killed Toxoplasma gondii Tachyzoites: Dendritic Cell Activation After Infection with Viable Parasites Results in CD28 and CD40 Ligand Signaling That Controls IL-12-Dependent and -Independent T Cell Production of IFN-{gamma} J. Immunol., August 1, 2000; 165(3): 1498 - 1505. [Abstract] [Full Text] [PDF]
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D. Fortsch, M. Rollinghoff, and S. Stenger IL-10 Converts Human Dendritic Cells into Macrophage-Like Cells with Increased Antibacterial Activity Against Virulent Mycobacterium tuberculosis J. Immunol., July 15, 2000; 165(2): 978 - 987. [Abstract] [Full Text] [PDF]
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F. Garcia-Del Portillo, H. Jungnitz, M. Rohde, and C. A. Guzman Interaction of Salmonella enterica Serotype Typhimurium with Dendritic Cells Is Defined by Targeting to Compartments Lacking Lysosomal Membrane Glycoproteins Infect. Immun., May 1, 2000; 68(5): 2985 - 2991. [Abstract] [Full Text] [PDF]
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J.-L. Perfettini, T. Darville, G. Gachelin, P. Souque, M. Huerre, A. Dautry-Varsat, and D. M. Ojcius Effect of Chlamydia trachomatis Infection and Subsequent Tumor Necrosis Factor Alpha Secretion on Apoptosis in the Murine Genital Tract Infect. Immun., April 1, 2000; 68(4): 2237 - 2244. [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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L. Van Overtvelt, N. Vanderheyde, V. Verhasselt, J. Ismaili, L. De Vos, M. Goldman, F. Willems, and B. Vray Trypanosoma cruzi Infects Human Dendritic Cells and Prevents Their Maturation: Inhibition of Cytokines, HLA-DR, And Costimulatory Molecules Infect. Immun., August 1, 1999; 67(8): 4033 - 4040. [Abstract] [Full Text] [PDF]
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S.-K. Kim, M. Angevine, K. Demick, L. Ortiz, R. Rudersdorf, D. Watkins, and R. DeMars Induction of HLA Class I-Restricted CD8+ CTLs Specific for the Major Outer Membrane Protein of Chlamydia trachomatis in Human Genital Tract Infections J. Immunol., June 1, 1999; 162(11): 6855 - 6866. [Abstract] [Full Text] [PDF]
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R. Coutinho-Silva, P. M. Persechini, R. D. C. Bisaggio, J.-L. Perfettini, A. C. T. D. S. Neto, J. M. Kanellopoulos, I. Motta-Ly, A. Dautry-Varsat, and D. M. Ojcius P2Z/P2X7 receptor-dependent apoptosis of dendritic cells Am J Physiol Cell Physiol, May 1, 1999; 276(5): C1139 - C1147. [Abstract] [Full Text] [PDF]
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D. Zhang, X. Yang, H. Lu, G. Zhong, and R. C. Brunham Immunity to Chlamydia trachomatis Mouse Pneumonitis Induced by Vaccination with Live Organisms Correlates with Early Granulocyte-Macrophage Colony-Stimulating Factor and Interleukin-12 Production and with Dendritic Cell-Like Maturation Infect. Immun., April 1, 1999; 67(4): 1606 - 1613. [Abstract] [Full Text] [PDF]
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H. Lu and G. Zhong Interleukin-12 Production Is Required for Chlamydial Antigen-Pulsed Dendritic Cells To Induce Protection against Live Chlamydia trachomatis Infection Infect. Immun., April 1, 1999; 67(4): 1763 - 1769. [Abstract] [Full Text] [PDF]
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 H Boleti, A Benmerah, D. Ojcius, N Cerf-Bensussan, and A Dautry-Varsat Chlamydia infection of epithelial cells expressing dynamin and Eps15 mutants: clathrin-independent entry into cells and dynamin-dependent productive growth J. Cell Sci., January 5, 1999; 112(10): 1487 - 1496. [Abstract] [PDF]
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D. M. Ojcius, P. Souque, J.-L. Perfettini, and A. Dautry-Varsat Apoptosis of Epithelial Cells and Macrophages Due to Infection with the Obligate Intracellular Pathogen Chlamydia psittaci J. Immunol., October 15, 1998; 161(8): 4220 - 4226. [Abstract] [Full Text] [PDF]
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 H. Su, R. Messer, W. Whitmire, E. Fischer, J. C. Portis, and H. D. Caldwell Vaccination against Chlamydial Genital Tract Infection after Immunization with Dendritic Cells Pulsed Ex Vivo with Nonviable Chlamydiae J. Exp. Med., September 7, 1998; 188(5): 809 - 818. [Abstract] [Full Text] [PDF]
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| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
) and HeLa cells (
) and
effect of heparin. A, Host cells were incubated with
C. psittaci for the indicated times, and internalized
bacteria were visualized as described in Materials and
Methods. B, Cells were incubated with C.
psittaci for 4 h in the presence of the indicated
concentrations of heparin.
