The JI
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     
 

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Fish, S. C.
Right arrow Articles by Kasaian, M. T.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Fish, S. C.
Right arrow Articles by Kasaian, M. T.
The Journal of Immunology, 2005, 174: 7716-7724.
Copyright © 2005 by The American Association of Immunologists

IgE Generation and Mast Cell Effector Function in Mice Deficient in IL-4 and IL-13

Susan C. Fish, Debra D. Donaldson1, Samuel J. Goldman, Cara M. M. Williams and Marion T. Kasaian2

Wyeth Research, Cambridge, MA 02140


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
 References
 
IL-4 and IL-13 are potent cytokines that drive production of IgE, which is critical to the development of atopic disease. In this study, we directly compared IgE generation and IgE-dependent mast cell effector function in mouse strains lacking IL-4, IL-13, IL-4 + IL-13, or their common receptor component, IL-4R{alpha}. Although serum IgE was undetectable under resting conditions in most animals deficient in one or both cytokines, peritoneal mast cells from mice lacking IL-4 or IL-13 had only partial reductions in surface IgE level. In contrast, peritoneal mast cells from IL-4/13–/– and IL-4R{alpha}–/– animals were severely deficient in surface IgE, and showed no detectable degranulation following treatment with anti-IgE in vitro. Surprisingly, however, intradermal challenge with high concentrations of anti-IgE Ab induced an ear-swelling response in these strains, implying some capacity for IgE-mediated effector function in tissue mast cells. Furthermore, upon specific immunization with OVA, both IL-4/IL-13–/– and IL-4R{alpha}–/– mice produced detectable levels of serum IgE and Ag-specific IgG1, and generated strong ear-swelling responses to intradermal administration of anti-IgE. These findings suggest that a mechanism for IgE production exists in vivo that is independent of IL-4 or IL-13.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 Disclosures
 References
 
Specific IgE is the underlying requirement for atopic disease, being necessary and sufficient to define atopy. This relationship is underscored by observations that genetic polymorphisms predisposing humans to high serum IgE levels are associated with the development of asthma (1, 2, 3). Several of these polymorphisms affect genes encoding IL-4, IL-13, or their receptor components, attesting to the critical importance of these cytokines in regulating IgE levels in vivo (4, 5, 6, 7, 8, 9, 10, 11, 12, 13). IL-4 and IL-13 are the only agents to date described to induce Ig class switch recombination to IgE. Both cytokines interact with a receptor consisting of IL-4R{alpha} paired with IL-13R{alpha}1. IL-4 may also act through IL-4R{alpha} complexed with the {gamma} common chain. In either case, signal transduction through IL-4R{alpha} results in the phosphorylation, dimerization, and nuclear translocation of STAT-6, which binds to a specific response element in the C{epsilon} promoter region, providing an essential signal for IgE production (14, 15).

Despite their shared receptor and overlapping activities, IL-4 and IL-13 do not appear to be interchangeable in vivo. In studies of murine allergic asthma, IL-13, but not IL-4, was found to be necessary and sufficient for generation of the asthmatic phenotype (16, 17). Although both cytokines are produced by activated Th2 cells, mast cells, and basophils, it is clear that their generation and function can be regulated independently. One major distinction is that T cells lack IL-13R{alpha}1 and thus can respond to IL-4, but not IL-13 (18). B cells respond to both cytokines, and both induce IgE production, but distinctions exist in vivo. Neither IL-4-deficient nor IL-13-deficient mice produce wild-type levels of IgE following immunization with OVA (19, 20, 21) or treatment with anti-IgD (22), suggesting that one cytokine does not fully compensate for lack of the other. In contrast, infection with the murine AIDS (MAIDS)3 virus generates a full IgE response in IL-4-deficient mice (22), indicating that IL-13 preferentially drives IgE production during this infection. Even more intriguing are observations that suggest IgE can be made in the absence of both IL-4 and IL-13, or in the absence of their shared signaling elements. Infection with MAIDS induces IgE in STAT6-deficient mice (23), and IL-4R{alpha}-deficient mice generate a serum IgE response, albeit weak and delayed, following immunization with OVA (19). Based on these observations, it is conceivable that additional pathway(s) exists to generate this important effector molecule.

In addition to that found in the circulation, IgE can exist in tissues, bound to the surface of mast cells. Mast cell-bound IgE has a longer t1/2 than the circulating Ab (24), and can be a more sensitive indicator of bioavailable IgE levels (25). Cross-linking mast cell-bound IgE triggers degranulation and downstream effector function, including anaphylactic reactions. Such responses have been described in mice lacking IL-4R{alpha} (19), indicating that IgE effector function could be elicited in animals that lack responsiveness to either IL-4 or IL-13. Nevertheless, the use of OVA rather than anti-IgE to trigger the response leaves some question as to its IgE dependence. Oettgen et al. (26) showed that anaphylactic responses to Ag can be IgE independent, as they persisted in mice containing a homozygous C{epsilon} null mutation. In contrast, anaphylaxis triggered by anti-IgE was absent. Finkelman and colleagues (27) confirmed these results, showing that in mice lacking IgE or those deficient in Fc{epsilon}RI, Ag-dependent anaphylaxis could still be seen, whereas anti-IgE-dependent anaphylaxis was abolished. Anti-IgE-induced anaphylactic responses were complement independent in both studies (26, 27). To confirm and extend these observations, we now report anti-IgE-induced in vitro and in vivo IgE effector function in mouse strains deficient in IL-4 or IL-13, both IL-4 and IL-13, or the shared IL-4R{alpha} chain.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 Disclosures
 References
 
Mice

BALB/c mice were obtained from The Jackson Laboratory. IL-13R{alpha}2–/– mice were derived by M. Grusby (Harvard School of Public Health Boston, MA) (28). Mice deficient in IL-13 (21) and those lacking both IL-4 and IL-13 (29) were licensed from the Medical Research Council (Cambridge, U.K.), and bred at Taconic Farms. Mice lacking IL-4 (30) were purchased from The Jackson Laboratory. IL-4R{alpha}-deficient mice (31) were obtained through the Taconic Emerging Models Program (Taconic Farms). All mutant strains were on the BALB/c background. Age- and sex-matched animals were used in all experiments. All animal experiments were reviewed and approved by the Institutional Animal Care and Use Committe at Wyeth Research.

ELISA for serum Abs

To assay IgE levels, ELISA plates (MaxiSorp; Nunc) were coated overnight at 4°C with rat anti-mouse IgE (BD Biosciences). Plates were blocked for 1 h at room temperature with 0.5% gelatin in PBS, washed in PBS containing 0.05% Tween 20 (PBS-Tween), and incubated 6 h, room temperature with purified mouse IgE (BD Biosciences) as standard or with serum dilutions. Binding was detected with biotinylated anti-mouse IgE (BD Biosciences) using mouse IgG (Sigma-Aldrich) as a blocker. Binding was detected with peroxidase-linked streptavidin (Southern Biotechnology Associates) and Sure Blue substrate (Kirkegaard & Perry Laboratories). For determination of total IgG1, plates were coated with goat anti-mouse IgG (Sigma-Aldrich), and bound serum Ab detected with biotinylated rat anti-mouse IgG1 (BD Biosciences), using purified mouse IgG1 (BD Biosciences) as standard. For determination of OVA-specific IgE or IgG1, plates were coated with OVA (Sigma-Aldrich). Bound IgE was detected with biotinylated rat anti-mouse IgE (BD Biosciences) in the presence of mouse IgG as a blocking agent, and bound IgG1 was detected with biotinylated rat anti-mouse IgG1 (BD Biosciences).

