Simon P. Hogan

IL-5 Promotes Eosinophil Trafficking to the Esophagus

Eosinophil infiltration into the esophagus occurs in a wide range of diseases; however, the underlying pathophysiological mechanisms involved are largely unknown. We now report that the Th2 cytokine, IL-5, is necessary and sufficient for the induction of eosinophil trafficking to the esophagus. We show that transgenic mice overexpressing IL-5 under the control of a T cell (CD2) or a small intestinal enterocyte (fatty acid-binding protein) promoter have markedly increased eosinophil numbers in the esophagus. For example, esophageal eosinophil levels are 1.9 ± 0.9 and 121 ± 14 eosinophils/mm2 in wild-type and CD2-IL-5-transgenic mice, respectively. Consistent with this effect being mediated by a systemic mechanism, pharmacological administration of IL-5 via a miniosmotic pump in the peritoneal cavity resulted in blood and esophageal eosinophilia. To examine the role of IL-5 in oral Ag-induced esophageal eosinophilia, eosinophilic esophagitis was induced by allergen exposure in IL-5-deficient and wild-type mice. Importantly, IL-5-deficient mice were resistant to eosinophilic esophagitis. Finally, we examined the role of eotaxin when IL-5 was overproduced in vivo. Esophageal eosinophil levels in CD2-IL-5-transgenic mice were found to decrease 15-fold in the absence of the eotaxin gene; however, esophageal eosinophil numbers in eotaxin-deficient IL-5-transgenic mice still remained higher than wild-type mice. In conclusion, these studies demonstrate a central role for IL-5 in inducing eosinophil trafficking to the esophagus.

Aeroallergen-induced eosinophilic inflammation, lung damage, and airways hyperreactivity in mice can occur independently of IL-4 and allergen-specific immunoglobulins

In this investigation we have used a mouse model containing certain phenotypic characteristics consistent with asthma and IL-4- and CD40-deficient mice to establish the role of this cytokine and allergen-specific immunoglobulins in the initiation of airways hyperreactivity and morphological changes to the airways in responses to aeroallergen challenge. Sensitization and aerosol challenge of mice with ovalbumin resulted in a severe airways inflammatory response which directly correlated with the induction of extensive airways damage and airways hyperreactivity to beta-methacholine. Inflammatory infiltrates were primarily characterized by the presence of CD4+ T cells and eosinophils. In IL-4-deficient mice, the recruitment of airways eosinophils was impaired, but not abolished in response to aeroallergen. Moreover, the characteristic airways damage and hyperreactivity normally resulting from allergen inhalation were not attenuated. Induction of these structural and functional changes to the airways occurred in the absence of ovalbumin-specific IgE and IgG1, but IgG2a and IgG3 were detected in the sera of IL-4-deficient mice. CD4+ T cells isolated from both wild-type and IL-4-deficient mice given ovalbumin produced significant levels of IL-5 after in vitro stimulation. Treatment of IL-4-deficient mice with anti-IL-5 mAb before aeroallergen challenge abolished blood and airways eosinophilia, lung damage, and airways hyperreactivity. These results indicate that IL-4 is not essential for the development of IL-5-producing CD4+ T cells or for the induction of eosinophilic inflammation and airways damage and hyperreactivity. In response to sensitization and aerosol challenge, CD40-deficient mice did not produce ovalbumin-specific IgE, IgG isotypes, or IgA, and airways inflammation and hyperreactivity were not attenuated. Our results suggest that allergic airways disease can occur via pathways which operate independently of IL-4 and allergen-specific immunoglobulins. Activation of these pathways is intimately associated with IL-5 and eosinophilic inflammation. Such pathways may play a substantive role in the etiology of asthma.

A pathological function for eotaxin and eosinophils in eosinophilic gastrointestinal inflammation

Although eosinophils have been implicated in the pathogenesis of gastrointestinal disorders, their function has not been established. Using a murine model of oral antigen–induced eosinophil-associated gastrointestinal disease, we report the pathological consequences of eosinophilic inflammation and the involvement of eotaxin and eosinophils. Exposure of mice to enteric-coated antigen promotes an extensive T helper 2–associated eosinophilic inflammatory response involving the esophagus, stomach, small intestine and Peyers patches as well as the development of gastric dysmotility, gastromegaly and cachexia. Electron microscopy shows eosinophils in proximity to damaged axons, which indicated that eosinophils were mediating a pathologic response. In addition, mice deficient in eotaxin have impaired eosinophil recruitment and are protected from gastromegaly and cachexia. These results establish a critical pathological function for eotaxin and eosinophils in gastrointestinal allergic hypersensitivity.

