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The esophagus is the only segment in the gastrointestinal (GI) tract that is devoid of eosinophils; whereas, most of the other leukocytes reside in the esophagus at baseline in healthy state (1, 2). The esophagus is lined with mucous membrane and muscles that act with peristaltic action to move swallowed food down to the stomach. The epithelium of the esophagus is squamous but not keratinized like skin; therefore keratinocytes are directly exposed to the esophageal content. This indicates that esophageal epithelium may have a significant role in the induction of esophageal inflammation. The accumulation of eosinophils in the esophageal mucosa is the cardinal pathological finding that occurs secondary to several unrelated diseases and is reported in a number of esophageal diseases such as hypereosinophilic syndrome, eosinophilic gastroenteritis, drug reactions, fungal/parasitic infections, gastroesophageal reflux (GERD), and eosinophilic esophagitis (EoE) (1-11). (EoE or EE- For consistency thru the monograph we would like to call eosinophilic esophagitis EoE. EoE has been increasingly confused with erosive esophagitis in the adult gi world-)) EoE is a commonly observed medical problem and is well documented in pediatric patients, but the adult form has only recently gained recognition as a distinct entity. EoE is characterized by an increase in esophageal eosinophilia, basal cell hyperplasia and the number of other esophageal abnormalities that include furrows, the formation of fine concentric mucosal rings (corrugated esophagus), and esophageal strictures (narrowing) (3-9), associated with extensive tissue remodeling and fibrosis (10, 11).
Esophageal eosinophils are not pathognomonic for EoE since eosinophil infiltration in the esophagus occurs in a variety of states including GERD. Differentiating EoE from other esophageal disorders, specifically GERD is often a challenge (23). Patients with primary EoE commonly report symptoms that include difficulty feeding, vomiting, chest pain, dysphagia, and food impaction (19, 24-26); these symptoms appear to occur sequentially as disease progresses from infancy into adulthood (27). Dysphagia and food impaction are commonly observed in adult EoE patients (28-30). However, longitudinal studies from childhood into adulthood are not yet available. EoE patients are predominantly young males (19, 24) and have relatively high levels of eosinophils in the esophageal mucosa, extensive epithelial hyperplasia, and a high rate of atopic disease compared to patients with GERD (22, 31). In particular, esophageal eosinophil levels of >24/hpf have been reported to correlate with lack of responsiveness to anti-GERD therapy (12, 32); these concentrations may be diagnostic of EoE rather than GERD, especially in patients already on anti-GERD therapy. A recent expert panel established as part of the First International Group of EoE Researchers (FIGERS) recommended that a cutoff of 15 eosinophils/hpf is sufficient for the diagnosis of EoE provided that GERD has been eliminated as the diagnosis (33); for research purposes a higher threshold level was recommended. Additionally, esophageal biopsies from EoE demonstrate a thickened mucosa with basal layer hyperplasia and papillary lengthening. Radiographic and endoscopic studies have shown many findings, including small caliber esophagus, strictures, mucosal rings, ulcerations, whitish papules and polyps (8, 21, 25, 26, 36, 37). EoE has been found to be associated with esophageal dysmotility but the etiology of the motor disturbances is unclear. The eosinophil and mast cell activation and degranulation has been postulated as a possible etiology of EoE pathogenesis (13, 34, 35).