Peritoneal mast cell degranulation

Peritoneal cavities of mice were lavaged with 5 ml of 25 mM PIPES, pH 7.2, containing 110 mM NaCl, 5 mM KCl, and 50 µM EDTA. Peritoneal cells were pelleted and washed into DMEM containing 10 mM HEPES, 50 µM 2-ME, 10% heat-inactivated FCS, 100 U/ml penicillin, 100 µg/ml streptomycin, 2 mM L-glutamine, and 20 ng/ml mouse rIL-3 (R&D Systems), and incubated overnight at 37°C and 10% CO2. Where indicated, 0.1 µg/ml mouse IgE anti-DNP (Sigma-Aldrich) was added. For [3H]serotonin release, 1 µCi/ml [3H]5-hydroxytryptamine, [{alpha},{beta}-[3H](N)]-creatine sulfate (PerkinElmer Life and Analytical Sciences), was added to the overnight incubation. For the degranulation assay, cells were harvested, washed into pipes, albumin, calcium, magnesium buffer (25 mM PIPES, pH 7.2, containing 110 mM NaCl, 5 mM KCl, 5 mM CaCl2, 2.5 mM MgCl2, and 0.005% BSA), and challenged with the indicated concentration of rat anti-mouse IgE (BD Biosciences), compound 48/80 (Sigma-Aldrich), murine stem cell factor (SCF; R&D Systems), or ionomycin (Sigma-Aldrich) for 30 min at 37°C. Supernatants were collected and frozen. [3H]Serotonin release was determined by liquid scintillation counting. Histamine release was assayed by ELISA (Corixa). Total (maximum) serotonin or histamine was determined by lysing the cells with 1% Triton X-100. Spontaneous release was assayed by challenge with pipes, albumin, calcium, magnsium buffer instead of degranulating agent. The percentage of total release was calculated according to the formula: ((unknown – spontaneous)/(total – spontaneous)) x 100.

Flow cytometry

Peritoneal lavage was harvested, as described above. Cells were washed into PBS containing 1% BSA. In some cases, cells were preincubated for 60 min, 4°C with 5 µg/ml mouse IgE anti-DNP (Sigma-Aldrich). Cells were stained with FITC-labeled Ab to mouse IgE and PE-labeled Ab to mouse c-kit (BD Biosciences) for 30 min on ice. Nonspecific binding was blocked by addition of Fc block (BD Biosciences) and rat IgG (Sigma-Aldrich). Cells were analyzed on a FACScan cytometer (BD Biosciences).

Passive cutaneous anaphylaxis

To induce mast cell degranulation, mice were challenged intradermally in the left ear with varying doses (0.02–100 ng in 20 µl of PBS) of rat anti-mouse IgE (clone R35-72; BD Biosciences). As a control, rat IgG (Sigma-Aldrich) was injected intradermally into the right ear. Ear swelling was measured with an engineer’s micrometer (Mitutoyo) at various time points postchallenge. Swelling was expressed as the change in ear thickness ({Delta}) from baseline preinjection values.

Active cutaneous anaphylaxis

Mice (n = 10/group) were actively sensitized on days 0 and 14 with an i.p. injection of 20 µg of OVA emulsified in 2.25 mg of alum (AlumImject; Pierce) for a total of 200 µl. On day 28, mice were challenged intradermally in the left ear with 10 ng of anti-mouse IgE for a total volume of 20 µl. As a control, mice received 10 ng of rat IgG (Sigma-Aldrich) into the right ear. Ear swelling was measured at various time points postchallenge, as described above, for the (PCA) studies. To assess serum IgE levels, blood was collected by retro-orbital puncture on days 0 and 28.

Tissue staining and confocal microscopy

Ears of mice were harvested, snap frozen in liquid nitrogen in OCT freezing medium (Tissue Tek), and stored at –80°C. Cryostat sections (4–5 µm) were fixed in acetone, air dried, then incubated for 1 h at room temperature with PBS containing 1% BSA and PE-labeled Ab to c-kit (BD Biosciences). Rat IgG (Sigma-Aldrich) and Fc block (BD Biosciences) were included as blocking agents, and biotinylated anti-VCAM (BD Biosciences) and allophycocyanin-streptavidin (Molecular Probes) were used as counterstains. Sections were fixed with 1% paraformaldehyde in PBS, and coverslips were mounted with ProLong antifade agent (Molecular Probes). Fluorescence was analyzed using a Nikon TE200 microscope with Radiance 2000 confocal system (Bio-Rad-Zeiss).


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 Disclosures
 References
 
Mice deficient in IL-4 and/or IL-13 lack serum IgE

IL-4 and IL-13 are critical cytokines for IgE production in response to Ag challenge (14, 15). To investigate the requirement for these cytokines to support resting IgE levels in naive mice, serum was examined from wild-type mice and from mice genetically deficient in IL-4, IL-13, IL-4 + IL-13, IL-4R{alpha}, and IL-13R{alpha}2. Mice deficient in IL-4, IL-13, both IL-4 and IL-13, or their common receptor component, IL-4R{alpha}, had greatly reduced or undetectable levels of serum IgE (Fig. 1) (21, 29, 32). For each of these strains, there was a distribution of serum IgE levels, such that some individual animals had detectable IgE, while the majority did not. In contrast, mice lacking the decoy receptor, IL-13R{alpha}2 (18), displayed elevated levels of serum IgE, in agreement with previous studies (28).



View larger version (17K):
[in this window]
[in a new window]
  		FIGURE 1. Mice lacking IL-4, IL-4/IL-13, or IL-4R have reduced serum IgE and IgG1 levels. Serum IgE and IgG1 concentrations were determined in naive BALB/c wild-type mice and in mutant strains lacking IL-4, IL-13, IL-4/13, IL-4R{alpha}, and IL-13R{alpha}2, as described in Materials and Methods. A, Serum IgE levels, determined from 35–50 mice of each strain. The broken line indicates the limit of assay sensitivity, 2.5 ng/ml. B, Total serum IgG1 levels, determined from 25 mice of each strain. Each point represents the mean of three separate determinations for an individual mouse. Horizontal bars represent the median value for each group. Values of p were determined by Student’s t test.

 
In addition to IgE, IL-4 acts as a switch factor for IgG1 (33, 34), although the dependence on IL-4 is not as strict as for IgE (35). Therefore, we examined IgG1 levels in naive mice. Mice lacking IL-4, IL-4/IL-13, or IL-4R{alpha} were all deficient in circulating IgG1 levels (Fig. 1B). Interestingly, IL-13–/– animals exhibited normal levels of serum IgG1 (Fig. 1B) combined with a profound deficiency in IgE (Fig. 1A), indicating that IL-13 may not fully share the IgG1 switch—promoting activity of IL-4.

