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Dalhousie Inflammation Group, Departments of Pathology and Microbiology & Immunology, Dalhousie University, Halifax, Nova Scotia, Canada
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Introduction |
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Langerhans cells (LC) are immature dendritic cells found in the epidermal layer of the skin. They are characterized by the presence of Birbeck granules, expression of high levels of MHC class II, and a characteristic dendritic morphology. Recently, Langerin/CD207, a transmembrane C-type lectin and an unconventional endocytic receptor associated with the formation of the Birbeck granules, has been identified as a LC marker (2). LC act as sentinels for immune responses by capturing Ag in the epidermis in response to appropriate signals and by maturing and migrating into the draining LN where they become very potent in Ag presentation (3, 4). A number of cytokines are important inducers of LC maturation and/or migration, including GM-CSF (5), TNF, and IL-1
(6, 7). Recombinant TNF and IL-1 induce a significant migration of LC from the epidermis and a consequent accumulation of dendritic cells in the draining LN (8, 9, 10).
Mast cells are also present in the skin and are mostly studied in the context of allergy, but also have an important role in host defense. They express high-affinity Fc
RI and IgE-mediated activation leads to the release of multiple preformed, newly formed, and lipid mediators. Preformed mediators, include histamine and granule-associated TNF, which induce dendritic cell maturation (11, 12). Mast cells produce a number of cytokines in the hours following IgE-mediated activation with known effects on dendritic cells including TNF, IL-1, and GM-CSF (13, 14, 15). CD40 ligand is also expressed by mast cells, and is important in regulating dendritic cell migration in vivo (16). Exosomes from IgE-activated human mast cells have recently been shown to induce human dendritic cell maturation in vitro (17). IL-1, TNF, and GM-CSF have all been clearly demonstrated to have the ability to induce dendritic cell maturation in vivo or in vitro (5, 6, 8, 9, 18). A study by Alard et al. (19) suggested that mast cell-derived TNF might inhibit contact sensitization through effects on dendritic cells in mice. A recent report has also demonstrated that mast cell-derived TNF is primarily responsible for LN swelling following bacterial or compound 48/80-induced mast cell activation (20), while histamine did not contribute significantly to this response. Dendritic cells have, however, been shown to express H1, H2, and H3 receptors and to undergo significant maturational changes as a result of histamine treatment (11, 21, 22, 23).
In this study, we examined the role of mast cells in LC migration from the skin to the draining auricular LN using a mouse model. We demonstrate that local IgE-mediated mast cell degranulation provides an effective signal for LC migration to the draining LN in vivo, in the absence of additional local Ag or TLR activator administration.
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Materials and Methods |
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C57BL/6 mice, as well as genetically mast cell-deficient WBB6F1-W/Wv (W/Wv) mice and their normal congenic WBB6F1+/+ (+/+) mice (The Jackson Laboratory, Bar Harbor, ME) were used. All mice were males, 6–10 wk of age. Mice were housed under conventional conditions with food and water provided ad libitum. All experiments were approved by the animal research ethics boards of Dalhousie University.
Induction of local skin reactions
Mice received an intradermal (i.d.) injection into the ear pinna of 0.07 mg of purified mouse anti-trinitrophenyl (TNP) IgE prepared from the B cell line TIB-141 (American Type Culture Collection, Manassas, VA) in 25 µl of PBS, mouse anti-dansyl IgE (BD Pharmingen, San Diego, CA) in 25 µl of PBS, or diluent control (as stated). After a resting period, to allow excess IgE to dissipate (7–14 days), mice were challenged with an i.v. injection of 5 mg/ml TNP-BSA (Biosearch Technologies, Novato, CA) in 250 µl of saline. Mice were sacrificed and their ear pinnae were removed after 18 h or draining auricular LN were excised after 24 h. For experiments using locally mast cell-reconstituted W/Wv mice both ear pinnae were injected with anti-TNP IgE 14 days before i.v. injection of TNP-BSA. In additional experiments, one ear pinna was injected with anti-TNP IgE and the other with anti-dansyl IgE 14 days before i.v. TNP administration. Some animals were also examined 14 days after i.d. IgE injection, but in the absence of i.v. TNP-BSA administration.
