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Centre d’Immunologie de Marseille-Luminy, Marseille, France
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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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Dendritic cells are thought to be the most efficient APC and as such are considered critical for the induction of immune responses (7). Lacking true Ag-specific receptors, immature dendritic cells can acquire Ags by fluid phase endocytosis and present them to T cells (8, 9, 10). This raises the question as to how an Ag is "perceived" by the dendritic cells in peripheral tissues, i.e., by what mechanisms Ag uptake induces the initiation of dendritic cell migration to secondary lymphoid organs (11) and up-regulation of costimulatory molecules (12, 13), which are events implicated in maturation of dendritic cells and necessary for their interaction with T cells. Dendritic cells express "pattern recognition receptors" (14), with affinity for repeating determinants, such as mannosyl groups, present on many bacteria (9, 15). These receptors clearly augment the efficiency of dendritic cells as APC for T cell stimulation, but the relative roles of Ag acquisition vs signals transmitted by these receptors have not been determined. Another strategy available for dendritic cells is to use the products of acquired immunity to recognize microorganisms expressing antigenic determinants identical to or cross-reactive with those for which Ab exists. The role of Fc receptor (FcR) in increasing the efficiency of presentation of opsonized Ag by dendritic cells has been reported (8, 16).
Both surface Ig and Fc
RIII signal intracellularly use immunoreceptor
tyrosine-associated motifs on cytoplasmic domains of the surface
Ig-associated Ig
- and Igß-chains and the -chain of FcRIII
(17), or closely related sequences for FcRII (18), suggesting
analogous downstream events following receptor engagement. We therefore
investigated the role of multivalent vs monovalent Ag targeted to
surface Ig of B cells and the FcR of dendritic cells, with respect to
the capacity of these cells to serve as APC to HEL-specific T cells
from TCR transgenic mice. We compared these molecules with MHC class I
and II determinants, which are present on both cells but are not known
to be receptors implicated in the uptake of exogenous Ag.
In this study we used HEL-containing liposomes with repeating hapten
determinants on their surfaces. Results presented in this paper
indicate that B cells transgenic for an anti-HEL surface Ig
presented Ag much more efficiently when the protein was encapsulated in
liposomes targeted to surface Ig than when HEL was free in solution.
HEL encapsulated in liposomes opsonized by IgG Abs to the hapten is
taken up and efficiently presented to T cells by dendritic cells via Ig
FcR (FcRII and/or FcRIII), while HEL in nonopsonized liposomes
was taken up and presented inefficiently. However, our results indicate
that other cell surface molecules of dendritic cells, namely class I
and class II MHC molecules, also permit efficient presentation of Ag in
targeted liposomes. This is in contrast to the reduced capacity of
these same molecules to serve an Ag-presenting function when B cells
are used as APC. These results underline the different strategies of
exclusive uptake by B cells of ligands corresponding to the specificity
of their surface Ig as compared with less fastidious uptake by
dendritic cells of particulate Ags with which they come into contact by
means of a more varied group of cell surface molecules.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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CBA/J mice transgenic for rearranged IgM and IgD Abs with variable regions derived from an Ab (HH10) with high affinity for HEL (19) were generated (20) using plasmids provided by A. Basten (Centenary Institute, Newtown, New South Wales, Australia). Characteristics of other transgenic lines made with these plasmids have been reported in detail (21). TCR transgenic mice (22), which express the same TCR as the 3A9 hybridoma (23), specific for the immunodominant HEL peptide 46–61 in the context of H-2 I-Ak, were kindly provided by M. Davis (Stanford University, Palo Alto, CA). Transgenic mice were maintained as heterozygotes backcrossed to CBA/J mice (IFFA-CREDO, L’Arbresle, France) and identified, by an ELISA for serum IgM anti-HEL for Ig transgenic mice (20) or by FACS analysis of TCR transgenic mice, as those in which a majority of peripheral blood CD4+ cells express Vß8.2, the family of the gene used for construction of the strain (22). Mice used were between 6 and 12 wk of age.