Food allergies affect an estimated 6% of children and 3.7% of adults in the US and during the past decade food allergy and its manifestations have substantially increased (53, 54). Food allergies can be classified into those that are IgE-mediated and those that are non-IgE-mediated. IgE-mediated reactions develop when food-specific IgE antibodies residing on mast cells and basophils come into contact with and bind to circulating food allergens and activate the cells to release potent mediators and cytokines. In non-IgE-mediated food allergic disorders, multiple inflammatory cells and their mediators play a role in immunopathogenesis. The majority of EoE patients (90%) have evidence of food and aeroallergen hypersensitivity yet only a subset (10-30%) has a history of food anaphylaxis (38). Recent literature on pediatric patients with EoE confirms that nearly all patients respond to an elemental diet with resolution of symptoms and normalization of biopsies (38); reintroduction of foods causes symptoms and esophageal eosinophilia to return (39). Interestingly, patients with EoE have also been reported to exhibit seasonal variations in their symptoms and changes in their esophageal eosinophil levels. The mucosal eosinophil counts were elevated during the spring and summer and were suppressed during the winter (40), indicating a role for aeroallergens. Additionally, studies of animal models have also linked EoE to aeroallergens and allergic diseases (41). These findings indicate that sensitization pathways could occur in human EoE and antigen presenting cells (APCs) may play an important role in the pathogenesis of EoE. An average of 3 to 6 foods per patient were directly linked to the development of esophageal eosinophilia and the common foods identified were milk, egg, soy, chicken, wheat, beef, corn, and peanuts (38, 39). Taken together, this provides supportive evidence that food and aeroallergen sensitization is causally involved in EE. (Figure 1)
The current understanding of the pathophysiology of EE comes from both basic immunological studies and clinical observation and treatment. A large number of eosinophils were detected in the esophagus following induction of EE in humans as well as in an experimental mouse model of EE. These eosinophils contain a number of toxic granular proteins. Eosinophil granules contain a crystalloid core composed of major basic protein (MBP-1 and -2), and a matrix composed of eosinophil cationic protein (ECP), eosinophil-derived neurotoxin (EDN), and eosinophil peroxides (EPO) (42). These cationic proteins share certain pro-inflammatory properties but differ in other ways. The MBP, EPO, and ECP have cytotoxic effects on epithelium and ECP and EDN possess anti-viral and ribonuclease activity (44). MBP directly increases smooth muscle reactivity (45) and also triggers degranulation of mast cells and basophils (46). Indeed, evidence is emerging that mast cells and eosinophils both are involved in EE pathogenesis (47, 48). Several studies have shown that mast cells are present in the normal esophagus and their number increases in esophageal biopsies of EE patients (47, 48). Interestingly, mast cell gene (like tryptase or carboxypeptidase's 3) induced expression is noticed in the esophageal biopsies of EE patients (47). Mast cells are known to release mediators (such as histamine and proteases) and upon activation they cause pathophysiological alterations in tissues (49, 50). Evidence indicates that both eosinophils and mast cells are present in an activated state in EE patients (35, 47, 52). Activated eosinophils and mast cells generate lipid mediators that stimulate smooth muscle contraction, increased vascular permeability and mucus secretion (51). Moreover, the leukotrienes released by these cells can also recruit inflammatory cells from the circulation, which would be an indirect pathway affecting the inflammatory response. Eosinophils and mast cells are also the source of transforming growth factor (TGF)-β that may induce tissue fibrosis. Of note, the amount of TGF-β is increased in the esophagus of human and mouse models of experimental EoE; therefore, induced TGF-β is capable of modifying clinical symptoms in the esophagus of EoE patients.
Eosinophil accumulation often occurs in the absence of infiltration by other inflammatory cells. As a result, there has been a considerable amount of research focused on identifying eosinophil specific chemoattractants (53). Eosinophils respond to a variety of chemoattractants including lipids, complement degradation products, and various chemokines including eotaxin, RANTES, and MCPs. Eotaxin was originally identified as the predominant eosinophil chemoattractant and a potent activator of eosinophils (54). The specificity of eotaxin for eosinophils is the result of the exclusive signaling of eotaxin through its receptor, CCR3, which is expressed predominantly on human and mouse eosinophils (53, 55-58). Eosinophils express several chemokine receptors, but CCR3 is expressed at the highest level per cell (55, 56, 59). CCR3 appears to function as the predominant chemokine receptor since CCR3 ligands are generally the most potent eosinophil chemoattractants. Consistent with the expression of CCR3, eosinophils respond to MIP-1, RANTES, MCP-2, MCP-3, MCP-4, eotaxin-1, eotaxin-2, and eotaxin-3. Both eotaxin-2 and -3 have 30% homology to eotaxin-1 (48, 60, 61) Recently, we reported that eotaxin-3 is a signature gene for EoE, which is also a powerful eosinophil activating protein (48, 61, 62). Eotaxin-3 is induced in EoE patients of all ages regardless of gender and the allergic status of the patient and is completely distinct from the gene expression in patients with reflux esophagitis. Previously, it has been shown that eotaxin-3 is a chemoattractant for eosinophils and esophageal epithelium is a rich source of eotaxin-3 in EoE patients (48). This indicates that eotaxin-3 has a critical role in the accumulation of eosinophils in the esophageal mucosa of EoE patients. We recently showed that an eotaxin-3 gene single nucleotide polymorphism is associated with EoE (48). Furthermore, the significance of eotaxin-3 is supported by our previous work demonstrating that mice with genes targeted for the eotaxin receptor, CCR3 were protected from the development of experimental EoE (48).