Examining the role of histamine receptors
To determine the role of histamine H1 and H2 receptors, 14 days following IgE or control injection in the ear pinnae mice were treated with either the H1 antagonist pyrilamine, 50 mg/kg i.p., or the H2 antagonist cimetidine, 50 mg/kg i.p (Sigma-Aldrich, St. Louis, MO). The doses of histamine receptor antagonists used were based on previous reports of a single dose in vivo used in mice (24, 25, 26). Ninety minutes later the animals were injected i.v. with TNP-BSA and responses were evaluated as described below following harvest of LN cells 24 h later.
Examining the role of serotonin
To determine the role of serotonin (5-hydroxytryptamine), 14 days following IgE or control injection in the ear pinnae mice were injected i.p. with 1.6 mg/kg ketanserin, a 5-hydroxytryptamine receptor antagonist (Research Biochemical International, Oakville, Ontario, Canada). Ninety minutes later the animals were injected i.v. with TNP-BSA and responses were evaluated as described below following harvest of LN cells 24 h later.
Induction of response to LPS
To examine the effect of LPS, mice were injected with 0.25 µg of Escherichia coli LPS serotype 055:B5 (Sigma-Aldrich) in 25 µl of saline in the ear pinna or with diluent control. Draining auricular nodes were examined after 18 h.
Preparation and analysis of epidermal sheets
Epidermal sheets were prepared as previously described by Baker et al. (27). Briefly, mice were sacrificed and their ears were split into dorsal and ventral halves by forceps. The dorsal halves were incubated for 2 h at 37°C with 10 mM disodium ethylenediamine tetraacetic acid (BDH, Toronto, Canada) dissolved in PBS. Ears were washed with PBS and then under a dissecting microscope the epidermis was pealed off using forceps. The epidermis was then fixed in cold acetone (–20°C) for 2 min, washed in PBS, and stored at 4°C until immunohistochemical staining.
Preparation of auricular LN cells
After the mice were challenged with TNP-BSA, the draining (auricular) LNs were excised and, in some experiments (as stated), pooled for each animal. A LN-derived single-cell suspension was prepared, washed, and resuspended in RPMI 1640 medium. Viable cell counts were performed using trypan blue, and the cell concentration was adjusted to 1 x 106 cells/ml. Cytocentrifuge preparations of the LN-derived single-cell suspension were made and cells were stained with anti-Langerin/CD207 or control Ab according to the immunohistochemical procedure described below.
Immunohistochemistry and cell counts
Immunostaining was performed on epidermal skin sheets, LN frozen sections, and cytocentrifuge preparation of LN cell suspensions. Following fixation, sheets or slides were washed with PBS and incubated with 3% BSA (Boehringer Mannheim, Mannheim, Germany) for 1 h. Sheets or slides were then incubated with either anti-Langerin Ab/CD207, clone 959F3 rat IgG1 (DCGM4) (0.75 µg/ml) (courtesy of Dr. S. Saeland, Schering-Plough, Kenilworth, NJ) or in some experiments (as stated) with an anti-mouse MHC class II Ab MRC OX-3 (12.5 µg/ml; Cedarlane Laboratories, Hornby, Ontario, Canada) or an isotype-matched control Ab for 16–18 h at room temperature. Sheets or slides were washed and incubated with biotinylated goat anti-mouse Ab, then detected using streptavidin-HRP (DAKO LSAB-2 System; DakoCytomation, Mississauga, Ontario, Canada) and aminoethylcarbazole substrate (Zymed Laboratories, San Francisco, CA). A randomly chosen 10 fields (0.25 mm2) of epidermal sheets were counted and at least 2000 cells were assessed per slide for examination of cell suspensions. All counts were performed on coded slides with the observer being unaware which slides belonged to which experimental group.