Mice deficient in IL-4 or IL-13 display mast cell-bound IgE

Mast cells isolated from the peritoneal cavity of wild-type mice have high levels of cell surface IgE, most likely bound to Fc{epsilon}RI, and detected by staining the cells with anti-IgE Ab. They also express some level of unoccupied Fc{epsilon}RI, which can be assayed by incubating the mast cells with exogenous IgE. Comparison of anti-IgE staining on mast cells in the presence or absence of exogenous IgE indicates the presence of free IgE receptors that are available to be occupied (Fig. 2A). A large body of data has shown that both the proportion of occupied receptors and the overall level of receptor expression can be modulated by IgE exposure (36, 37, 38, 39), and thus will reflect mast cell accessibility to IgE in vivo.



View larger version (26K):
[in this window]
[in a new window]
  		FIGURE 2. Peritoneal mast cells from mice lacking IL-4 or IL-13 have reduced levels of cell surface-bound IgE. Peritoneal lavage was collected from BALB/c wild-type mice and from mutant strains lacking IL-4, IL-13, IL-4/13, IL-4R{alpha}, and IL-13R{alpha}2. Untreated cells and cells loaded with exogenous IgE were stained with FITC-labeled Ab to mouse IgE and PE-labeled Ab to c-kit. Mast cells were identified by forward and side scatter profile in combination with high intensity staining for c-kit, and the mean fluorescence intensity (MFI) of anti- IgE binding was determined by flow cytometry. A, Histograms showing binding of FITC-labeled anti- IgE by pooled cells from the indicated mouse strain, with or without preloading with exogenous IgE. B, MFI of anti- IgE binding to peritoneal mast cells of each strain. Each point represents an individual mouse. The broken line indicates MFI produced by binding an isotype control Ab to the cells. Values of p were determined by Student’s t test. C, Average MFI (+SEM) for anti- IgE binding to individual mice in each strain. {cjs2108}, No added IgE; {blacksquare}, preloaded with exogenous IgE.

 
Peritoneal mast cells from wild-type and IL-13R{alpha}2–/– mice were found to have a high density of surface IgE receptor, which was almost fully occupied with endogenous IgE (Fig. 2, A and B). Peritoneal mast cells from IL-4–/– and IL-13–/– mice had intermediate levels of mast cell-bound IgE, and could capture additional exogenous IgE. Peritoneal mast cells from IL-4/13–/– and IL-4R{alpha}–/– mice had little, if any, surface IgE staining above the negative control level, and a large proportion of empty Fc{epsilon}RI, indicated by their capacity to bind exogenous IgE (Fig. 2, A and B). Interestingly, for mice lacking IL-4, IL-13, IL-4/13, or IL-4R{alpha}, total binding capacity following loading with exogenous IgE was highly comparable (Fig. 2C), indicating that the total IgE receptor density does not fall below a minimum level, even with low IgE exposure.

Mice deficient in IL-4 or IL-13 have normal numbers of peritoneal mast cells

In addition to IgE receptor expression, it has been proposed that the level of circulating IgE modulates mast cell number in vivo by affecting cell survival (40, 41). Mast cells were quantitated as a percentage of total peritoneal leukocytes in wild-type and mutant mice. Total cell numbers recovered in peritoneal lavage, as well as the percentages of lymphocytes and monocytes, were comparable for all groups (data not shown). IL-13R{alpha}2–/– mice had an increased percentage of mast cells in the peritoneal lavage (Fig. 3). IL-4–/– mice also had a higher percentage of peritoneal mast cells (Fig. 3), in agreement with findings reported by Chen et al. (42). IL-13–/–, IL-4/13–/–, and IL-4R{alpha}–/– strains all had peritoneal mast cell frequency comparable with wild type (Fig. 3). Thus, a compensatory increase in mast cell number was not a general feature of those strains that had impaired IgE responses.



View larger version (15K):
[in this window]
[in a new window]
  		FIGURE 3. IL-13R{alpha}2–/– and IL-4–/– strains have increased numbers of mast cells in peritoneal lavage. Peritoneal lavage was collected from BALB/c wild-type mice and from mutant strains lacking IL-4, IL-13, IL-4/13, IL-4R{alpha}, and IL-13R{alpha}2. Cells were stained with PE-labeled Ab to c-kit. Mast cells were identified by forward and side scatter profile in combination with high intensity staining for c-kit, and enumerated by flow cytometry as a percentage of total cells in peritoneal lavage. Each point represents an individual mouse. Values of p were determined by Student’s t test.

 
Peritoneal mast cells from mice deficient in IL-4 or IL-13 show IgE-dependent degranulation in vitro

Peritoneal mast cells can be degranulated in vitro by treatment with anti-IgE to cross-link IgE bound to Fc{epsilon}RI. As histamine releasability has been shown to correlate with the number of IgE molecules on mast cells (43), this response was compared with or without exposure of the cells to exogenous IgE in vitro. Maximum release was quantitated following solubilization of the cell membranes with Triton X-100. Wild-type mast cells achieved 22% maximum degranulation with 1000 ng/ml anti-IgE, which increased to 30% when the cells were preloaded with IgE (Fig. 4). Consistent with their higher levels of bound IgE (Fig. 2), mast cells from IL-13R{alpha}2–/– mice reached 30% maximum degranulation with 1000 ng/ml anti-IgE, which increased to 40% upon loading with exogenous IgE (Fig. 4). In contrast, mice lacking either IL-4 or IL-13 had reduced degranulation responses, achieving only 8 and 18% maximum degranulation, respectively, after challenge with 1000 ng/ml anti-IgE, even after exposure to exogenous IgE (Fig. 4). Peritoneal mast cells isolated from mice lacking both IL-4 and IL-13, or those lacking IL-4R{alpha}, degranulated only after IgE was added exogenously to fill empty receptors (Fig. 4), reaching 6 and 2% maximum degranulation, respectively, with 1000 ng/ml anti-IgE. Even though they showed a low level of degranulation in response to anti-IgE, peritoneal mast cells from IL-4/IL-13- or IL-4R{alpha}-deficient mice did not have a general defect in degranulation, as they displayed normal responses to the IgE-independent stimuli compound 48/80, ionomycin, and SCF (Fig. 5). Table I shows that the levels of total and spontaneous degranulation did not differ significantly between wild-type and cytokine-deficient strains.



View larger version (26K):
[in this window]
[in a new window]
  		FIGURE 4. Peritoneal mast cells from mice lacking IL-4 or IL-13 are deficient in degranulation with anti- IgE. Peritoneal lavage was collected from BALB/c wild-type mice and from mutant strains lacking IL-4, IL-13, IL-4/13, IL-4R{alpha}, and IL-13R{alpha}2. Lavage from four to eight individual mice was pooled for each strain. Untreated cells (•) or cells loaded with exogenous IgE ({circ}) were challenged with the indicated concentration of anti- mouse IgE. Degranulation was measured, as described in Materials and Methods. Data are shown as mean ± SD for four replicate challenge cultures per point.

 


View larger version (25K):
[in this window]
[in a new window]
  		FIGURE 5. Peritoneal mast cells of all mutant strains show full degranulation responses to IgE-independent agents. Peritoneal lavage was collected from BALB/c wild-type mice and from mutant strains lacking IL-4, IL-13, IL-4/13, IL-4R{alpha}, and IL-13R{alpha}2. Lavage from four to eight individual mice was pooled for each strain. Cells were challenged with the indicated concentration of: A, compound 48/80; B, ionomycin; and C, SCF. Data are shown as mean ± SD for four replicate challenge cultures per treatment.