Elemental signals regulating eosinophil accumulation in the lung

Summary: In this review we identify the elemental signals that regulate eosinophil accumulation in the allergic lung. We show that there are two interwoven mechanisms for the accumulation of eosinophils in pulmonary tissues and that these mechanisms are linked to the development of airways hyperreactivity (AHR). Interleukin‐(IL)‐5 plays a critical role in the expansion of eosinophil pools in both the bone marrow and blood in response to allergen provocation of the airways. Secondly, IL‐4 and IL‐13 operate within the allergic lung to control the transmigration of eosinophils across the vascular bed into pulmonary tissues. This process exclusively promotes tissue accumulation of eosinophils. IL‐13 and IL‐4 probably act by activating eosinophil‐specific adhesion pathways and by regulating the production of IL‐5 and eotaxin in the lung compartment. IL‐5 and eotaxin co‐operate locally in pulmonary tissues to selectively and synergistically promote eosinophilia. Thus, IL‐5 acts systemically to induce eosinophilia and within tissues to promote local chemotactic signals. Regulation of IL‐5 and eotaxin levels within the lung by IL‐4 and IL‐13 allows Th2 cells to elegantly co‐ordinate tissue and peripheral eosinophilia. Whilst the inhibition of either the IL‐4/IL‐13 or IL‐5/ eotaxin pathways resulted in the abolition of tissue eosinophils and AHR, only depletion of IL‐5 and eotaxin concurrently results in marked attenuation of pulmonary inflammation. These data highlight the importance of targeting both IL‐5 and CCR3 signalling systems for the resolution of inflammation and AHR associated with asthma.

Murine Eotaxin-2: A Constitutive Eosinophil Chemokine Induced by Allergen Challenge and IL-4 Overexpression

The generation of tissue eosinophilia is governed in part by chemokines; initial investigation has identified three chemokines in the human genome with eosinophil selectivity, referred to as eotaxin-1, -2, and -3. Elucidation of the role of these chemokines is dependent in part upon analysis of murine homologues; however, only one murine homologue, eotaxin-1, has been identified. We now report the characterization of the murine eotaxin-2 cDNA, gene and protein. The eotaxin-2 cDNA contains an open reading frame that encodes for a 119-amino acid protein. The mature protein, which is predicted to contain 93 amino acids, is most homologous to human eotaxin-2 (59.1% identity), but is only 38.9% identical with murine eotaxin-1. Northern blot analysis reveals three predominant mRNA species and highest constitutive expression in the jejunum and spleen. Additionally, allergen challenge in the lung with Asperigillus fumigatus or OVA revealed marked induction of eotaxin-2 mRNA. Furthermore, eotaxin-2 mRNA was strongly induced by both transgenic over-expression of IL-4 in the lung and administration of intranasal IL-4. Analysis of eotaxin-2 mRNA expression in mice transgenic for IL-4 but genetically deficient in STAT-6 revealed that the IL-4-induced expression was STAT-6 dependent. Recombinant eotaxin-2 protein induced dose-dependent chemotactic responses on murine eosinophils at concentrations between 1–1000 ng/ml, whereas no activity was displayed on murine macrophages or neutrophils. Functional analysis of recombinant protein variants revealed a critical role for the amino terminus. Thus, murine eotaxin-2 is a constitutively expressed eosinophil chemokine likely to be involved in homeostatic, allergen-induced, and IL-4-associated immune responses.

Immunopathogenesis of Experimental Ulcerative Colitis Is Mediated by Eosinophil Peroxidase

The precise role that individual inflammatory cells and mediators play in the development of gastrointestinal (GI) dysfunction and extraintestinal clinical manifestations of ulcerative colitis (UC) is unknown. In this study, we have used a mouse model of UC to establish a central role for eotaxin and, in turn, eosinophils in the development of the immunopathogenesis of this disease. In this model the administration of dextran sodium sulfate (DSS) induces a prominent colonic eosinophilic inflammation and GI dysfunction (diarrhea with blood and shortening of the colon) that resembles UC in patients. GI dysfunction was associated with evidence of eosinophilic cytolytic degranulation and the release of eosinophil peroxidase (EPO) into the colon lumen. By using IL-5 or eotaxin-deficient mice, we show an important role for eotaxin in eosinophil recruitment into the colon during experimental UC. Furthermore, using EPO-deficient mice and an EPO inhibitor resorcinol we demonstrate that eosinophil-derived peroxidase is critical in the development of GI dysfunction in experimental UC. These findings provide direct evidence of a central role for eosinophils and EPO in GI dysfunction and potentially the immunopathogenesis of UC.