Mast cell reconstitution
Bone marrow-derived mast cells were generated from the bone marrow of WBB6F1+/+ (+/+) mice according to the method of Tertian et al. (28), a minimum of 98% pure cells was used. W/Wv mice were locally reconstituted as previously described (29). Cells derived from +/+ mice (0.5 x 106) were locally injected into the right ear pinna of W/Wv mice and allowed to mature for 10 wk.
Evaluation of anti-TNP Ab responses
Serum Ab responses against TNP were evaluated by ELISA. Briefly, the wells of ELISA plates were coated with TNP-keyhole limpet hemocyanin (Calbiochem, San Diego, CA) and nonspecific binding was blocked by incubation with 1% BSA. A series of dilutions of serum samples from immunized and nonimmunized mice were added to the washed plate and incubated for 16–18 h. Binding of Abs was detected using a biotinylated goat anti-mouse IgG/IgA/IgM Ab (Cedarlane Laboratories) followed by a commercial amplification substrate system (Invitrogen Life Technologies, Grand Island, NY).
Flow cytometry
Cells were washed twice with 2% FBS/PBS, fixed with 1% paraformaldehyde/PBS for 30 min, and washed with 2% FBS/PBS. The cells were permeabilized with 0.1% saponin/2% FBS/PBS for 20 min and stained with anti-Langerin (CD207) Ab, anti-fascin Ab, or an isotype control for 20 min. Following staining the cells were washed twice by gentle shaking in 0.1% saponin/2% FBS/PBS for 5 min before centrifugation. Cells were incubated with the secondary Ab, an FITC-labeled mouse anti-rat IgG Ab, and a PE-labeled goat anti-mouse IgG Ab for 20 min, followed by the same washing procedure. To block free Ab binding sites, the cells were incubated with 5% rat serum and 20 µg/ml hamster IgG. Cells were washed twice with 0.1% saponin/2% FBS/PBS, once with 2% FBS/PBS, and then stained for cell surface markers with a PE-conjugated anti-CD86, CD40, or isotype control as well as an allophycocyanin-conjugated CD11c or isotype control. Finally, the cells were washed twice with 2% FBS/PBS. For flow cytometric analysis, 3 x 104 cells were acquired with a FACSCalibur (BD Biosciences, Mountain View, CA) and analyzed with CellQuest (BD Biosciences). All Abs were purchased from (BD Biosciences) except for anti-fascin, which was purchased from DakoCytomation and the PE-labeled goat anti-mouse IgG Ab which was purchased from Molecular Probes (Leiden, The Netherlands).
Statistical methods
Following confirmation of appropriate data distribution, differences between left and right epidermal sheets or left and right LNs were evaluated using a paired Student’s t test. Differences between groups of animals were assessed using an unpaired Student’s t test.
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Results |
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The consequences of functional c-kit deficiency on the normal distribution of LC in the skin of W/Wv mice were examined to confirm the appropriateness of this mouse model for these studies. The number of Langerin/CD207-positive cells in the epidermis of untreated mast cell-deficient W/Wv mice was compared with those in the wild-type control +/+ mice. No significant differences in LC distribution or morphology were observed (Fig. 1).
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IgE-mediated mast cell activation was used to selectively activate mast cells in one ear pinna. C57BL/6 mice were injected with anti-TNP IgE Ab into the right ear pinna and saline into the left ear pinna 14 days before Ag challenge with an i.v. injection of TNP-BSA. After another 18 h, selected on the basis of pilot experiments, the number of LC in the epidermal sheets was assessed. A significant (p < 0.05) reduction in the number of Langerin/CD207-positive LC was consistently observed in the right (IgE-injected) ears of the mice compared with their left (control) ears (Fig. 2, A and B). These results were confirmed using MHC class II as a second marker of LC (data not shown). The mean reduction in LC in the IgE-injected skin compared with the control skin was 16.4% (Fig. 2C) using Langerin/CD207 staining and 16.7% using MHC class II staining.