Cells
Dendritic cells were obtained from femur bone marrow of normal CBA/J mice cultured in Iscove’s modified Dulbecco’s medium with 10% FCS, antibiotics, 2 x 10-5 M 2-ME, glutamine, and supernatants from NIH3T3 cells supplemented with 10–20 ng/ml murine granulocyte macrophage-CSF (24). After 3 days of culture, cells were diluted 1:1 in the same medium, and, after an additional 4–6 days of culture, plastic nonadherent cells were used as APC. At this time, the majority of these had immature dendritic cell morphology. B cells were purified by treatment of spleen cells with Abs to Thy-1, CD4 and CD8, and rabbit complement. T cells from the spleen were purified by passage over nylon wool columns, as described (25).
Antibodies
Abs used are described in Table I
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The protein A-anti-DNP (PA-anti-DNP) conjugate was prepared by
covalent coupling of Fab fragments of the mouse IgG1 anti-DNP mAb
265.5 (Immunotech, Marseille, France) to equimolar quantities of
protein A (Pharmacia, Piscataway, NJ), modified with
N-succinimidyl-6-maleimido caproate (Sigma, St. Louis, MO).
The product was separated from multimeric conjugates and noncoupled
species on Superdex 200 columns (Pharmacia). Monoclonal IgG1 and IgG2a
anti-DNP Abs were produced from hybridoma cells kindly provided by
Zelig Eshhar (Weizmann Institute of Science, Rehovot, Israel). All
anti-DNP Abs were purified on protein G-Sepharose (Pharmacia). Ab
7.6 is an anti-Id (20) specific for HH10, the anti-HEL Ab
expressed by the transgenic B cells used in this study. Other Abs were
provided by the groups that published them or by the American Type
Culture Collection (Manassas, VA).
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Liposomes (80 µmol with respect to lipids) were made from 65% (mol/mol) dimyristoyl phosphatidylcholine and 34.5% cholesterol (both from Sigma) and 0.5% DNP-caproylphosphatidylethanolamine (Molecular Probes, Eugene, OR). Liposomes were formed by exposing lipids evaporated from chloroform/methanol (9:1 v/v) to an aqueous solution containing HEL (Sigma) (10 mg/ml; 700 µM) and 10 mM carboxyfluorescein (CF) (Molecular Probes) in PBS. For some experiments, liposomes of the same composition were made in CF/PBS or PBS alone. Following repeated cycles of freezing and thawing, liposomes were formed by extrusion (Extruder; Lipex Biomembranes, Vancouver, British Columbia, Canada) (26) through polycarbonate filters of 200-nm pore size (Corning Costar, Oneonta, NY) at 45°C, followed by gel filtration over Sepharose 4B columns (Pharmacia) to eliminate unencapsulated solute. Lipid vesicles were sterilized by filtration through 0.45 µm filters (Gelman, Ann Arbor, MI). Different HEL concentrations in lipid vesicles were obtained by dilution of this stock solution. HEL levels were confirmed by fluorescence of dilutions of free and liposome-entrapped CF with reference to the stock solution of HEL and CF used for liposome preparation. Use of 125I-labeled HEL has confirmed that the fraction of encapsulated CF accurately reflects passive HEL entrapment (4). The internal volume of the liposomes used is about 4.2 x 10-18 liters, so at 700 µM Ag used, each liposome contains about 1750 HEL molecules.
Tests of Ag presentation
A schematic of the liposomes, the PA-anti-DNP conjugate, and
the protein A-binding Ab that are used to target determinants other
than the FcR is presented in Fig. 1A. Direct opsonization of the
same liposomes by anti-DNP Abs, used to target the FcR, is
presented in Fig. 1B. Twenty thousand (or as specified) B or
dendritic cells were distributed in wells of 96-well flat-bottom
microtiter plates (Costar, Cambridge, MA) in RPMI 1640
supplemented with 5% FCS, 2 x 10-5 M 2-ME,
glutamine, and antibiotics. Tenfold dilutions from the above stock
solutions of free HEL or DNP liposomes containing HEL were added to the
wells together with the PA-anti-DNP conjugate and/or Abs at a final
concentration of 5 µg/ml. In some experiments, a washing step
separated the exposure of cells to Ag and Abs or liposomes. Cells and
Ags were incubated overnight at 37°C in a total volume of 100
µl, after which 20,000 T cells from transgenic or normal mice were
added and incubated for an additional 48 h in a total volume of
110 µl. At this time, the concentration of IL-2 in supernatant fluids
was determined by adding them to IL-2-dependent CTLL cells
(10,000 cells/well). Following a 16-h incubation, proliferation of CTLL
was assessed by measurement of radiolabeled thymidine incorporation
into DNA at the end of an additional 8-h incubation in the presence of
0.5 µCi [3H]thymidine. For studies of T cell
proliferation, dendritic cells were irradiated (2000 rads) before Ag
addition and [3H]thymidine incorporation by T cells
during an 8-h period measured following 96-h exposure to APC. All
points are the means of duplicate determinations.