IL-5 is the most specific cytokine for eosinophil growth, differentiation, activation, and survival and primes eosinophils to respond to chemoattractants such as eotaxin, an eosinophil selective CC chemokine (63-65). IL-5 also facilitates eosinophil migration from bone marrow to the blood (69). Recent experiments have demonstrated that IL-5 is over-expressed in the esophagus of patients with EoE (66). Additionally, systemic over-expression of IL-5 (via pharmacological or transgenic approaches) promotes eosinophil trafficking to the esophagus in mice (67) and neutralizing anti-IL-5 (TRFK5) treatment in a murine model of EoE restricts eosinophil trafficking to the lung and esophagus (41). Determining the role of IL-5 in EoE is of importance because two different studies have shown an improvement in the clinical and pathological symptoms of EoE after anti-IL-5 treatment (68). Animal studies have documented that dietary and environmental antigens such as enteric-coated ovalbumin or Aspergillus antigen induced esophageal eosinophilia (41, 70) was ablated in mice deficient in IL-5 (41, 70). Conversely, transgenic mice overexpressing IL-5 had constitutively elevated number of esophageal eosinophils (71). Eosinophils also produce IL-5; therefore, it is important to understand whether local induction of IL-5 has a role in EoE pathogenesis. We recently showed that local expression of IL-5 is required for the induction of esophageal remodeling in human and experimental EoE.
IL-13 has also been implicated in a variety of allergic conditions including asthma (72), atopic dermatitis (35, 73, 74), and allergic rhinitis (75-77). The over-expression of IL-13 by pharmacological administration or transgenic approaches induces multiple features of asthma, including eosinophilia, mucus over-production, and airway hyperresponsiveness (78, 79). Based on the importance of IL-13 in allergic diseases and the high concordance between asthma and EoE in humans, we delivered IL-13 to the lung by intratracheal administration (Figure 2). Our experimentation established that pulmonary inflammation, triggered by IL-13, is associated with the development of EoE. In humans IL-13 induced lung and esophageal eosinophilia was reduced by pretreatment with human neutralizing IL-13 (CAT-354) antibody (80). Use of experimental models in mice has established that Th2 signaling is required for induction of experimental EoE (41) and this will be further proven by our findings that STAT6 gene-deficient mice are protected from allergen- and IL-13-induced experimental EoE (81, 82). The emerging clinical and experimental data support a central role for IL-5 and IL-13 in EoE pathogenesis (71, 81). Furthermore, a clinical report indicated that stimulation with food and environmental aeroallergens induced Th2 cytokine (IL-5 and IL-13) production by peripheral blood mononuclear cells in adult EoE patients (83). Collectively, these findings demonstrate that not only external allergic triggers but also intrinsic Th2 cytokines and eosinophil effector function are critical in EoE pathogenesis. The intimate connection between the lung and the esophagus was previously shown in experimental models of EoE; however, recent findings indicate that allergen-induced EoE is independent of the lung inflammation (84). Following intranasal allergen challenge, esophageal eosinophilic inflammation is induced in CD4 gene-deficient mice without inducing eosinophilic lung inflammation (84), although lung and esophageal inflammation is a T cell dependent process (84). This indicates a different mechanism is operational in the induction of esophageal and lung eosinophilic inflammation that requires further research to provide a better understanding of the mechanistic pathway of the induction of EoE and its relation to other allergic diseases.
Recently, we and others showed that eosinophil recruitment to the esophagus induces esophageal remodeling, specifically in the lamina propria in the pediatric EoE population (10, 11). An impressive collagen accumulation in the epithelial and subepithelial mucosa, increased vascularity, basal cell hyperplasia and vascular activation along with increased expression of TGF-β, VCAM-1, SMAD2/3, MUC5AC and IL-5 genes was observed in esophageal biopsies of EoE patients (10). The induced gene expression of TGF-β and its signaling molecule SMAD2/3 in the esophagus of human and experimental EoE indicates that the TGF-β pathway may play a critical role in the development of esophageal pathophysiological abnormalities including the formation of esophageal rings and strictures in EoE patients. By using mouse models of EoE, we demonstrated that local IL-5 expression-induced esophageal eosinophilia is critical in the development of esophageal remodeling (11). Fibroblast proliferation is observed in the esophageal biopsies of EoE patients, and previously it has been reported that eosinophil-fibroblast interactions generate subepithelial fibrosis in tissue in inflammatory states. Our recent observation that local expression of IL-5 is critical for esophageal remodeling is in accordance with a previous report showing that IL-5 induces fibroblast to myofibroblast transdifferentiation and the expression of α-smooth muscle actin and ?? extracellular matrix (85). Although a few recent reports indicate that esophageal remodeling and fibrosis occurs in EoE, the mechanism of the induction of esophageal fibrogenesis remains unclear. In order to define the changes in esophageal epithelial cells, fibroblast proliferation, differentiation, extracellular matrix accumulation and smooth muscle hyperplasia in EoE more experimental studies are needed.