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The decrease in LC at skin sites of IgE-mediated activation was mirrored by an increase in the number of Langerin-positive dendritic cells within the draining LNs (Fig. 2, E and F). A 2.3-fold increase (p < 0.05) was observed in the number of Langerin/CD207-positive cells/LN draining sites of IgE-mediated activation compared with control sites (Fig. 2, G and H). The specificity of Langerin/CD207 staining was confirmed using isotype control Ab in place of anti-Langerin/CD207; no positively stained dendritic cells were observed under these conditions (data not shown).
LC migration in W/Wv mast cell-deficient mice
To determine the role of mast cells in IgE-mediated LC migration, both mast cell-deficient W/Wv mice and their wild-type control +/+ mice were injected with anti-TNP IgE Ab and challenged with i.v. Ag as described above. No significant decrease in the numbers of Langerin-positive cells was observed in the IgE-injected ear pinnae of W/Wv mice (Fig. 3A). However, a small but consistent increase in the number of Langerin/CD207-positive LC cells was observed. In control +/+ mice, a 23% decrease in Langerin/CD207-positive LC cells (p < 0.01) was observed in the IgE-injected ear pinna compared with the control site (Fig. 3B). Consistent with these data, only a 25% increase (p < 0.05) in total LN cellularity was observed in W/Wv mice in the LNs draining the IgE-activated ear pinnae compared with controls (Fig. 3C). In contrast, a 2-fold increase (p < 0.0001) in total cellularity was observed in +/+ mice in the LNs draining the IgE-activated side compared with control (Fig. 3D). No significant increase was observed in the numbers of Langerin/CD207-positive cells in the LNs of W/Wv mice (Fig. 3E). However, a 2.5-fold increase (p < 0.01) in the numbers of Langerin/CD207-positive cells was observed in control +/+ mice (Fig. 3F). These observations suggested that the induction of LC migration following local IgE-mediated activation is mast cell dependent.
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LC cells observed in draining LNs have undergone maturation
To examine the maturation status of LC within LNs draining sites of IgE-mediated mast cell activation, a flow cytometric approach was used. The dual Langerin-positive and CD11c-positive cells were examined for their expression of the well-defined maturation markers CD86, fascin, and CD40. LC at these sites were found to be on average (n = 6) 85% CD86 positive, 87% fascin positive, and 45% CD40 positive (data not shown). This marker profile is in keeping with a matured LC phenotype. The marker profile of LC was not significantly different between nodes draining sites of IgE-mediated mast cell activation and control sites.
LC migration in W/Wv mice with local mast cell reconstitution
To more directly address the role of the mast cell, W/Wv mice were locally reconstituted by i.d. injection with bone marrow-derived mast cells in the right ear only. After a 10-wk resting period, to allow mast cells to mature in vivo, mice were injected with anti-TNP IgE Ab in both ear pinnae. One week later, mice were challenged with TNP-BSA Ag i.v. and both epidermal sheets (18 h) and auricular nodes (24 h) were examined for their numbers of Langerin/CD207-positive dendritic cells. Examination of the epidermal sheets from the ear pinnae revealed that Langerin/CD207-positive cell numbers were significantly (p < 0.01) decreased in the mast cell-reconstituted site compared with the nonreconstituted site (Fig. 4A). A significant (p < 0.01) increase in total cellularity was observed in the LNs draining the mast cell-reconstituted ear pinna (Fig. 4B). A significant (p < 0.02) increase in the number of Langerin/CD207-positive dendritic cells/LNs was also observed at this site (Fig. 4C).
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LPS is a known inducer of dendritic cell migration (7) via interactions with TLRs, but does not induce mast cell degranulation (30). Both W/Wv mice and their wild-type control +/+ mice were injected with LPS into their right ear and saline into the left. After 18 h, mice were sacrificed and LNs were examined. In the draining LNs, there was a similar significant (p < 0.01) increase in the number of total cells in the LPS draining node compared with the control in both W/Wv mice (2-fold; Fig. 5A) and control +/+ mice (2.6-fold; Fig. 5B). In addition, the numbers of Langerin/CD207-positive dendritic cells showed a significant (p < 0.05 and p < 0.01) increase in the sites draining LPS-injected skin compared with control in both W/Wv mice (3.4-fold; Fig. 5C) and +/+ mice (4-fold; Fig. 5D).