 

View this table:
[in this window]
[in a new window]
  		Table I. Levels of total and spontaneous [3H]serotonin release from peritoneal cellsa

 
Mice deficient in IL-4/IL-13 or IL-4R{alpha} exhibit IgE-dependent PCA

To examine the influence of IL-4 and IL-13 on IgE effector function in vivo, PCA was induced in wild-type and mutant mice. Injection of anti-IgE, but not control Ab, into the ears of naive wild-type mice induced a swelling response that was strong at 1 h, and persisted for over 6 h. The induction of this response with anti-IgE ensured that a mast cell-dependent event was being studied. Compared with wild-type controls, IL-13R{alpha}2–/– mice displayed enhanced PCA responses (Fig. 6A), consistent with their high levels of serum IgE (Fig. 1), mast cell-bound IgE (Fig. 2), and IgE-mediated mast cell degranulation (Fig. 4). A dose titration revealed that a ~10-fold lower concentration of anti-IgE was required to elicit a comparable ear-swelling response in IL-13R{alpha}2–/– mice as compared with wild-type controls (Fig. 6B).



View larger version (22K):
[in this window]
[in a new window]
  		FIGURE 6. IL-4/13–/– and IL-4R{alpha}–/– mice are deficient in anti- IgE-induced PCA. Naive mice from the indicated strain were challenged intradermally in the right ear with anti- mouse IgE Ab (rat IgG1), and ear thickness was determined 1 h later. Mice were challenged in the left ear with a control rat IgG1 of irrelevant specificity, which produced no detectable ear swelling. A, Ear thickness at 1 h postchallenge with 10 ng of anti- IgE, with prechallenge values subtracted. B, Anti-IgE dose response for PCA of naive mice from the indicated strain. Values shown are mean ± SD for five animals per group.

 
Surprisingly, despite their lack of serum IgE and low levels of peritoneal mast cell-bound IgE, mice lacking either IL-4 or IL-13 displayed full ear-swelling responses as compared with wild-type controls, using a 10 ng/ear challenge dose of anti-IgE (Fig. 6A). These observations suggest that the ear-swelling response is induced by very low levels of mast cell-bound IgE. Animals deficient in both IL-4 and IL-13, however, failed to show a PCA response at the 10 ng/ear dose and required 100 ng/ear anti-IgE to elicit detectable ear swelling above background (Fig. 6B). We estimate that >100-fold higher concentration of anti-IgE was required to elicit ear-swelling responses in IL-4/13–/– or IL-4R{alpha}–/– mice comparable to those seen in wild-type controls (Fig. 6B). Staining of ear tissue sections with PE-labeled Ab to c-kit revealed that tissue mast cell content was comparable in wild-type, IL-4/13–/–, and IL-4R{alpha}–/– mice (21.1 ± 4.2, 23.6 ± 6.7, and 22.5 ± 5.8 mast cells per field, repectively, at x20 magnification; n = 12 fields/strain), arguing against the possibility that low responses were due to decreased mast cell numbers in the IL-4/13–/– or IL-4R{alpha}–/– strains. Because the response of IL-4/13–/– and IL-4R{alpha}–/– mice required such high doses of anti-IgE injected into the ear, however, we cannot rule out that the ear swelling was due to mast cell degranulation resulting from complement activation, as opposed to an IgE receptor-mediated mechanism. Nevertheless, ear swelling above background was seen in each of four separate experiments with these strains.

Mice deficient in IL-4/IL-13 or IL-4R{alpha} have enhanced IgE responses following immunization

To examine IgE responses following Ag exposure in wild-type, IL-4/13–/–, and IL-4R{alpha} –/– mice, animals were immunized and boosted with OVA. Immunization increased total IgE levels in wild-type mice ~100-fold (Fig. 7A). Serum IgE levels in IL-4/13–/– and IL-4R{alpha}–/– strains also increased, to detectable levels in all animals (Fig. 7A). Furthermore, OVA-specific IgG1 was detectable in both mutant strains (Fig. 7B), confirming their Ag responsiveness. OVA-specific IgE could also be found in sera from wild-type and IL-4R{alpha}–/– mice following immunization, although it was undetectable in sera of IL-4/13–/– animals (Fig. 8). Because detection of IgE activity in the in vivo PCA response proved to be much more sensitive than serum IgE measurements by ELISA, we also examined effects of OVA immunization on the anti-IgE-induced ear swelling. Immunization with OVA induced a striking increase in ear-swelling responses to anti-IgE in IL-4/13–/– and IL-4R{alpha}–/– mice, such that challenge with 10 ng of Ab elicited a strong PCA reaction (Fig. 9). Thus, a modest increase in serum IgE levels was associated with a strong boost in IgE-dependent ear-swelling responses in IL-4/13–/– and IL-4R{alpha}–/– mice.



View larger version (16K):
[in this window]
[in a new window]
  		FIGURE 7. OVA immunization induces serum IgE and OVA-specific IgG1 in IL-4/13–/– and IL-4R{alpha}–/– mice. BALB/c wild-type, IL-4/13–/–, and IL-4R{alpha}–/– mice were immunized i.p. with OVA in alum on day 0, then boosted on day 14. Sera were collected on day 22. A, Total serum IgE levels, in ng/ml. B, OVA-specific IgG1 levels, expressed as serum dilution required to reach OD of 0.5 in a specific ELISA. The dashed line represents the limit of assay sensitivity. Each point represents the mean of three separate determinations for an individual mouse. Horizontal bars represent the median value for each group. Values of p were determined by Student’s t test.

 


View larger version (18K):
[in this window]
[in a new window]
  		FIGURE 8. OVA immunization induces OVA-specific IgE in sera of wild-type and IL-4R{alpha}–/– mice. BALB/c wild-type, IL-4/13–/–, and IL-4R{alpha}–/– mice were immunized i.p. with OVA in alum on day 0, then boosted on days 14 and 28. Sera were collected on day 34 and tested for the presence of OVA-specific IgE by ELISA. Filled symbols, sera from individual immune mice. Open symbols, average values for sera of same mice before immunization.

 


View larger version (15K):
[in this window]
[in a new window]
  		FIGURE 9. OVA immunization boosts IgE-dependent PCA reactions in IL-4/13–/– and IL-4R{alpha}–/– mice. BALB/c wild-type (A), IL-4/13 –/– (B), and IL-4R{alpha}–/– (C) mice were immunized i.p. with OVA in alum on day 0, then boosted on day 14. On day 28, mice were challenged in the right ear with 10 ng of anti- mouse IgE Ab (rat IgG1) and in the left ear with a control rat IgG1 of irrelevant specificity. Ear thickness was measured 1 h later, and prechallenge values were subtracted. Values shown are mean ± SEM for five animals per group.