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Dendritic cells or B cells were incubated on ice with unlabeled or fluorescein-modified Abs (5 µg/ml), the PA-anti-DNP conjugate (5 µg/ml), and/or fluorescent or nonfluorescent liposomes corresponding to an HEL concentration of 100 nM before fixation. Analyses were performed on a FACScan using Lysis II and CellQuest software (Becton Dickinson, Mountain View, CA).
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Ag presentation to transgenic T cells was evaluated using B cells
from anti-HEL transgenic or nontransgenic CBA/J mice as APC (Fig. 2). HEL was either free in solution or
encapsulated in hapten-bearing liposomes. B cells from nontransgenic
mice were unable to present Ag in either free or lipid
vesicle-encapsulated form at HEL concentrations below 100 nM (1.5
µg/ml; Fig. 2A). As expected, B cells from anti-HEL Ig
transgenic mice efficiently presented soluble HEL at concentrations as
low as 10 pM (Fig. 2B). HEL encapsulated in liposomes was
only slightly more efficiently presented by these transgenic mice than
by nontransgenic mice in the absence of a cell-targeting Ab. Since
HEL-specific B cells presented free HEL at 100-fold lower
concentrations, this indicates that maximal leakage of HEL from lipid
vesicles did not exceed 1% of the encapsulated Ag. Anti-hapten Fab
coupled to protein A permitted targeting liposomes to surface IgM and
IgD of Ig transgenic mice via the protein A binding anti-Id Ab 7.6
(20). In this situation, HEL in liposomes was presented at
concentrations that never exceeded 1 pM HEL, a concentration 100-fold
less than that required for presentation of free HEL directly taken up
by the HEL-specific surface Ig and 100,000-fold less than free HEL or
HEL in liposomes taken up by nonspecific B cells (Fig. 2A).
The increased presentation of HEL in ligand-bearing liposomes, as
compared with a monovalent HEL bound to the same receptor, could be
related to increased delivery of the HEL into specialized intracellular
compartments, up-regulation of costimulatory molecules due to the
cross-linking of surface Ig, and/or an increased quantity of HEL in
liposomes associated with the cells. Two series of experiments were
performed to attempt to differentiate these possibilities.
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. Responses to B cells incubated with
HEL and then washed were comparable with responses without a washing
step (data not shown). When cells were subsequently incubated with the
7.6 anti-Id a marked response was noted at an Ag concentration
100-fold lower than for cells incubated by Ag alone. This response was
slightly increased by the addition of the PA-anti-DNP complex and
was not further increased when DNP-bearing empty liposomes were added.
No increased response was seen for liposomes in the presence of a
control Ab, and no response at all seen for empty anti-Id targeted
liposomes in the absence of free HEL. Thus, a fixed Ag dose was
presented more efficiently when the surface Ig to which it bound was
cross-linked. However, when targeted liposomes contained Ag the
response was at plateau levels for Ag concentrations 100 times lower.
Since induction of costimulation induced by cross-linking of surface Ig
would be expected to be the same under conditions in which empty or
HEL-containing liposomes were used, our results may be explained
by a surface Ig-signaling-dependent modification of HEL
presentation.
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A), while differences in
binding of liposomes to these different determinants were minor (Fig. 4B). Ag targeted by an Ab to the I-Ak molecule
was not presented, since this Ab is present in excess during the
experiment and blocks recognition of the I-Ak-HEL peptide
complex recognized by the TCR. Ag targeted to H-2Kk or
I-Ek was presented with the same efficiency whether
transgenic or nontransgenic B cells were used (data not shown). Thus,
the advantage for Ag presentation for HEL in liposomes targeted to
surface Ig may not be explained only by increased Ag binding.