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To investigate the mechanism of these responses in more detail, we focused on the changes in Langerin-positive cells and total cellularity observed in the auricular LNs draining sites of IgE-mediated mast cell activation. Murine LC do not express FcRI (31); however, they do express the low-affinity FcRII/CD23. There was no significant difference in the number of Langerin/CD207-positive cells found within the LNs draining IgE-injected or control (saline-injected) ear pinnae (Fig. 6, A and B). As further confirmation that the substantial dendritic cell responses observed in the LNs were the result of Ag-specific IgE-mediated mast cell activation, one ear pinna of a group of six C57BL/6 mice was injected i.d. with anti-dansyl IgE while the other was injected with anti-TNP IgE. Fourteen days later, the mice were injected i.v. with TNP-BSA and after 24 h the draining auricular LNs were excised and examined for the number of Langerin/CD207-positive cells. Again, the LNs draining the site of IgE-mediated mast cell activation (injected with anti-TNP IgE) showed significantly increased numbers of Langerin/CD207-positive cells compared with the site that received anti-dansyl IgE injection alone (Fig. 6, C and D).
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In view of the known effects of histamine on dendritic cell maturation and function, the consequences of pharmacologic blockade of either H1 receptors using pyrilamine or H2 receptors using cimetidine were examined. LNs draining sites of IgE-mediated mast cell activation were examined in C57BL/6 mice that had been pretreated with cimetidine, pyrilamine, or diluent (control) injection i.p. 90 min before i.v. Ag challenge of IgE-sensitized mice (anti-TNP IgE 14 days before Ag injection). The numbers of Langerin/CD207-positive cells migrating to the LN draining sites of IgE-mediated activation were similar in diluent-treated animals and following H1 blockade (Fig. 7, B and D). In marked contrast, following H2 blockade, increases in Langerin/CD207-positive cells in the auricular node draining sites of IgE-mediated activation were almost completely blocked (Fig. 7F; 90.5% inhibition compared with diluent-treated control group). The increase in total LN cellularity, as a result of IgE-mediated mast cell activation, was not significantly inhibited by cimetidine pretreatment of the animals. The effects of combined cimetidine and pyrilmaine treatment were similar to those of cimetidine alone (Fig. 7, G and H).
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Serotonin is also found in large amounts in rodent mast cells. To examine the ability of this mediator to participate in the observed LC response, the serotonin receptor antagonist ketanserin was used. Ketanserin was completely ineffective in blocking LC migration in response to mast cell activation when used at in vivo doses that have been shown to be highly effective in blocking other serotonin-dependent events (32). Ketanserin was injected 90 min before Ag challenge, and both the number of total cells and Langerin-positive cells were determined. Total cells in the IgE-injected side were 5.6 x 106 cells vs 3.2 x 106 cells in the control side. Langerin-positive cells were 17 x 103 cells in the IgE-injected side vs 8.3 x 103 cells in the control-injected side with a p < 0.0001.
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Discussion |
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Several different approaches confirmed that changes in LC populations were the result of mast cell activation and not IgE administration alone or IgE-Ag interactions on other cell types. First, when W/Wv mice were selectively reconstituted in one ear pinna with mast cells and then injected in both ear pinnae with IgE, before i.v. Ag challenge, a significant (p < 0.01) difference was observed between the mast cell-containing site and the nonreconstituted site. Both a decrease in Langerin/CD207-positive cells in the epidermal sheets where mast cells were present and an increase in Langerin/CD207-positive cells in the draining auricular LNs were observed when compared with the mast cell-deficient ear sites within the same animals (Fig. 4). Second, we confirmed that IgE injection into the ear pinna alone, in the absence of i.v. Ag challenge, did not induce changes in the number of Langerin/CD207-positive cells within the draining LNs when compared with saline-injected control ears in the same animals (Fig. 6B). As a further control for potential effects of IgE injection alone on LC migration, some animals were injected in both ears with IgEs of differing specificity. When Ag was injected i.v., only the LC populations from sites injected with IgE of appropriate specificity were observed to substantially migrate from the ear epidermis to the draining node (Fig. 6D).