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
Disclosures
 References
 
The cytokines IL-4 and IL-13 are key regulators of B cell switch recombination to IgE. Thus, mice deficient in either IL-4 or IL-13 are impaired in serum IgE production following OVA immunization (20, 21), Nippostrongylus brasiliensis infection (21), infection with Plasmodium chabaudi (44), or treatment with anti-IgD (22), indicating that both cytokines normally contribute to IgE production in vivo. Mice deficient in both IL-4 and IL-13, those lacking IL-4R{alpha}, and those deficient in STAT6 would be expected to lack IgE completely, and most studies investigating serum IgE levels support this prediction (29, 45, 46, 47). Surprisingly, however, IgE has been detected in IL-4R{alpha} knockout mice following immunization with OVA (19) and in STAT6-deficient animals infected with MAIDS (23), suggesting that under some conditions, IgE could be produced independently of IL-4 and IL-13.

Because mast cell-bound IgE could be a more sensitive indicator of IgE production in vivo than serum IgE levels (25), we investigated mast cell-mediated IgE effector function in mice lacking IL-4 and/or IL-13, or those lacking the shared IL-4R{alpha} chain. Although the presence of either IL-4 or IL-13 alone was not sufficient to maintain wild-type levels of serum IgE, peritoneal mast cells recovered from IL-4–/– or IL-13–/– animals had detectable IgE bound to the cell surface. In contrast, mice deficient in both IL-4 and IL-13, or in IL-4R{alpha}, had no detectable peritoneal mast cell-bound IgE. If challenged with high levels of anti-IgE in the ear, however, both IL-4/13–/– and IL-4R{alpha}–/– mice exhibited a PCA response. Thus, PCA was the most sensitive indicator of IgE responsiveness, and implied that some level of IgE exists, even in the absence of IL-4 or IL-13 function.

Analysis of peritoneal mast cells from IL-4–/– and IL-13–/– animals revealed that IgE could be bound to the cell surface even when no IgE was detectable in serum, as has been noted previously (25). In serum, the t1/2 of IgE is only 8–12 h (48, 49), but in tissues, it extends to ~6 days (24). IgE bound to peritoneal mast cells was readily detectable in IL-4–/– or IL-13–/– mice, but at levels clearly lower than those found on wild-type mast cells. Although others have failed to detect IgE bound to mast cells of IL-4–/– mice (42, 50), our demonstration of mast cell-bound IgE by flow cytometry is supported by the ability of peritoneal mast cells from IL-4–/– and IL-13–/– animals to degranulate in response to anti-IgE in vitro, and by the induction of a PCA reaction following administration of anti-IgE in these strains.

By adding IgE back to peritoneal mast cells, the surface IgE receptor can be saturated (50). Although this method is not quantitative, it gives an indication of the total surface receptor density and relative levels of occupied vs empty receptors. IgE receptors were almost completely occupied on peritoneal mast cells of wild-type mice. In contrast, for mice lacking IL-4, IL-13, or both, the presence of empty receptors could be demonstrated. Along with decreased receptor occupancy, peritoneal mast cells from these strains had an apparent decrease in IgE receptor expression, as inferred by their reduced capacity to bind exogenous IgE. Thus, low serum IgE levels were associated with low IgE receptor expression. In contrast, the high levels of serum IgE in IL-13R{alpha}2–/– mice were associated with increased mast cell IgE-binding capacity. These findings support the model that exposure to IgE regulates mast cell surface IgE receptor expression (36, 37, 38, 39).

Mice lacking both IL-4 and IL-13, and those lacking IL-4R{alpha}, did not have detectable IgE in the serum or bound to peritoneal mast cells, and showed no peritoneal mast cell degranulation in vitro when challenged with anti-IgE. Therefore, we were surprised to find that anti-IgE elicited a mild PCA reaction even in naive IL-4/13–/– and IL-4R{alpha}–/– mice, although this required >100x higher anti-IgE concentration than that which produced ear swelling in wild-type mice. Thus, it appears that anti-IgE could trigger activation of skin mast cells, but not peritoneal mast cells in these strains. One explanation for these findings is that in vitro determination of histamine or serotonin release following peritoneal mast cell degranulation is simply less sensitive than the in vivo PCA response. It is also possible that tissue mast cells have a lower activation threshold in terms of IgE receptor cross-linking than peritoneal mast cells, or that more IgE is bound to mast cells in skin than to those in the peritoneum. A reliable quantitative comparison could not be made using current methods. Nevertheless, in tissues, mast cells effectively localize and concentrate IgE bound to the high affinity receptor (51, 52), and in this way could maximize effector responses even when low levels of IgE are available.

Furthermore, mast cells in tissues could be exposed to higher levels of IgE than those in the peritoneum, and could derive IgE from sources outside the circulation. It has been suggested that tissue mast cells may acquire IgE directly from lymphatics rather than from the serum (51). In addition, local production of IgE at mucosal and tissue sites has been demonstrated conclusively by the detection of switch circles, which are obligatory intermediates in the switch recombination process (53). Switch recombination occurring at local tissue sites may be driven in part by IL-4 and IL-13 produced by the mast cells themselves (54, 55, 56, 57). IgE produced in a local tissue environment rich in mast cells would most likely bind to those mast cells without ever being released into the circulation. Consistent with this, mast cells in tissues demonstrate IgE-mediated effector function even in the absence of detectable serum IgE (58), and levels of cell-bound IgE are more responsive to allergen challenge or parasite infection than serum IgE (59, 60, 61).

Upon immunization with OVA, total serum IgE became readily detectable in IL-4/13–/– and IL-4R{alpha}–/– mice, and PCA responses were greatly enhanced. The relatively strong induction of anti-IgE-mediated PCA responses in association with only marginal increases in serum IgE following immunization is consistent with observations that even low level IgE receptor occupancy can support the degranulation of basophils (62, 63) or mast cells (43). The increased IgE levels and effector responses following immunization confirm and extend previous reports of OVA-induced anaphylactic responses in IL-4R{alpha}–/– mice (19), by indicating that neither IL-4 nor IL-13 is absolutely required for IgE production or for IgE-dependent mast cell activation in vivo. These observations suggest the existence of an additional pathway to IgE, the exact nature of which remains to be elucidated. IL-4 or IL-13 interacts with a receptor form containing the IL-4R{alpha} chain, triggering the phosphorylation, dimerization, and nuclear translocation of STAT6, which promotes C{epsilon} gene expression and isotype switch recombination to IgE (14, 15). It is conceivable that IgE could be produced in the absence of IL-4R{alpha} if another receptor form could interact with STAT6. Such a model has been proposed by Mattes et al. (64), who reported that IL-13 can promote airways hyperreactivity in mice lacking IL-4R{alpha}, but not in those lacking STAT6. In addition, the finding that IL-15 can trigger STAT6 phosphorylation in mast cells (65) raises the possibility that cytokines other than IL-4 or IL-13 could use STAT6 to drive IgE production. Others have shown that mice lacking STAT6 exhibit various IL-13-mediated allergic-type responses (66) and can even produce IgE under certain activation conditions (23), raising the alternative possibility of a STAT6-independent pathway leading to IgE in mice lacking IL-4 and IL-13. Finally, even though IL-4 and IL-13 may facilitate IgE switch recombination, the event may occur at some low rate even in the absence of these agents.

