IgE-associated food allergy alters the presentation of paediatric eosinophilic esophagitis.

BACKGROUND: Links between food allergens and eosinophilic esophagitis (EoE) have been established, but the interplay between EoE- and IgE-associated immediate hypersensitivity to foods remains unclear. OBJECTIVE: We sought to determine the prevalence of IgE-associated food allergy at the time of diagnosis of EoE in children and to determine whether differences existed in presentation and disease compared to subjects with EoE alone. METHODS: Eosinophilic esophagitis patients were stratified based on the diagnosis of IgE-associated immediate hypersensitivity (EoE + IH vs. EoE-IH). Clinical, histologic, pathologic, and endoscopic differences were investigated using a retrospective database. RESULTS: We found that 29% of the 198 EoE patients in our cohort had EoE + IH. These subjects presented at a younger age than those without IH (6.05 vs. 8.09 years, P = 0.013) and were more likely to have comorbid allergic disease. Surprisingly, the EoE + IH group presented with significantly different clinical symptoms, with increased dysphagia, gagging, cough, and poor appetite compared to their counterparts in the EoE-IH group. Male gender, allergic rhinitis, the presence of dysphagia, and younger age were independently associated with having EoE + IH. Specific IgE levels to common EoE-associated foods were higher in EoE + IH, regardless of eliciting immediate hypersensitivity symptoms. In contrast, IgE levels for specific foods triggering EoE were relatively lower in both the groups than IgE levels for immediate reactions. CONCLUSIONS AND CLINICAL RELEVANCE: Immediate hypersensitivity is common in children with EoE and identifies a population of EoE patients with distinct clinical characteristics. Our study describes a subtype of EoE in which IgE-mediated food allergy may impact the presentation of paediatric EoE.

The rat c-kit ligand, stem cell factor, induces c-kit receptor-dependent mouse mast cell activation in vivo. Evidence that signaling through the c-kit receptor can induce expression of cellular function.

Interactions between products of the mouse W locus, which encodes the c-kit tyrosine kinase receptor, and the Sl locus, which encodes a ligand for c-kit receptor, which we have designated stem cell factor (SCF), have a critical role in the development of mast cells. Mice homozygous for mutations at either locus exhibit several phenotypic abnormalities including a virtual absence of mast cells. Moreover, the c-kit ligand SCF can induce the proliferation and maturation of normal mast cells in vitro or in vivo, and also can result in repair of the mast cell deficiency of Sl/Sld mice in vivo. We now report that administration of SCF intradermally in vivo results in dermal mast cell activation and a mast cell-dependent acute inflammatory response. This effect is c-kit receptor dependent, in that it is not observed when SCF is administered to mice containing dermal mast cells expressing functionally inactive c-kit receptors, is observed with both glycosylated and nonglycosylated forms of SCF, and occurs at doses of SCF at least 10-fold lower on a molar basis than the minimally effective dose of the classical dermal mast cell-activating agent substance P. These findings represent the first demonstration in vivo that a c-kit ligand can result in the functional activation of any cellular lineage expressing the c-kit receptor, and suggest that interactions between the c-kit receptor and its ligand may influence mast cell biology through complex effects on proliferation, maturation, and function.

Distinct expression and function of the novel mouse chemokine monocyte chemotactic protein-5 in lung allergic inflammation.

We have cloned a novel mouse CC chemokine cDNA from the lung during an allergic inflammatory reaction. The protein encoded by this cDNA is chemotactic for eosinophils, monocytes, and lymphocytes in vitro and in vivo. Based on its similarities in sequence and function with other CC chemokines, we have named it mouse monocyte chemotactic protein-5 (mMCP-5). Under noninflammatory conditions, expression of mMCP-5 in the lymph nodes and thymus is constitutive and is generally restricted to stromal cells. Neutralization of mMCP-5 protein with specific antibodies during an allergic inflammatory reaction in vivo resulted in a reduction in the number of eosinophils that accumulated in the lung. Moreover, mMCP-5 mRNA expression in vivo is regulated differently from that of other major CC chemokines in the lung during the allergic reaction, including Eotaxin. The presence of lymphocytes is essential for expression of mMCP-5 by alveolar macrophages and smooth muscle cells in the lung, and the induction of mMCP-5 RNA occurs earlier than that of the eosinophil chemokine Eotaxin during allergic inflammation. In contrast to Eotaxin, mRNA for mMCP-5 can be produced by mast cells. From these results, we postulate that mMCP-5 plays a pivotal role during the early stages of allergic lung inflammation.