Mucosal IL-12 gene delivery inhibits allergic airways disease and restores local antiviral immunity

Allergic asthma strongly correlates with airways inflammation driven by interleukin (IL)‐4 and IL‐5 secreted by allergen‐specific CD4+ T cells. It is possible that over‐production of these factors in the lungs may render asthmatic individuals less able to resolve virus infection of the respiratory tract by down‐regulating type 1 cytokine‐driven immune responses. IL‐12 is important for the establishment of cell‐mediated immunity (CMI) and may also inhibit responses driven by type 2 cytokine production. Sustained expression of IL‐12 in the airways may, therefore, represent an effective preventive treatment or therapy for allergic asthma and any adverse consequences of excessive production of type 2 cytokines for the development of local CMI. Here, we show that allergic responses in airways profoundly inhibit the development of antiviral CMI in mice following local immunization with vaccinia virus (VV) leading to persistent lung infection. However, mucosal gene transfer of IL‐12 in the lung, via a VV vector, inhibited local type 2 cytokine production, both prevented the development of allergic disease and airways hyperreactivity in a manner largely dependent on endogenous interferon‐γ expression and suppressed established allergic disease, and reversed the suppression of local antiviral CMI responses resulting in rapid resolution of virus infection. Our study provides the first direct demonstration that allergic conditions, particularly in airways, may inhibit immune responses to concomitant virus infection and suggests that transient mucosal IL‐12 gene therapy represents an effective approach to both the prevention and treatment of allergic airways disease and associated immunosuppression of CMI.

Chemokines and Chemokine Receptors: Their Role in Allergic Airway Disease

One of the hallmarks of allergic pulmonary disorders is the accumulation of an abnormally large number of leukocytes including eosinophils, neutrophils, lymphocytes, basophils, and macrophages in the lung (1). There is now substantial evidence that eosinophils, under the control of T lymphocytes, are major effector cells in the pathogenesis of asthma. Therefore, understanding the mechanisms by which eosinophils accumulate and are activated in tissues is a fundamental question very relevant to allergic diseases. Another characteristic of allergic inflammation is the activation of leukocytes resulting in the release of biologically active mediators, such as histamine from mast cells and basophils. It is now apparent that chemokines are potent leukocyte chemoattractants, cellular activating factors, histamine releasing factors, and regulators of homeostatic immunity, making them particularly important in the pathogenesis of airway inflammation in asthma (2). In this regard, chemokines are attractive new therapeutic targets for the treatment of allergic disease. This article focuses on recently emerging data on the importance of chemokines and their receptors in allergic airway inflammation.

Interleukin-5 and eosinophils as therapeutic targets for asthma

Extensive clinical investigations have implicated eosinophils in the pathogenesis of asthma. In a recent clinical trial, humanized monoclonal antibody to interleukin (IL)-5 significantly limited eosinophil migration to the lung. However, treatment did not affect the development of the late-phase response or airways hyperresponsiveness in experimental asthma. Although IL-5 is a key regulator of eosinophilia and attenuation of its actions without signs of clinical improvement raises questions about the contribution of these cells to disease, further studies are warranted to define the effects of anti-IL-5 in the processes that lead to chronic asthma. Furthermore, eosinophil accumulation into allergic tissues should not be viewed as a process that is exclusively regulated by IL-5 but one in which IL-5 greatly contributes. Indeed, data on anti-IL-5 treatments (human and animal models) are confounded by the failure of this approach to completely resolve tissue eosinophilia and the belief that IL-5 alone is the critical molecular switch for eosinophil development and migration. The contribution of these IL-5-independent pathways should be considered when assessing the role of eosinophils in disease processes.

A Plant-Based Allergy Vaccine Suppresses Experimental Asthma Via an IFN-γ and CD4+CD45RBlow T Cell-Dependent Mechanism

Allergic asthma is currently considered a chronic airway inflammatory disorder associated with the presence of activated CD4+ Th2-type lymphocytes, eosinophils, and mast cells. Interestingly, therapeutic strategies based on immune deviation and suppression have been shown to successfully attenuate the development of the asthma phenotype. In this investigation, we have for the first time used a genetically modified (GM) plant, narrow leaf lupin (Lupinus angustifolius L.), expressing a gene for a potential allergen (sunflower seed albumin) (SSA-lupin) to examine whether a GM plant/food-based vaccine strategy can be used to suppress the development of experimental asthma. We show that oral consumption of SSA-lupin promoted the induction of an Ag-specific IgG2a Ab response. Furthermore, we demonstrate that the plant-based vaccine attenuated the induction of delayed-type hypersensitivity responses and pathological features of experimental asthma (mucus hypersecretion, eosinophilic inflammation, and enhanced bronchial reactivity (airways hyperreactivity). The suppression of experimental asthma by SSA-lupin was associated with the production of CD4+ T cell-derived IFN-γ and IL-10. Furthermore, we show that the specific inhibition of experimental asthma was mediated via CD4+CD45RBlow regulatory T cells and IFN-γ. Thus, our data demonstrate that a GM plant-based vaccine can promote a protective immune response and attenuate experimental asthma, suggesting that plant-based vaccines may be potentially therapeutic for the protection against allergic diseases.