These findings have important implications for the treatment of IgE-mediated atopic diseases. IL-4 and IL-13 are clearly the most critical inducers of IgE in vivo, and numerous approaches targeting these cytokines are being developed as therapies for asthma and allergy (67). The present results suggest that even in the absence of IL-4 and IL-13, alternative pathways to IgE may exist, enhanced by immunization, and resulting in IgE effector responses in vivo. Additional investigation is required to fully understand the nature of these pathways and their contribution to atopic reactions.


   Acknowledgments
 
We thank Lawrence Mason for technical expertise, and Dr. Joe Sypek for help with coordinating the animal experiments.


   Disclosures
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
References
 
The authors have no financial conflict of interest.


   Footnotes
 
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

1 Current address: Millennium Pharmaceuticals, Inc. Cambridge, MA 02139. Back

2 Address correspondence and reprint requests to Dr. Marion T. Kasaian, Wyeth Research, 200 Cambridge Park Drive, Cambridge, MA 02140. Back

3 Abbreviations used in this paper: MAIDS, murine AIDS; PCA, passive cutaneous anaphylaxis; SCF, stem cell factor. Back

Received for publication December 9, 2004. Accepted for publication April 12, 2005.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
 References
 

  1. Halapi, E., H. Hakonarson. 2004. Recent development in genomic and proteomic research for asthma. Curr. Opin. Pulm. Med. 10: 22-30.[Medline]
  2. Cookson, W. O.. 2002. Asthma genetics. Chest 121: 7S-13S.[Abstract/Free Full Text]
  3. Nanavaty, U., A. D. Goldstein, S. J. Levine. 2001. Polymorphisms in candidate asthma genes. Am. J. Med. Sci. 321: 11-16.[Medline]
  4. Vercelli, D.. 2002. Genetics of IL-13 and functional relevance of IL-13 variants. Curr. Opin. Allergy Clin. Immunol. 2: 389-393.[Medline]
  5. Shirakawa, I., K. A. Deichmann, I. Izuhara, I. Mao, C. N. Adra, J. M. Hopkin. 2000. Atopy and asthma: genetic variants of IL-4 and IL-13 signalling. Immunol. Today 21: 60-64.[Medline]
  6. Tamura, K., M. Suzuki, H. Arakawa, K. Tokuyama, A. Morikawa. 2003. Linkage and association studies of STAT6 gene polymorphisms and allergic diseases. Int. Arch. Allergy Immunol. 131: 33-38.[Medline]
  7. Howard, T. D., G. H. Koppelman, J. Xu, S. L. Zheng, D. S. Postma, D. A. Meyers, E. R. Bleecker. 2002. Gene-gene interaction in asthma: IL-4RA and IL-13 in a Dutch population with asthma. Am. J. Hum. Genet. 70: 230-236.[Medline]
  8. Marsh, D. G., J. D. Neely, D. R. Breazeale, B. Ghosh, L. R. Freidhoff, E. Ehrlich-Kautzky, C. Schou, G. Krishnaswamy, T. H. Beaty. 1994. Linkage analysis of IL-4 and other chromosome 5q31.1 markers and total serum immunoglobulin E concentrations. Science 264: 1152-1156.[Abstract/Free Full Text]
  9. Van der Pouw Kraan, T. C., A. van Veen, L. C. Boeije, S. A. van Tuyl, E. R. de Groot, S. O. Stapel, A. Bakker, C. L. Verweij, L. A. Aarden, J. S. van der Zee. 1999. An IL-13 promoter polymorphism associated with increased risk of allergic asthma. Genes Immun. 1: 61-65.[Medline]
  10. Andrews, R. P., L. Burrell, L. Rosa-Rosa, C. M. Cunningham, J. L. Brzezinski, J. A. Bernstein, G. K. Khurana Hershey. 2001. Analysis of the Ser786Pro interleukin-4 receptor {alpha} allelic variant in allergic and nonallergic asthma and its functional consequences. Clin. Immunol. 100: 298-304.[Medline]
  11. Liu, X., R. Nickel, K. Beyer, U. Wahn, E. Ehrlich, L. R. Freidhoff, B. Bjorksten, T. H. Beaty, S. K. Huang. 2000. An IL-13 coding region variant is associated with a high total serum IgE level and atopic dermatitis in the German multicenter atopy study (MAS-90). J. Allergy Clin. Immunol. 106: 167-170.[Medline]
  12. Graves, P. E., M. Kabesch, M. Halonen, C. J. Holberg, M. Baldini, C. Fritzsch, S. K. Weiland, R. P. Erickson, E. von Mutius, F. D. Martinez. 2000. A cluster of seven tightly linked polymorphisms in the IL-13 gene is associated with total serum IgE levels in three populations of white children. J. Allergy Clin. Immunol. 105: 506-513.[Medline]
  13. Heinzmann, A., X. Q. Mao, M. Akaiwa, R. T. Kreomer, P. S. Gao, K. Ohshima, R. Umeshita, Y. Abe, S. Braun, T. Yamashita, et al 2000. Genetic variants of IL-13 signalling and human asthma and atopy. Hum. Mol. Genet. 9: 549-559.[Abstract/Free Full Text]
  14. Geha, R. S., H. H. Jabara, S. R. Brodeur. 2003. The regulation of immunoglobulin E class-switch recombination. Nat. Rev. Immunol. 3: 721-732.[Medline]
  15. Monticelli, S., D. Vercelli. 2001. Molecular regulation of class switch recombination to IgE through {epsilon} germline transcription. Allergy 56: 270-278.[Medline]
  16. Wills-Karp, M., J. Luyimbazi, X. Xu, B. Schofield, T. Y. Neben, C. L. Karp, D. D. Donaldson. 1998. Interleukin-13: central mediator of allergic asthma. Science 282: 2258-2261.[Abstract/Free Full Text]
  17. Grunig, G., M. Warnock, A. E. Wakil, R. Venkayya, F. Brombacher, D. M. Rennick, D. Sheppard, M. Mohrs, D. D. Donaldson, R. M. Locksley, D. B. Corry. 1998. Requirement for IL-13 independently of IL-4 in experimental asthma. Science 282: 2261-2263.[Abstract/Free Full Text]
  18. Hershey, G. K.. 2003. IL-13 receptors and signaling pathways: an evolving web. J. Allergy Clin. Immunol. 111: 677-690.[Medline]
  19. Grunewald, S. M., M. Teufel, K. Erb, A. Nelde, M. Mohrs, F. Brombacher, E. B. Brocker, W. Sebald, A. Duschl. 2001. Upon prolonged allergen exposure IL-4 and IL-4R{alpha} knockout mice produce specific IgE leading to anaphylaxis. Int. Arch. Allergy Immunol. 125: 322-328.[Medline]
  20. Walter, D. M., J. J. McIntire, G. Berry, A. N. McKenzie, D. D. Donaldson, R. H. DeKruyff, D. T. Umetsu. 2001. Critical role for IL-13 in the development of allergen-induced airway hyperreactivity. J. Immunol. 167: 4668-4675.[Abstract/Free Full Text]