It is notable that the observed changes in Langerin/CD207-positive cells within the epidermal sheets were lower than might have been expected given the substantial increases in Langerin/CD207-positive cells within the draining auricular node. The scale of change in both the epidermal site and the draining node are consistent with that observed by others using direct injection of TNF and/or IL-1 into the skin (9, 10). The small overall reduction in LC numbers may, in part, be explained by recruitment of new LC to the site of mast cell activation (33). Henri et al. (34) have demonstrated that the numbers of Langerin-positive cells in LNs draining the skin are normally regulated by a steady state of migration, consistent with the presence of some Langerin/CD207-positive cells in the control LNs. In addition, Stoitzner et al. (35) have shown that Langerin/CD207-positive cells in the skin-draining LNs are mostly mature. Changes in LC numbers in both the ear epidermis and draining LNs were consistent with IgE-mediated mast cell activation inducing LC migration in the context of a background level of continuous recruitment of immature LC to the skin and subsequent migration of cells to the draining LNs. The appropriateness of the specific auricular node selected for our study was confirmed by pilot experiments in which India ink was injected i.d. into the ear pinnae of mice (data not shown). This enabled us to isolate a single LN for study, reducing any contribution from other tissues to the changes in LC populations we examined. However, we cannot completely rule out the possibility that Langerin/CD207-positive cells from neighboring sites might contribute to the increase in Langerin/CD207-positive cells within the node.
The LC that migrated to draining LNs following IgE-mediated mast cell activation appear to have a similar phenotype to those found normally at this site and to have undergone some degree of maturation. A high percentage of Langerin/CD207-positive cells in these LNs were fascin and CD86 positive, a profile typical of a mature dendritic cell. It has been suggested that as LC mature in the LNs they will lose their expression of Langerin/CD207 so that both the numbers of LC migrating to the draining nodes and the degree of maturity, we are reporting, might be underestimates.
The ability of LPS administration to induce LC migration in both W/Wv and control +/+ mice, to a similar extent, illustrates that mast cell activation is not required for LC migration in response to all stimuli. Although murine mast cells express the TLR4 receptor and have been shown to respond functionally by producing multiple cytokines in response to LPS (36, 37, 38, 39), the mast cell contribution to LC mobilization, under our experimental conditions, was not significant. These data also illustrate that the LC in W/Wv mice are capable of being mobilized normally following appropriate stimulation, further confirming that the functional c-kit deficiency in W/Wv mice has not substantially altered the ability of LC to migrate normally. It is also notable that although changes in auricular LN cellularity in response to IgE/Ag-mediated activation were highly mast cell dependent, LPS injection induced similar changes in LN cellularity in both mast cell-containing and mast cell-deficient mice. These data suggest that mast cell-independent mechanisms for LN activation are mobilized following LPS injection in addition to the mast cell-derived TNF-dependent mechanisms that have been recently described (20).