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Dendritic cells are known to be excellent APC and are also known
to have active fluid phase endocytosis (10). This endocytic activity is
presumably related to the APC function, but few studies have compared
presentation of Ag taken up by fluid phase or receptor-mediated
processes. Dendritic cells were incubated with HEL or
DNP-bearing liposomes in the absence of cell-targeting Abs or in the
presence of IgG1 or IgG2a monoclonal anti-DNP Abs, together with
HEL-specific T cells. Results for IgG1 are presented in Fig. 5A. Neither free HEL nor lipid
vesicle-encapsulated HEL incubated with an irrelevant Ab were presented
at concentrations below 10 nM. This minimum concentration is similar to
those of published results for the presentation of free HEL by splenic
dendritic cells to HEL-specific T hybridomas (27). In the presence of
IgG1 or IgG2a anti-DNP Ab, DNP-bearing liposome-encapsulated HEL
was as efficient for T cell stimulation at 10 pM, at least 1000-fold
less than free HEL or nontargeted lipid vesicles. The anti-DNP Abs
had no effect when HEL-containing liposomes were made without DNP on
their membranes, or when DNP-bearing liposomes were made without
encapsulated HEL (data not shown). The IgG FcR dependence of
presentation mediated by the anti-DNP Ab was shown by the absence
of efficient presentation in the presence of Fab (or
F(ab')2, data not shown) fragments of the IgG1 Ab and by
inhibition of presentation of liposomes incubated with IgG1 (or IgG2a,
not shown) by preincubation with the anti-FcR Ab 2.4G2 (Fig. 5B). In parallel experiments, HEL in liposomes opsonized by
intact anti-DNP Abs was not presented by B cells (data not shown),
consistent with the inability of the B cell isoform of the FcR to take
up Ag (28).
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C). Binding of fluorescence-labeled anti-DNP Abs was
low in the absence of DNP-bearing liposomes. Their binding increased
markedly in the presence of nonfluorescent DNP liposomes. The binding
and presentation mediated by the 265.5 IgG1 Ab was similar to that of
U7.6.3, an independently derived monoclonal IgG1 anti-DNP Ab (data
not shown). Similar patterns of binding were seen when we used
nonfluorescent anti-DNP Abs and DNP-bearing fluorescent liposomes
(data not shown). Inhibition by 2.4G2 of binding of liposomes to the
cells mediated by IgG1 and IgG2a Abs (Fig. 5) correlated with
inhibition of Ag presentation via those isotypes (data not shown).
Since Ab 2.4G2 is specific for FcR II/III (29), all of the
FcR-mediated binding and presentation are due to targeting to one or
both of these receptors. The homologue of the high affinity (CD64) Ig
FcR reported to be present on dendritic cells from human blood (30, 31)
consequently does not appear to participate in the uptake of Ag
by mouse dendritic cells derived from bone marrow under these culture
conditions. Ag presentation by dendritic cells is also efficient for multivalent ligands targeted to several cell surface determinants other than the FcR
The above results indicate that the FcR can play a role for
dendritic cells analogous to that of surface Ig for B cells. In
contrast to the situation for B cells, increased Ag presentation by
dendritic cells was also seen when the PA-anti-DNP conjugate was
used for targeting together with anti-class I (H-2Kk)
or II (I-Ek) Abs (Fig. 6B); a separate experiment
targeting I-Ek is presented in Fig. 6A. As for
the situation with B cells, Ag presentation was not seen when the
restricting I-Ak molecule was targeted (Fig. 6B). The anti-FcR Ab 2.4G2 did not inhibit
presentation of Ag delivered in the presence of protein A-bearing Abs
(data not shown). Thus, dendritic cells are not efficient APC for HEL
present free in solution or in neutral liposomes at biologically
relevant concentrations (
150 ng/ml) but may present Ag to T cells
about as efficiently as Ag-specific B cells (15–150 pg/ml), provided
that the Ag makes multivalent contact with the cell. The same class I
and II molecules that were inefficient for Ag presentation when
expressed on B cells were more efficient when dendritic cells were
targeted.
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We asked whether the more efficient Ag presentation by
dendritic cells for FcR targeted as compared with nontargeted Ag
correlated with a more efficient Ag presentation by each
cell, thus reducing the number of APC required. The number of dendritic
cells necessary for the stimulation of IL-2 production by transgenic T
cells was evaluated as a function of the mode of Ag acquisition.
Variable numbers of irradiated dendritic cells were incubated with 10
nM free or 1 nM liposome-encapsulated Ag targeted by intact or Fab
fragments of an IgG1 anti-DNP Ab. As few as 1000 dendritic cells
generated a plateau level of IL-2 production when the FcR was targeted,
whereas 10 times more dendritic cells were required for plateau
stimulation of T cells at a 10-fold higher concentration of free Ag.
Dendritic cells incubated under conditions of optimal Ag stimulation
but in the absence of transgenic T cells failed to produce any factor
capable of stimulating [3H]thymidine incorporation by
CTLL cells (Fig. 7A).
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B). |
Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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B). Alternatively, the use of these liposomes with
PA-anti-DNP Fab conjugates permitted intact protein A-binding Abs
to be oriented correctly toward other target molecules (Fig. 1A) while simultaneously blocking association with Fc
receptors (32). Liposomes of the type used in this study are
consequently models for infectious agents, which may interact with any
of a number of cell surface determinants via multivalent ligands (13),
and which contain multiple copies of Ag behind the barrier of a
membrane or capsule. In the present experiments, each liposome
expresses more than 1500 DNP molecules, permitting a high density of
targeting ligands, based on DNP binding by intact anti-DNP Abs to
dendritic cells or by PA-anti-DNP conjugates to other determinants
on B cells or dendritic cells.
Differences in stimulation of Ag-specific T cells will depend on the
consequences of interaction between Ag, or the liposome in which it is
entrapped, and the particular APC surface molecule to which the Ag is
targeted. These will be determined by 1) the density of the target
molecules; 2) the signals generated by binding of Ag to the receptor;
3) the rate and extent of their endocytosis; 4) the quantity of the Ag
delivered; and 5) the intracellular fate of the Ag, including
degradation and access to class II molecules. Intact monovalent Ag has
been shown to be able to permit transgenic B cells to stimulate
transgenic T cells bearing the same TCR as those used in the present
study, at comparable Ag concentrations (6, 22). This monovalent form of
Ag presentation by B cells is shown in the short term in vitro studies
performed here to be much less efficient for stimulation than the same
quantity of Ag bound to surface Ig that is subsequently cross-linked by
Ab or empty liposomes. This difference for the same target molecule and
APC is consistent with modulation of signals that are transmitted by
surface Ig as a consequence of cross-linking, as has already been
demonstrated in studies in which mutated Ig or Igß molecules
associated with surface Ig on B cells were transfected into myeloma or
lymphoma cells (33, 34). However, cross-linking of surface Ig in the
presence of a fixed amount of bound free HEL increased Ag presentation
to a much lesser extent than when liposomes bound to surface Ig
contained HEL (Fig. 3). HEL is passively entrapped within the aqueous
spaces of these liposomes and presumably behaves like free HEL when
released in cells. This is in contrast to experiments in which Ags have
been covalently hapten modified for binding to anti-hapten surface
Ig (35, 36) or directly bound to anti-Ag Abs for targeting
purposes. These may alter Ag processing, or result in modification of
their presentation, depending on the site on the Ag to which the Ab
binds (37, 38). In other experiments we have studied Ag presentation to
T hybridomas specific for the 46–61 determinant, which requires
extensive processing for presentation and is associated primarily with
newly synthesized class II molecules, as well as T hybridomas specific
for peripheral peptides included within the sequence of HEL, which are
presented by recycling class II molecules. These results indicate that
HEL in liposomes behaves like free HEL when taken up by fluid phase
endocytosis and shares the markedly enhanced presentation of HEL when
targeted to surface Ig, whatever the peptide specificity of the
hybridoma.5 There is thus no
evidence that a specialized processing compartment is required for
release of liposome-entrapped Ag.
It has been estimated that fewer than 300 HEL peptide-IAk complexes may activate class II- plus peptide-specific T cell hybridomas (39), whereas about 1 HEL molecule in 750 taken up by fluid phase endocytosis by lymphoma cells generated I-Ak-associated peptides recognized by a hybridoma with the same TCR as that expressed by our TCR transgenic T cells (40). Although naive transgenic T cells are reported to require a somewhat higher number of class II-peptide complexes for IL-2 production than primed T cells or hybridomas (41), the efficiency of class II association of Ag taken up by a receptor on APC may permit this level of peptide binding to be achieved at low external Ag concentrations, as suggested by the present studies. The efficiency of this process will also depend on the form of the Ag. Each liposome used here contains hundreds of HEL molecules, so that surface Ig targeted by anti-Id will internalize much more Ag than the two HEL molecules in free form taken up by the same receptor. Thus, the internalization of Ag in monovalent form could be considered as an "analogue" event, with the Ag dose internalized linearly with increasing numbers of receptors engaged, whereas internalization of multivalent Ag is a "quantal" event, in which one or only a few liposomes may contain sufficient Ag to induce a response, if delivered into a suitable compartment. This is an appropriate response in nature, in which quanta of Ag will normally be contained within infectious organisms. However, this sensitivity is manifest only in the context of cross-linking of surface Ig. Indeed, targeting of liposomes and the same PA-anti-DNP conjugates to I-Ek or H-2Kk, of which the density of expression on B cells is comparable with that of surface Ig, resulted in Ag presentation to a level not greater than that seen for the uptake of soluble HEL by anti-HEL surface Ig. This strongly suggests that binding of Ag to these molecules does not result in signaling that permits up-regulation of costimulatory molecules and access of Ag to class II-containing compartments. The recent development of Abs specific for HEL peptide-IAk conjugates (40, 42) will permit quantification of the actual I-Ak association of Ag presented in monovalent or multivalent forms targeted to different cell surface molecules.
Several studies have emphasized the endocytic capacities of dendritic cells (9, 10). The liposomes we use are small enough to be taken up by fluid phase endocytosis. Free HEL and nontargeted liposome-encapsulated HEL were presented by dendritic cells equally well at equivalent Ag concentrations, ruling out any protective effect of the liposome membranes with respect to Ag processing. The presentation by dendritic cells of Ag associated with phagocytosis of micrometer-sized latex particles has been shown to be orders of magnitude more efficient than uptake of the same Ag in solution (43). Nonrecognition of the 200 nm DNP-bearing neutral liposomes used here in the absence of a targeting ligand permitted an unambiguous evaluation of the role of defined receptors in the uptake process. These lipid vesicles could be used to bind intact anti-DNP Ab, resulting in Ag presentation by IgG FcR-bearing dendritic cells. Stimulation of Ag-specific T cells by liposome-encapsulated HEL targeted to the FcR of dendritic cells by anti-DNP Abs required 1000-fold less Ag than when the cells were exposed to free HEL or nonopsonized liposomes containing HEL. Ag delivered by liposomes opsonized by PA-anti-DNP conjugates in the presence of Abs specific for class I or nonrestricting class II molecules on dendritic cells also resulted in presentation of Ag, resulting in IL-2 production at low Ag concentrations. This contrasts with the reduced efficacy of Ag presentation from liposomes targeted via class I and II molecules on B cells, as compared with surface Ig-targeted liposomes. The fact that we targeted identical class I and II molecules on B and dendritic cells rules out any possible effects of the affinity of receptor-ligand interactions on processing of the liposome-encapsulated Ag. On the other hand, this study compares B cells directly obtained from the spleen with dendritic cells maintained in culture. The phenotype of cultured dendritic cells may change rapidly, with down-regulation of Fc receptors and up-regulation of class II molecules, although the culture conditions used here are reported to maintain an immature phenotype (44). The respective roles of different dendritic cell surface molecules in Ag presentation in vivo remain to be tested.
B cells derived from mice in which the class II-associated invariant chain has been deleted cannot present class II-restricted Ag, while dendritic cells from the same mice present these Ags normally (24). These results indicate that Ag-processing pathways of dendritic and B cells differ. Recent studies indicate that cross-linking of surface Ig may alter intracellular transport of class II molecules and increase its association with endosomal compartments to which Ag is transported by surface Ig (45, 46). The experiments reported here suggest that Ags targeted to surface Ig on B cells are efficiently presented as a consequence of alteration of transport induced by Ig cross-linking. The FcR serves a similar role on dendritic cells, but its participation appears less predominant for dendritic cells than that of surface Ig for B cells. The function of dendritic cells as "sentinels" exposes them to infection by numerous microorganisms. This parasitism of APC is useful for host defense, provided that the act of entering the dendritic cell results in T cell activation. The capacity of dendritic cells to present Ags that cross-link molecules, including MHC class I and II molecules that are not nominal cell surface Ag receptors, may counter the potential use of these molecules as receptors by microorganisms. While the presence of immunoreceptor tyrosine-associated motifs in molecules associated with both surface Ig on B cells and the FcR of dendritic cells suggests common downstream pathways potentially responsible for both efficient presentation and activation (47), the mechanisms permitting Ag presentation or activation via other molecules on dendritic cells remain to be determined.
The question as to whether B cells can initiate immune responses, or are better or worse as APC for T cells than dendritic cells, has been long posed. Early studies indicating that B cells cannot initiate responses are based on experiments in which surface Ig is not stimulated at all, as when exogenous or constitutive peptides are presented (48, 49). Subsequent studies indicated that this presentation defect could be partially overcome by monovalent Ag (22). In the present study the efficiency of Ag presentation by B cells the surface Ig of which was targeted by multivalent Ag was further augmented by orders of magnitude. Furthermore, our data indicate that contact with multivalent Ag may be as important for Ag presentation by dendritic cells as it is for Ag-specific B cells. Thus, these results suggest that when access to Ag is assured, Ag-specific B cells and dendritic cells may have equivalent capacity for Ag presentation to Ag-specific CD4+ T cells. In fact, presentation of Ag from relevant pathogens may be a cooperative process between B cells and dendritic cells under physiologic conditions, since by secreting Ab into the circulation, B cells can permit dendritic cells to identify Ag via their Fc receptors. The requirement that Ab pre-exist to promote processes that augment production of Abs of equivalent specificity does not necessarily pose a dilemma because of the neonatal availability of IgG Ab of maternal origin, as also suggested by others (31). Maternal Ab will permit dendritic cells to present environmental Ags, increasing the frequency of Ag-specific T cells and compensating for the rarity of Ag-specific B cells.
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Acknowledgments |
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Footnotes |
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2 Current address: Laboratory of Theoretical and Physical Biology, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892.
3 Address correspondence and reprint requests to Dr. Lee Leserman, CIML, Case 906, 13288 Marseille Cedex 9, France. E-mail address:
4 Abbreviations used in this paper: HEL, hen egg lysozyme; FcR, Fc receptor; PA-anti-DNP, protein A anti-dinitrophenyl Ab conjugate; CF, carboxyfluorescein.
5 F. Forquet, N. Barois, P. Machy, J. Trucy, V. S. Zimmermann, L. Leserman, and J. Davoust. Submitted for publication.
Received for publication February 19, 1998. Accepted for publication August 10, 1998.