EoE is a newly recognized disease and is an emerging entity throughout developing and developed countries including the US (5, 6, 12-16, 19, 22, 86, 87). Therefore, understanding of the causes, natural history, diagnosis and management is important for future theraputic interventions for EoE. The diagnosis of EoE is generally made by performing an esophageal biopsy, with evidence of eosinophils infiltrating the esophageal mucosa. Once EoE is diagnosed, an allergist will typically perform extensive allergy testing looking for evidence of food and environmental allergen sensitization. Currently, therapy for esophageal inflammation is based on antigen elimination trials, anti-inflammatory approaches, and physical dilatation when strictures are present. The latter approach, which has been the strategy reported for adults, temporarily reduces symptoms of dysphagia; however, it involves the significant risk of rupture and hemorrhage, needs to be repeated regularly, and does not reduce the underlying inflammation (88). An antigen elimination approach in sensitized individuals (e.g. aeroallergen avoidance and food elimination diets) is typically unsatisfactory or practically difficult (when patients are sensitized to many allergens), probably because the current allergen sensitization tests (skin prick and patch tests, as well as allergen-specific plasma IgE levels) are not optimal for detecting sensitization. A diet consisting exclusively of an elemental (amino acid based) formula frequently improves symptoms and normalizes esophageal pathology (89-91). However, this approach is often not tolerated (especially in older individuals), frequently requires a feeding tube, and can be expensive. Systemic steroids are used for acute exacerbations, while topical glucocorticoids are used to provide long-term control (92, 93). Glucocorticoid treatment shows significant effect in reducing esophageal eosinophila. Newer glucocorticoids with decreased systemic effect may improve the care of EoE patients. Although some treatments are effective in EoE, the molecular mechanisms involved in the remission have still not been established. Humanized antibody therapy designed to block IL-5 and IL-13 is currently in testing and has demonstrated great promise. Recent reports indicate that anti-IL-5 therapy has a significant effect on EoE in patients and CAT-354, an IL-13 blocking antibody has been shown to prevent lung and esophageal eosinophilia in mice (68, 80). The anti-IL-5 therapeutic trials have been conducted in a small number of patients and more extensive studies are needed. The development of an experimental model of EoE is the most useful tool to dissect the molecular mechanisms involved in remission or resistance to therapy. The current knowledge suggests that targeting IL-5, IL-13 and eotaxin-3 may be promising therapeutic strategies for the treatment of EoE.
EoE is linked to allergic responses to food or aeroallergens, but cases have also been reported in which patients have EE without detectable food allergies by patch or prick skin testing. This indicates that EE could also be associated with immune dysregulation and these tests might not reflect hypersensitivity driven by discrete antigens. The pathogenesis of EE is still not clear, but a growing body of evidence has established that this condition represents a T cell mediated immune response involving a number of pro-inflammatory mediators and chemoattractants known to regulate eosinophilic accumulation in the esophagus, such as IL-4, IL-5, IL-13 and eotaxin-1, -2 and -3. Our recent findings indicate that the most highly expressed gene in EoE patients is eotaxin-3, whereas, eotaxin-1 and -2 are modestly increased. In addition, esophageal eosinophilia correlates with eotaxin-3 mRNA and protein levels in esophageal biopsies of EoE patients. It has been recently reported that the esophageal epithelium from esophageal biopsies of EE patients expresses eotaxin-3 and exposure of esophageal epithelial cells to IL-13 induces eotaxin-3. These data indicate that IL-5, IL-13 and eotaxin-3 are the major mediators that regulate eosinophil numbers in the esophagus. Determining the mechanism(s) through which human esophageal cell-derived factors ultimately induce the functional abnormalties observed, and to which antigens EoE patients are sensitized that lead to the manifestation of symptoms is of significant interest. Esophageal dysmotility is a recognized complication of GERD, but has not yet been studied in EoE. Therefore, experimental models need to be designed to identify and evaluate the key factors in the inflammatory pathways of EoE that are directly responsible for altered smooth muscle contractility and eventually clinical symptoms. These studies will lead to a better understanding of the mechanisms of the development of EoE and likely provide further treatment options.
This work is supported by NIH RO1 DK067255.
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