The mechanism by which mast cells induce LC migration appears to be highly dependent upon histamine and H2 receptors. Our data demonstrate a critical role for H2 receptors in the migration of LC to draining LNs following IgE-mediated mast cell activation. It is notable, however, that H2 blockade was not effective as an inhibitor of mast cell-dependent increases in total LN cellularity. These data are in keeping with recent studies suggesting that this process is mainly TNF dependent (20). H1 blockade did not significantly alter total LN cellularity or LC migration to draining LNs at sites of IgE-mediated activation. However, there was a slight trend toward reduced LC numbers in the activated LNs, which may reflect some cross-inhibition of H2 receptors by the relatively high doses of pyrilamine used in this study. Histamine up-regulates the expression of CD86, CD40, CD54, CD80, and MHC class II, as well as induces proinflammatory cytokine and chemokine production by human immature dendritic cells through H1 and H2 receptors (11). However, histamine alone does not up-regulate CCR7, which is thought to be crucial for dendritic cell migration and can be regulated by IL-1 or TNF (40, 41). Although histamine is clearly required, it is probably not the only necessary mast cell-derived factor involved in the process of mast cell-dependent LC mobilization. Indeed, administration of histamine alone into ear skin did not induce LC migration, suggesting a role for other factors induced from mast cells or neighboring cells as a result of IgE-mediated mast cell activation. One potentially important factor is TNF, which is found both preformed in mast cells and newly synthesized following activation (14). Intradermal administration of TNF induces LC migration and dendritic cell accumulation in the draining LNs in mice (8, 9). Other additional candidate inducers of mast cell-mediated LC migration include IL-1, GM-CSF, and CD40 ligand, which have been implicated in dendritic cell migration or maturation and are expressed by activated human and rodent mast cells (42, 43). Mast cell exosomes may provide many of these mediators, or additional signals and have been shown to up-regulate dendritic cell MHC class II, CD80, CD86, and CD40 molecules. Mast cell exosomes, containing histamine, also induce dendritic cells to become efficient APCs, eliciting both IgG1 and IgG2a Abs in mice following immunization with exosome-associated Ags (17).
Complement activation has also been implicated as an important signal for mast cell activation and subsequent interaction with dendritic cells. Tsuji et al. (44) have shown that C5 is required early for the elicitation phase of contact sensitivity and delayed-type hypersensitivity. C5 was required for macrophage chemotactic activity and the late production of IFN-
. C5a has also been shown to induce serotonin release from mast cells which induces endothelial cells activation and Th1 cells extravasation (45). The design of the current studies largely preclude an important role for complement locally within one ear as compared with the contralateral ear. Mouse IgE, particularly when bound to FcRI, is not considered an effective complement activator. Free IgE has a relatively short (12-h) half-life in rodents and is injected 7–14 days before i.v. Ag administration in our experiments.
The data reported here do not directly implicate mast cells in the process of Ag presentation or contact sensitization, which has previously been implied (46, 47). For example, Bryce et al. (48) have shown that mast cells are required for contact sensitivity and dendritic cell reduction in the epidermis following hapten exposure. However, our results indicate that LC migration to draining LNs can be induced by IgE-mediated mast cell activation. Further signals may then be required to allow such LC to present Ag effectively. The process of mast cell-induced LC migration suggests a potential role in the perpetuation of chronic allergic disease. Once an IgE response to a specific environmental Ag has been established, subsequent exposure may lead to mast cell activation and release of mediators. Potentially, these mediators may enhance the migration of further local LC, carrying further environmental Ags to the draining LNs. This process could increase the possibility that additional IgE responses would occur in response to common environmental Ags. Histamine has been demonstrated to enhance Th2 responses through effects on dendritic cells (21). Our current studies demonstrate that mast cell-derived signals, including histamine, also provide key stimuli for inducing LC migration in vivo. The importance of mast cell-dependent mobilization of LC to both allergic sensitization and the rapid development of effective immunity to pathogens will be important topics for future studies.
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Acknowledgments |
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Footnotes |
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1 This work was supported by the Canadian Institutes for Health Research. D.M.J. is supported by a scholarship from the Cultural Bureau of Saudi Arabia. 
2 Address correspondence and reprint requests to Dr. Jean S. Marshall, Department of Pathology, Sir Charles Tupper Medical Building, Dalhousie University, Halifax, Nova Scotia, B3H 1X5 Canada. E-mail address: Jean.Marshall{at}Dal.ca
3 Abbreviations used in this paper: LN, lymph node; LC, Langerhans cell; i.d., intradermal; TNP, trinitrophenyl.
Received for publication December 19, 2003. Accepted for publication August 17, 2004.
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A. L. Christy and M. A. Brown The Multitasking Mast Cell: Positive and Negative Roles in the Progression of Autoimmunity J. Immunol., September 1, 2007; 179(5): 2673 - 2679. [Abstract] [Full Text] [PDF]
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V. Heib, M. Becker, T. Warger, G. Rechtsteiner, C. Tertilt, M. Klein, T. Bopp, C. Taube, H. Schild, E. Schmitt, et al. Mast cells are crucial for early inflammation, migration of Langerhans cells, and CTL responses following topical application of TLR7 ligand in mice Blood, August 1, 2007; 110(3): 946 - 953. [Abstract] [Full Text] [PDF]
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J. P. McGlade, S. Gorman, J. C. Lenzo, J. W. Tan, T. Watanabe, J. J. Finlay-Jones, W. R. Thomas, and P. H. Hart Effect of Both Ultraviolet B Irradiation and Histamine Receptor Function on Allergic Responses to an Inhaled Antigen J. Immunol., March 1, 2007; 178(5): 2794 - 2802. [Abstract] [Full Text] [PDF]
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 A. Majdalawieh, L. Zhang, and H.-S. Ro Adipocyte Enhancer-binding Protein-1 Promotes Macrophage Inflammatory Responsiveness by Up-Regulating NF-{kappa}B via I{kappa}B{alpha} Negative Regulation Mol. Biol. Cell, March 1, 2007; 18(3): 930 - 942. [Abstract] [Full Text] [PDF]
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A. Mazzoni, R. P. Siraganian, C. A. Leifer, and D. M. Segal Dendritic Cell Modulation by Mast Cells Controls the Th1/Th2 Balance in Responding T Cells J. Immunol., September 15, 2006; 177(6): 3577 - 3581. [Abstract] [Full Text] [PDF]
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T. Satoh, R. Moroi, K. Aritake, Y. Urade, Y. Kanai, K. Sumi, H. Yokozeki, H. Hirai, K. Nagata, T. Hara, et al. Prostaglandin D2 Plays an Essential Role in Chronic Allergic Inflammation of the Skin via CRTH2 Receptor J. Immunol., August 15, 2006; 177(4): 2621 - 2629. [Abstract] [Full Text] [PDF]
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D. M. Jawdat, G. Rowden, and J. S. Marshall Mast Cells Have a Pivotal Role in TNF-Independent Lymph Node Hypertrophy and the Mobilization of Langerhans Cells in Response to Bacterial Peptidoglycan J. Immunol., August 1, 2006; 177(3): 1755 - 1762. [Abstract] [Full Text] [PDF]
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H. Suto, S. Nakae, M. Kakurai, J. D. Sedgwick, M. Tsai, and S. J. Galli Mast Cell-Associated TNF Promotes Dendritic Cell Migration J. Immunol., April 1, 2006; 176(7): 4102 - 4112. [Abstract] [Full Text] [PDF]
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S. Nakae, H. Suto, M. Iikura, M. Kakurai, J. D. Sedgwick, M. Tsai, and S. J. Galli Mast Cells Enhance T Cell Activation: Importance of Mast Cell Costimulatory Molecules and Secreted TNF J. Immunol., February 15, 2006; 176(4): 2238 - 2248. [Abstract] [Full Text] [PDF]
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P. J. O'Connell, X. Wang, M. Leon-Ponte, C. Griffiths, S. C. Pingle, and G. P. Ahern A novel form of immune signaling revealed by transmission of the inflammatory mediator serotonin between dendritic cells and T cells Blood, February 1, 2006; 107(3): 1010 - 1017. [Abstract] [Full Text] [PDF]
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C. E. Demeure, K. Brahimi, F. Hacini, F. Marchand, R. Peronet, M. Huerre, P. St.-Mezard, J.-F. Nicolas, P. Brey, G. Delespesse, et al. Anopheles Mosquito Bites Activate Cutaneous Mast Cells Leading to a Local Inflammatory Response and Lymph Node Hyperplasia J. Immunol., April 1, 2005; 174(7): 3932 - 3940. [Abstract] [Full Text] [PDF]
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