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V. Buatois, M. Baillet, S. Becart, N. Mooney, L. Leserman, and P. Machy MHC Class II-Peptide Complexes in Dendritic Cell Lipid Microdomains Initiate the CD4 Th1 Phenotype J. Immunol., December 1, 2003; 171(11): 5812 - 5819. [Abstract] [Full Text] [PDF]
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 U. Wahl, E. Nossner, K. Kronenberger, R. Gangnus, H. Pohla, M. S. Staege, H.-J. Kolb, M. Hallek, and R. Mocikat Vaccination against B-cell Chronic Lymphocytic Leukemia with Trioma Cells: Preclinical Evaluation Clin. Cancer Res., September 15, 2003; 9(11): 4240 - 4246. [Abstract] [Full Text] [PDF]
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 C. Arigita, L. Bevaart, L. A. Everse, G. A. Koning, W. E. Hennink, D. J. A. Crommelin, J. G. J. van de Winkel, M. J. van Vugt, G. F. A. Kersten, and W. Jiskoot Liposomal Meningococcal B Vaccination: Role of Dendritic Cell Targeting in the Development of a Protective Immune Response Infect. Immun., September 1, 2003; 71(9): 5210 - 5218. [Abstract] [Full Text] [PDF]
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 K. Kronenberger, A. Dieckmann, M. Selmayr, J. Strehl, U. Wahl, H. Lindhofer, G. Kraal, and R. Mocikat Impact of the lymphoma idiotype on in vivo tumor protection in a vaccination model based on targeting antigens to antigen-presenting cells Blood, February 15, 2002; 99(4): 1327 - 1331. [Abstract] [Full Text] [PDF]
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P. Machy, K. Serre, M. Baillet, and L. Leserman Induction of MHC Class I Presentation of Exogenous Antigen by Dendritic Cells Is Controlled by CD4+ T Cells Engaging Class II Molecules in Cholesterol-Rich Domains J. Immunol., February 1, 2002; 168(3): 1172 - 1180. [Abstract] [Full Text] [PDF]
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 A. Rivera, C.-C. Chen, N. Ron, J. P. Dougherty, and Y. Ron Role of B cells as antigen-presenting cells in vivo revisited: antigen-specific B cells are essential for T cell expansion in lymph nodes and for systemic T cell responses to low antigen concentrations Int. Immunol., December 1, 2001; 13(12): 1583 - 1593. [Abstract] [Full Text] [PDF]
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G. G. Chikh, S. Kong, M. B. Bally, J.-C. Meunier, and M.-P. M. Schutze-Redelmeier Efficient Delivery of Antennapedia Homeodomain Fused to CTL Epitope with Liposomes into Dendritic Cells Results in the Activation of CD8+ T Cells J. Immunol., December 1, 2001; 167(11): 6462 - 6470. [Abstract] [Full Text] [PDF]
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Z. You, J. Hester, L. Rollins, G. C. Spagnoli, P. van der Bruggen, and S.-Y. Chen A Retrogen Strategy for Presentation of an Intracellular Tumor Antigen as an Exogenous Antigen by Dendritic Cells Induces Potent Antitumor T Helper and CTL Responses Cancer Res., January 1, 2001; 61(1): 197 - 205. [Abstract] [Full Text]
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T. Keler, P. M. Guyre, L. A. Vitale, K. Sundarapandiyan, J. G. J. van de Winkel, Y. M. Deo, and R. F. Graziano Targeting Weak Antigens to CD64 Elicits Potent Humoral Responses in Human CD64 Transgenic Mice J. Immunol., December 15, 2000; 165(12): 6738 - 6742. [Abstract] [Full Text] [PDF]
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R. Ignatius, K. Mahnke, M. Rivera, K. Hong, F. Isdell, R. M. Steinman, M. Pope, and L. Stamatatos Presentation of proteins encapsulated in sterically stabilized liposomes by dendritic cells initiates CD8+ T-cell responses in vivo Blood, November 15, 2000; 96(10): 3505 - 3513. [Abstract] [Full Text] [PDF]
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Z. You, X. F. Huang, J. Hester, L. Rollins, C. Rooney, and S.-Y. Chen Induction of Vigorous Helper and Cytotoxic T Cell as well as B Cell Responses by Dendritic Cells Expressing a Modified Antigen Targeting Receptor-Mediated Internalization Pathway J. Immunol., October 15, 2000; 165(8): 4581 - 4591. [Abstract] [Full Text] [PDF]
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S. Corinti, D. Medaglini, C. Prezzi, A. Cavani, G. Pozzi, and G. Girolomoni Human Dendritic Cells Are Superior to B Cells at Presenting a Major Histocompatibility Complex Class II-Restricted Heterologous Antigen Expressed on Recombinant Streptococcus gordonii Infect. Immun., April 1, 2000; 68(4): 1879 - 1883. [Abstract] [Full Text] [PDF]
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V. S. Zimmermann, P. Rovere, J. Trucy, K. Serre, P. Machy, F. Forquet, L. Leserman, and J. Davoust Engagement of B Cell Receptor Regulates the Invariant Chain-Dependent MHC Class II Presentation Pathway J. Immunol., March 1, 1999; 162(5): 2495 - 2502. [Abstract] [Full Text] [PDF]
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