  21. McKenzie, G. J., C. L. Emson, S. E. Bell, S. Anderson, P. Fallon, G. Zurawski, R. Murray, R. Grencis, A. N. McKenzie. 1998. Impaired development of Th2 cells in IL-13-deficient mice. Immunity 9: 423-432.[Medline]
  22. Morawetz, R. A., L. Gabriele, L. V. Rizzo, N. Noben-Trauth, R. Kuhn, K. Rajewsky, W. Muller, T. M. Doherty, F. Finkelman, R. L. Coffman, H. C. Morse, III. 1996. Interleukin (IL)-4-independent immunoglobulin class switch to immunoglobulin (Ig)E in the mouse. J. Exp. Med. 184: 1651-1661.[Abstract/Free Full Text]
  23. Morse, H. C., III, T. McCarty, N. A. Giese, L. Taddesse-Heath, M. J. Grusby. 1999. STAT6-deficient mice exhibit normal induction of murine AIDS and expression of immunoglobulin E following infection with LP-BM5 murine leukemia viruses. J. Virol. 73: 7093-7095.[Abstract/Free Full Text]
  24. Hirano, T., C. Hom, Z. Ovary. 1983. Half-life of murine IgE antibodies in the mouse. Int. Arch. Allergy Appl. Immunol. 71: 182-184.[Medline]
  25. Chen, X. J., U. Wiedermann, U. Dahlgren, L. A. Hanson, L. Enerback. 1995. T-cell-independent and T-cell-dependent IgE responses to the nematode Nippostrongylus brasiliensis: comparison of serum IgE and mast-cell-bound IgE. Immunology 86: 351-355.[Medline]
  26. Oettgen, H. C., T. R. Martin, A. Wynshaw-Boris, C. Deng, J. M. Drazen, P. Leder. 1994. Active anaphylaxis in IgE-deficient mice. Nature 370: 367-370.[Medline]
  27. Strait, R. T., S. C. Morris, M. Yang, X. W. Qu, F. D. Finkelman. 2002. Pathways of anaphylaxis in the mouse. J. Allergy Clin. Immunol. 109: 658-668.[Medline]
  28. Wood, N., M. J. Whitters, B. A. Jacobson, J. Witek, J. P. Sypek, M. Kasaian, M. J. Eppihimer, M. Unger, T. Tanaka, S. J. Goldman, et al 2003. Enhanced interleukin (IL)-13 responses in mice lacking IL-13 receptor {alpha}2. J. Exp. Med. 197: 703-709.[Abstract/Free Full Text]
  29. McKenzie, G. J., P. G. Fallon, C. L. Emson, R. K. Grencis, A. N. McKenzie. 1999. Simultaneous disruption of interleukin (IL)-4 and IL-13 defines individual roles in T helper cell type 2-mediated responses. J. Exp. Med. 189: 1565-1572.[Abstract/Free Full Text]
  30. Noben-Trauth, N., G. Kohler, K. Burki, B. Ledermann. 1996. Efficient targeting of the IL-4 gene in a BALB/c embryonic stem cell line. Transgenic Res. 5: 487-491.[Medline]
  31. Noben-Trauth, N., L. D. Shultz, F. Brombacher, J. F. Urban, Jr, H. Gu, W. E. Paul. 1997. An interleukin 4 (IL-4)-independent pathway for CD4+ T cell IL-4 production is revealed in IL-4 receptor-deficient mice. Proc. Natl. Acad. Sci. USA 94: 10838-10843.[Abstract/Free Full Text]
  32. Hogan, S. P., A. Mould, H. Kikutani, A. J. Ramsay, P. S. Foster. 1997. Aeroallergen-induced eosinophilic inflammation, lung damage, and airways hyperreactivity in mice can occur independently of IL-4 and allergen-specific immunoglobulins. J. Clin. Invest. 99: 1329-1339.[Medline]
  33. Bergstedt-Lindqvist, S., H. B. Moon, U. Persson, G. Moller, C. Heusser, E. Severinson. 1988. Interleukin 4 instructs uncommitted B lymphocytes to switch to IgG1 and IgE. Eur. J. Immunol. 18: 1073-1077.[Medline]
  34. Esser, C., A. Radbruch. 1989. Rapid induction of transcription of unrearranged S{gamma}1 switch regions in activated murine B cells by interleukin 4. EMBO J. 8: 483-488.[Medline]
  35. Mao, C. S., J. Stavnezer. 2001. Differential regulation of mouse germline Ig{gamma}1 and {epsilon} promoters by IL-4 and CD40. J. Immunol. 167: 1522-1534.[Abstract/Free Full Text]
  36. Hsu, C., D. MacGlashan, Jr. 1996. IgE antibody up-regulates high affinity IgE binding on murine bone marrow-derived mast cells. Immunol. Lett. 52: 129-134.[Medline]
  37. Yamaguchi, M., C. S. Lantz, H.C. Oettgen, I. M. Katona, T. Fleming, I. Miyajima, J. P. Kinet, S. J. Galli. 1997. IgE enhances mouse mast cell Fc{epsilon}RI expression in vitro and in vivo: evidence for a novel amplification mechanism in IgE-dependent reactions. J. Exp. Med. 185: 663-672.[Abstract/Free Full Text]
  38. Yamaguchi, M., K. Sayama, K. Yano, C. S. Lantz, N. Noben-Trauth, C. Ra, J. J. Costa, S. J. Galli. 1999. IgE enhances Fc{epsilon} receptor I expression and IgE-dependent release of histamine and lipid mediators from human umbilical cord blood-derived mast cells: synergistic effect of IL-4 and IgE on human mast cell Fc{epsilon} receptor I expression and mediator release. J. Immunol. 162: 5455-5465.[Abstract/Free Full Text]
  39. Borkowski, T. A., M. H. Jouvin, S. Y. Lin, J. P. Kinet. 2001. Minimal requirements for IgE-mediated regulation of surface Fc{epsilon}RI. J. Immunol. 167: 1290-1296.[Abstract/Free Full Text]
  40. Kawakami, T., S. J. Galli. 2002. Regulation of mast-cell and basophil function and survival by IgE. Nat. Rev. Immunol. 2: 773-786.[Medline]
  41. Asai, K., J. Kitaura, Y. Kawakami, N. Yamagata, M. Tsai, D. P. Carbone, F. T. Liu, S. J. Galli, T. Kawakami. 2001. Regulation of mast cell survival by IgE. Immunity 14: 791-800.[Medline]
  42. Chen, X. J., N. Lycke, L. Enerback. 1999. Surface and gene expression of immunoglobulin E receptors on mast cells and mast-cell numbers in interleukin-4-gene knockout mice. Immunology 96: 544-550.[Medline]
  43. Chen, X. J., L. Enerback. 1995. Influence of genetic and environmental factors on surface expression and occupancy of IgE receptors and on histamine releasability of mast cells. Int. Arch. Allergy Immunol. 106: 241-249.[Medline]
  44. Von der Weid, T., M. Kopf, G. Kohler, J. Langhorne. 1994. The immune response to Plasmodium chabaudi malaria in interleukin-4-deficient mice. Eur. J. Immunol. 24: 2285-2293.[Medline]
  45. Brewer, J. M., M. Conacher, C. A. Hunter, M. Mohrs, F. Brombacher, J. Alexander. 1999. Aluminum hydroxide adjuvant initiates strong antigenspecific Th2 responses in the absence of IL-4- or IL-13-mediated signaling. J. Immunol. 163: 6448-6454.[Abstract/Free Full Text]