Histamine drives severity of innate inflammation via histamine 4 receptor in murine experimental colitis.

Ulcerative colitis (UC) patients exhibit elevated histamine, but how histamine exacerbates disease is unclear as targeting histamine 1 receptor (H1R) or H2R is clinically ineffective. We hypothesized that histamine functioned instead through the other colon-expressed histamine receptor, H4R. In humans, UC patient biopsies exhibited increased H4R RNA and protein expression over control tissue, and immunohistochemistry showed that H4R was in proximity to immunopathogenic myeloperoxidase-positive neutrophils. To characterize this association further, we employed both the oxazolone (Ox)- and dextran sulfate sodium (DSS)-induced experimental colitis mouse models and also found upregulated H4R expression. Mast cell (MC)-derived histamine and H4R drove experimental colitis, as H4R-/- mice had lower symptom scores, neutrophil-recruitment mediators (colonic interleukin-6 (IL-6), CXCL1, CXCL2), and mucosal neutrophil infiltration than wild-type (WT) mice, as did MC-deficient KitW-sh/W-sh mice reconstituted with histidine decarboxylase-deficient (HDC-/-) bone marrow-derived MCs compared with WT-reconstituted mice; adaptive responses remained intact. Furthermore, Rag2-/- × H4R-/- mice had reduced survival, exacerbated colitis, and increased bacterial translocation than Rag2-/- mice, revealing an innate protective antibacterial role for H4R. Taken together, colonic MC-derived histamine initiates granulocyte infiltration into the colonic mucosa through H4R, suggesting alternative therapeutic targets beyond adaptive immunity for UC.

Substance P-induced augmentation of cutaneous vascular permeability and granulocyte infiltration in mice is mast cell dependent.

The undecapeptide substance P is thought to mediate both vasodilatation and augmented vascular permeability when released from sensory nerve endings in the skin. Substance P also induces mast cell degranulation in vitro or in vivo. However, the extent to which substance P-induced changes in vascular permeability are mast cell-dependent is unclear. We investigated this issue by injecting substance P and certain related peptides (substance P1-4, substance P4-11) into the skin of genetically mast cell-deficient WBB6F1-W/W or WCB6F1- SI/SId mice the congenic normal (+/+) mice, and W/W mice which had undergone selective local repair of their mast cell deficiency by intradermal injection of IL-3-dependent mast cells generated in vitro from the bone marrow cells of the congenic +/+ mice. Substance P induced significant augmentation of vascular permeability and significant cutaneous swelling when injected into normal mice at doses as low as 2 pmol i.d. Substance P also induced granulocyte infiltration, although the infiltrate were modest and were seen at doses of peptide from 5 to more than 20-fold higher than those required for induction of tissue swelling. The effects of substance P on tissue swelling, vascular permeability, and granulocyte infiltration were virtually entirely mast cell dependent. By contrast, substance P1-4 was inactive in our assays at 25 nmol/site, and substance P4-11 induced modest augmentation of vascular permeability, which was at least in part mast cell independent.

Recruitment of neutrophils during IgE-dependent cutaneous late phase reactions in the mouse is mast cell-dependent. Partial inhibition of the reaction with antiserum against tumor necrosis factor-alpha.

Much of the clinically important pathology associated with IgE-dependent disorders is thought to reflect the actions of the blood-borne leukocytes recruited during these responses. To evaluate the extent to which mast cells are responsible for the leukocyte infiltration associated with IgE-dependent cutaneous reactions, we attempted to elicit these responses in normal mice, genetically mast cell-deficient W/Wv mice, and in W/Wv mice selectively repaired of their mast cell deficiency by the intradermal injection of cultured mast cells derived from the congenic normal (+/+) mice. We found that the tissue swelling associated with IgE-dependent passive cutaneous anaphylaxis reactions developed rapidly and diminished markedly from 2 to 4 h after antigen challenge, but remained detectable for at least 24 h after elicitation of the responses. Infiltration of leukocytes (predominantly neutrophils) also occurred at these sites, but reached maximal levels 6-12 h after antigen challenge, persisted at high levels for 24 h, and largely waned by 48 h. Virtually all of the tissue swelling and leukocyte infiltration associated with IgE-dependent cutaneous reactions was mast cell dependent. Intradermal injection of 40 U of recombinant murine TNF-alpha (rmTNF-alpha) elicited neutrophil infiltration similar in magnitude and kinetics to that observed after IgE-dependent mast cell degranulation. A rabbit anti-rmTNF-alpha (R anti-rmTNF-alpha) antiserum, which was able to inhibit 84% of the neutrophil infiltration observed after i.d. injection of rmTNF-alpha, inhibited IgE-, and mast cell-dependent leukocyte infiltration by 47 +/- 7% in three separate experiments. These findings indicate that TNF-alpha contributes to mast cell-dependent recruitment of leukocytes during IgE-dependent cutaneous late phase reactions, but suggest that other mast cell-associated mediators probably also contribute to this response.

Role of mast cells in ion transport abnormalities associated with intestinal anaphylaxis. Correction of the diminished secretory response in genetically mast cell-deficient W/Wv mice by bone marrow transplantation.

To investigate the role of mast cells in transport abnormalities during intestinal anaphylaxis, we examined responses to antigen in isolated intestinal preparations from ovalbumin-sensitized genetically mast cell-deficient WBB6F1-W/Wv (W/Wv) mice and congenic normal WBBGF1(-)+/+ (+/+) mice. Changes in ion transport (primarily secretion of chloride ions) were indicated by increases in short-circuit current (Isc). In tissues from +/+ mice, antigen caused increases in Isc which were significantly inhibited by antagonists to histamine (diphenhydramine) and serotonin (ketanserin), by a cyclooxygenase inhibitor (piroxicam) and by a neurotoxin (tetrodotoxin). In preparations from W/Wv mice, antigen-stimulated responses were approximately 30% of that in +/+ mice and were inhibited only by piroxicam. Responses to electrical transmural stimulation of nerves were approximately 50% in W/Wv versus +/+ mice, and were inhibited by antagonists of mast cell mediators in +/+ but not W/Wv mice. Reconstitution of mast cells in W/Wv mice by intravenous injection of +/+ bone marrow cells restored the normal responses to both antigen and nerve stimulation. Our results indicate that mast cell-dependent mechanisms are primarily responsible for the ion secretion associated with intestinal anaphylaxis, but that other cells are also involved. In addition, our data provide evidence for the functional importance of bidirectional communication between nerves and mast cells in the regulation of ion transport in the gastrointestinal tract.

Cytokine production by mast cells and basophils.

Mast cells and/or basophils have been implicated in the expression of a wide variety of biological responses, including immediate hypersensitivity reactions, host responses to parasites and neoplasms, angiogenesis, tissue remodeling, and immunologically non-specific inflammatory and fibrotic conditions. Recent findings suggest that an important mechanism by which mast cells influence such biological responses is through the production of a broad panel of multifunctional cytokines. In contrast, the extent to which basophils can produce cytokines is uncertain.

Eotaxin influences the development of embryonic hematopoietic progenitors in the mouse.

Eotaxin is a potent chemoattractant for eosinophils during inflammation and allergic reactions in the adult, but its role during development has not been studied. We report that eotaxin and its receptor, CCR-3, are expressed by embryonic tissues responsible for blood development, including the yolk sac, fetal liver, and fetal blood. We also found that eotaxin acts synergistically with stem cell factor (SCF) to accelerate the differentiation of embryonic mast cell progenitors and to promote the growth of Mac-1+/Gr-1- cells from progenitors isolated at 10-12 days of gestation. This response is diminished by Pertussis toxin, the Gi alpha inhibitor. These studies suggest that eotaxin is involved in the growth of myeloid cell progenitors and the differentiation of mast cells during embryogenic development.

Mast cell regulation of inflammation and gene expression during antigen-induced bladder inflammation in mice.

Mast cell numbers are significantly increased in bladder disorders including malignancy and interstitial cystitis, but their precise role has been difficult to determine. We characterized the role of mast cells on gene regulation associated with antigen-induced bladder inflammation in mice. For this purpose, we examined the responses in mast cell-deficient (Kit(W)/Kit(W-v)), congenic normal (+/+), and Kit(W)/Kit(W-v) mice that were reconstituted with bone marrow stem cells (BMR) to restore mast cells. All mice were actively sensitized and challenged intravesically with either saline or specific antigen. Bladder inflammation occurred in +/+ and BMR but not the Kit(W)/Kit(W-v) mice. Gene expression was determined using mouse cDNA expression arrays. Self-organizing maps, performed without preconditions, indicated gene expression changes dependent on the presence of mast cells. These genes were upregulated in bladders isolated from antigen challenge of +/+, not altered in Kit(W)/Kit(W-v), and were upregulated in BMR mice. Taken together these results demonstrate an important role for mast cells in allergic cystitis and indicate that mast cells can alter their environment by regulating tissue gene expression.

Analyzing mast cell development and function using mice carrying mutations at W/c-kit or Sl/MGF (SCF) loci.

Mast cells have been implicated in a wide variety of biological responses, but identifying the nature and importance of the mast cell's specific contributions to these reactions has been difficult. W/Wv mice have mutations affecting the c-kit tyrosine kinase receptor which is encoded at the W locus and which is necessary for normal mast cell development. In W/Wv mice, the cells which ordinarily give rise to normal mast cell populations do not adequately respond to a major migration, survival, proliferation and maturation factor expressed in the microenvironments where mast cells ordinarily develop: the c-kit receptor ligand, SCF. As a result, W/Wv mice virtually lack tissue mast cells. However, adoptive transfer to W/Wv mice of immature mast cells derived in vitro from the bone marrow cells of the congenic normal (+/+) mice selectively repairs the mast cell deficiency of the W/Wv recipients. These "mast cell knock-in" mice can be used to analyze the expression of biological responses in tissues which differ only because they do or do not contain populations of mast cells. This approach permits identification and quantification of the specific contributions of the mast cell to biological responses expressed in the skin, gastrointestinal tract and other anatomical sites, and also greatly facilitates analysis of the mechanisms by which mast cells influence these responses.

Mast cells contribute to PACAP-induced dermal oedema in mice.

In the present study the effect of intradermal PACAP-injection on dermal oedema in mice was investigated and the contribution of mast cells to this response was assessed. The injection of PACAP 1-38 into the ears of C57BL/6 mice evoked a dose-dependent response, which, after higher doses of PACAP 1-38, lasted at least 24 h. Histological examination showed significant mast cell degranulation induced by PACAP. Using mast cell-deficient WBB6F1-Kit(W)/Kit(W-v) mice and the congenic mice, we demonstrated that the the early phase (30 min to 6 h) of PACAP-induced ear swelling response was significantly diminished in mast cell-deficient mice, suggesting that mast cell degranulation contributes to this phase of the response. When mast cell-deficient WBB6F1-Kit(W)/Kit(W-v) mice were locally and selectively reconstituted by adoptive mast cell transfer, the dermal oedema was almost equal to that of control animals in the early phase of PACAP injection. These results show that mast cell degranulation contributes to PACAP-induced dermal oedema in mice.

Stem cell factor influences neuro-immune interactions: the response of mast cells to pituitary adenylate cyclase activating polypeptide is altered by stem cell factor.

Mast cells degranulation can be elicited by a number of biologically important neuropeptides, but the mechanisms involved in mast cell-neuropeptide interactions have not been fully elucidated. Stem cell factor (SCF), also known as c-kit or kit ligand, induces multiple effects on mast cells, including proliferation, differentiation, maturation, and prevents apoptosis. We investigated the ability of SCF to affect mast cell responsiveness to the neuropeptides pituitary adenylate cyclase activating polypeptide (PACAP) and vasoactive intestinal peptide (VIP). PACAP 1-27, PACAP1-38, or VIP failed to induced preformed mediator release from mouse bone-marrow-cultured mast cells (BMCMC) derived in concanavalin A-stimulated spleen conditioned medium (CM). By contrast, BMCMC grown in SCF-containing medium or freshly isolated peritoneal mast cells exhibited significant 3H-hydroxytrypamine (5-HT) release in response to PACAP peptides or VIP. Deoxyglucose and the mitochondrial inhibitor antimycin significantly inhibited PACAP-induced 5-HT release indicating that the central event induced by PACAP peptides was exocytosis. The G(alpha)i inhibitor, pertussis toxin, significantly diminished PACAP-induced 5-HT release from BMCMCs in SCF suggesting the involvement of heterotrimeric G-proteins. Western blot analysis using antibodies directed against the human VIP type I/PACAP type II receptor demonstrated a 70-72 kD immunoreactive protein expressed in greater amounts in BMCMC grown in SCF compared with BMCMC in CM. We conclude that SCF induces a mast cell population that is responsive to PACAPs and VIP involving a heterotrimeric G-protein-dependent mechanism.

IX. Mast cell-deficient mice and intestinal biology.

Mutant mice that express abnormalities in mast cell development represent a powerful tool for the investigation of multiple aspects of mast cell biology. In addition, the identification of the genes affected by these mutations has not only increased our knowledge about the mast cell but has opened new areas of investigation as to the role of these gene products in gastrointestinal pathology, immunology, and physiology.

Gastrointestinal mast cells. New approaches for analyzing their function in vivo.

Mast cells have been implicated in a variety of physiologic or pathologic responses in the gastrointestinal tract; however, the precise contributions of the mast cell to these reactions have been difficult to define. This article discusses general aspects of mast cell biology, outlines new approaches for the analysis of mast cell function in vivo, and reviews recent evidence indicating that mast cells can produce a wide spectrum of multifunctional cytokines.

The mucosal barrier, IgE-mediated gastrointestinal events, and eosinophilic gastroenteritis.

The "mucosal barrier" plays an important role in regulating immune responses in the gastrointestinal tract. Although the precise mechanisms controlling mucosal immune responses have not been defined, when an abnormal response such as IgE production is initiated, a variety of complex and incompletely understood pathophysiologic events occur and can result in gastrointestinal pathology. This article attempts to present the complexity of such abnormal responses. Although more questions have emerged than were answered, in recent years our knowledge of the pathophysiology of IgE-mediated events in the gastrointestinal tract has advanced significantly. Our understanding of the pathophysiology of these events will ultimately allow the development of more directed and effective therapeutic interventions for allergic gastrointestinal diseases. In addition, through further clinical and basic research, we should begin to unravel the more enigmatic conditions affecting the gastrointestinal tract, such as EG.

Mouse mast cell cytokine production: rôle in cutaneous inflammatory and immunological responses.

The rôle of mast cells in cutaneous physiology and pathology has long been a subject of intense speculation and great clinical interest. In this brief review, which is focused primarily on murine systems, we outline certain important aspects of the biology of the mast cell, including their ability to produce a variety of cytokines, describe the development of a model system (the "mast cell knock-in mouse") for studying the roles of cutaneous mast cells in biological responses in vivo, and illustrate a few examples of how this approach has been used to investigate the contributions of mast cells and their cytokines to cutaneous inflammatory or immune responses.

Gastric inflammation during systemic anaphylaxis: neutrophil recruitment in stomach wall of mice does not require mast cell participation.

We determined whether neutrophil infiltration into the stomach wall occurred during systemic anaphylaxis in mice and assessed the participation of mast cells in the response. Normal mice sensitized and challenged with antigen exhibited significant neutrophil infiltration in the gastric mucosa and submucosa compared with saline-challenged mice. The development of clinical signs of anaphylaxis and extent of gastric neutrophil infiltration was similar in mast cell-deficient Kit(W)/Kit(W-v) or Mgf(Sl)/Mgf(Sl-d) mice and the respective normal congenic mice. Pretreatment with sodium cromoglycate prevented the clinical signs of anaphylaxis and significantly diminished the infiltration of neutrophils in +/+ or Kit(W)/Kit(W-v) mice. Systemic anaphylaxis is associated with neutrophil infiltration into the stomach wall in mice, and mast cells are not required for the development of either the clinical manifestations or gastric neutrophil infiltration observed in the response.

Mast cell-dependent amplification of an immunologically nonspecific inflammatory response. Mast cells are required for the full expression of cutaneous acute inflammation induced by phorbol 12-myristate 13-acetate.

Mast cells clearly are critical for the expression of some IgE-dependent responses, but their roles in other forms of inflammation are uncertain. We previously described a new model system for defining the unique contribution of mast cells to biologic responses in vivo, genetically mast cell-deficient WBB6F1-W/Wv mice that have undergone selective local repair of their mast cell deficiency by the injection of IL-3-dependent cultured mast cells derived from the congenic normal (WBB6F1-+/+) mice. Using this approach, we analyzed the contribution of mast cells to the acute inflammation induced by the epicutaneous application of PMA. Even though PMA can activate a wide variety of cell types that may contribute to acute inflammation, we found that mast cells were required for the full expression of the tissue swelling and leukocyte infiltration associated with the response to the agent in vivo. Thus, in WBB6F1-W/Wv mice selectively reconstituted with dermal mast cells by intradermal injection of cultured WBB6F1-+/+ mast cells into the left ear only, PMA induced approximately twice the tissue swelling and neutrophil infiltration in the mast cell-reconstituted left ears as in the contralateral control ears. This represents the first use of W/Wv mice locally reconstituted with mast cells to confirm the hypothesis that mast cells can represent an important amplification mechanism in acute inflammatory responses of nonimmunologic origin. It also defines a model system that may be generally useful for investigating mast cell-dependent and -independent aspects of acute inflammatory responses.