  46. Mountford, A. P., K. G. Hogg, P. S. Coulson, F. Brombacher. 2001. Signaling via interleukin-4 receptor {alpha} chain is required for successful vaccination against schistosomiasis in BALB/c mice. Infect. Immun. 69: 228-236.[Abstract/Free Full Text]
  47. Kaplan, M. H., U. Schindler, S. T. Smiley, M. J. Grusby. 1996. Stat6 is required for mediating responses to IL-4 and for development of Th2 cells. Immunity 4: 313-319.[Medline]
  48. Haba, S., Z. Ovary, A. Nisonoff. 1985. Clearance of IgE from serum of normal and hybridoma-bearing mice. J. Immunol. 134: 3291-3297.[Abstract]
  49. Vieira, P., K. Rajewsky. 1988. The half-lives of serum immunoglobulins in adult mice. Eur. J. Immunol. 18: 313-316.[Medline]
  50. Banks, E. M., J. W. Coleman. 1996. A comparative study of peritoneal mast cells from mutant IL-4 deficient and normal mice: evidence that IL-4 is not essential for mast cell development but enhances secretion via control of IgE binding and passive sensitization. Cytokine 8: 190-196.[Medline]
  51. Mayrhofer, G.. 1977. Sites of synthesis and localization of IgE in rats infested with Nippostrongylus brasiliensis. Ciba Found. Symp. 46: 155-182.
  52. Lee, T. D., A. Sterk, T. Ishizaka, J. Bienenstock, A. D. Befus. 1985. Number and affinity of receptors for IgE on enriched populations of isolated rat intestinal mast cells. Immunology 55: 363-366.[Medline]
  53. Cameron, L., A. S. Gounni, S. Frenkiel, F. Lavigne, D. Vercelli, Q. Hamid. 2003. S{epsilon}Sµ and S{epsilon}S{gamma} switch circles in human nasal mucosa following ex vivo allergen challenge: evidence for direct as well as sequential class switch recombination. J. Immunol. 171: 3816-3822.[Abstract/Free Full Text]
  54. Pawankar, R., S. Yamagishi, T. Yagi. 2000. Revisiting the roles of mast cells in allergic rhinitis and its relation to local IgE synthesis. Am. J. Rhinol. 14: 309-317.[Medline]
  55. Fokkens, W. J., J. G. Vinke, A. KleinJan. 2000. Local IgE production in the nasal mucosa: a review. Am. J. Rhinol. 14: 299-303.[Medline]
  56. Ghaffar, O., S. R. Durham, K. Al-Ghamdi, E. Wright, P. Small, S. Frenkiel, H. J. Gould, Q. Hamid. 1998. Expression of IgE heavy chain transcripts in the sinus mucosa of atopic and nonatopic patients with chronic sinusitis. Am. J. Respir. Cell Mol. Biol. 18: 706-711.[Abstract/Free Full Text]
  57. Durham, S. R., H. J. Gould, Q. A. Hamid. 1997. Local IgE production in nasal allergy. Int. Arch. Allergy Immunol. 113: 128-130.[Medline]
  58. Coleman, J. W., G. T. Layton, D. R. Stanworth. 1983. The kinetics of in vivo sensitization of rat peritoneal and lung mast cells: temporal dissociation from circulating levels of IgE. Eur. J. Immunol. 13: 994-998.[Medline]
  59. Marcucci, F., L. G. Sensi, E. Migali, G. Coniglio. 2001. Eosinophil cationic protein and specific IgE in serum and nasal mucosa of patients with grass-pollen-allergic rhinitis and asthma. Allergy 56: 231-236.[Medline]
  60. Igarashi, Y., M. S. Goldrich, M. A. Kaliner, A. M. Irani, L. B. Schwartz, M. V. White. 1995. Quantitation of inflammatory cells in the nasal mucosa of patients with allergic rhinitis and normal subjects. J. Allergy Clin. Immunol. 95: 716-725.[Medline]
  61. Negrao-Correa, D., L. S. Adams, R. G. Bell. 1999. Variability of the intestinal immunoglobulin E response of rats to infection with Trichinella spiralis, Heligmosomoides polygyrus or Nippostrongylus brasiliensis. Parasite Immunol. 21: 287-297.[Medline]
  62. MacGlashan, D. W., Jr. 1993. Releasability of human basophils: cellular sensitivity and maximal histamine release are independent variables. J. Allergy Clin. Immunol. 91: 605-615.[Medline]
  63. MacGlashan, D. W., Jr, S. P. Peters, J. Warner, L. M. Lichtenstein. 1986. Characteristics of human basophil sulfidopeptide leukotriene release: releasability defined as the ability of the basophil to respond to dimeric cross-links. J. Immunol. 136: 2231-2239.[Abstract]
  64. Mattes, J., M. Yang, A. Siqueira, K. Clark, J. MacKenzie, A. N. McKenzie, D. C. Webb, K. I. Matthaei, P. S. Foster. 2001. IL-13 induces airways hyperreactivity independently of the IL-4R{alpha} chain in the allergic lung. J. Immunol. 167: 1683-1692.[Abstract/Free Full Text]
  65. Masuda, A., T. Matsuguchi, K. Yamaki, T. Hayakawa, M. Kubo, W. J. LaRochelle, Y. Yoshikai. 2000. Interleukin-15 induces rapid tyrosine phosphorylation of STAT6 and the expression of interleukin-4 in mouse mast cells. J. Biol. Chem. 275: 29331-29337.[Abstract/Free Full Text]
  66. Blease, K., J. M. Schuh, C. Jakubzick, N. W. Lukacs, S. L. Kunkel, B. H. Joshi, R. K. Puri, M. H. Kaplan, C. M. Hogaboam. 2002. Stat6-deficient mice develop airway hyperresponsiveness and peribronchial fibrosis during chronic fungal asthma. Am. J. Pathol. 160: 481-490.[Abstract/Free Full Text]
  67. Chung, K. F.. 2003. Individual cytokines contributing to asthma pathophysiology: valid targets for asthma therapy?. Curr. Opin. Investig. Drugs 4: 1320-1326.[Medline]



This article has been cited by other articles:


Home page
 Am J Trop Med HygHome page
L. M. GANLEY-LEAL, P. N. M. MWINZI, C. B. CETRE-SOSSAH, J. ANDOVE, A. W. HIGHTOWER, D. M. S. KARANJA, D. G. COLLEY, and W. E. SECOR
HIGHER PERCENTAGES OF CIRCULATING MAST CELL PRECURSORS CORRELATE WITH SUSCEPTIBILITY TO REINFECTION WITH SCHISTOSOMA MANSONI
Am J Trop Med Hyg, December 1, 2006; 75(6): 1053 - 1057.
[Abstract] [Full Text] [PDF]

Home page
 Toxicol SciHome page
K. Mizota and H. Ueda
Endocrine Disrupting Chemical Atrazine Causes Degranulation through Gq/11 Protein-Coupled Neurosteroid Receptor in Mast Cells
Toxicol. Sci., April 1, 2006; 90(2): 362 - 368.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Fish, S. C.
Right arrow Articles by Kasaian, M. T.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Fish, S. C.
Right arrow Articles by Kasaian, M. T.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS