New insights into the role of cytokines in asthma.(cytokines as targets in asthma therapeutics)

Abstract

Asthma is a triad of intermittent airway obstruction, bronchial smooth muscle cell hyperreactivity to bronchoconstrictors, and chronic bronchial inflammation. From an aetiological standpoint, asthma is a heterogenous disease, but often appears as a form of immediate hypersensitivity. Many patients with asthma have other manifestations of atopy, such as rhinitis or eczema. Even among nonatopic patients with asthma, the pathophysiology of airway constriction is similar, raising the hypothesis that alternative mechanisms of mast cell degranulation may underlie the disease. The primary inflammatory lesion of asthma consists of accumulation of CD4+ T helper type 2 (TH2) lymphocytes and eosinophils in the airway mucosa. TH2 cells orchestrate the asthmatic inflammation through the secretion of a series of cytokines, particularly interleukin 4 (IL-4), IL-13, IL-5, and IL-9. IL-4 is the major factor regulating IgE production by B cells, and is required for optimal TH2 differentiation. However, blocking IL-4 is not suf ficient to inhibit the development of asthma in experimental models. In contrast, inhibition of IL-13, another TH2 cytokine whose signal transduction pathway overlaps with that of IL-4, completely blocks airway hyperreactivity in mouse asthma models. IL-5 is a key factor for eosinophilia and could therefore be responsible for some of the tissue damage seen in chronic asthma. IL-9 has pleiotropic activities on allergic mediators such as mast cells, eosinophils, B cells and epithelial cells, and might be a good target for therapeutic interventions. Finally, chemokines, which can be produced by many cell types from inflamed lungs, play a major role in recruiting the mediators of asthmatic inflammation. Genetic studies have demonstrated that multiple genes are involved in asthma. Several genome wide screens point to chromosome 5q31-33 as a major susceptibility locus for asthma and high IgE values. This region includes a cluster of cytokine genes, and genes encoding IL-3, IL-4, IL-5, IL-9, IL-13, granulocyte macr ophage colony stimulating factor, and the [beta] chain of IL-12. Interestingly, for some of these cytokines, a linkage was also established between asthma and their receptor. Another susceptibility locus has been mapped on chromosome 12 in a region that contains other potential candidate cytokine genes, including the gene encoding interferon [gamma], the prototypical TH1 cytokine with inhibitory activities for TH2 lymphocytes. Taken together, both experimental and genetic studies point to TH2 cytokines, such as IL-4, IL-13, IL-5, and IL-9, as important targets for therapeutic applications in patients with asthma.

(J Clin Pathol 2001;54:577--589)

Keywords: asthma; cytokines; interleukins; treatment of asthma; interferon [gamma]

Asthma is one of the most common disorders encountered in clinical medicine in both children and adults. It affects approximately 5% of the adult population in the Western world and its reported incidence is increasing dramatically in many developed nations. The cost of the disease is substantial, and the market for the pharmaceutical industry is estimated at $5.5 billion/year. [1 2] Rather than being a single disease, asthma is currently considered to be a group of different disorders characterised by three major features: (1) intermittent and reversible airway obstruction leading to recurrent episodes of wheezing, breathlessness, chest tightness, and cough; (2) bronchohyperresponsiveness (BHR), which is defined as an increased sensitivity to bronchoconstrictors such as histamine or cholinergic agonists; and (3) airway inflammation.

This syndrome arises as a result of interactions between multiple genetic and environmental factors. [1] Most patients also exhibit acute immediate hypersensitivity responses to common inhaled proteins, known as allergens, of which very small amounts trigger IgE dependent mast cell degranulation, leading to reversible airway obstruction. Typical allergen sources include grass pollens and animal danders, but the most important to those with asthma is house dust mite. [2 3] However, a large proportion of patients with asthma present with no personal or family history of allergy, with negative skin tests, and with normal serum concentrations of IgE, and therefore have disease that cannot be classified on the basis of defined immunological mechanisms. In these non-atopic patients, the pathophysiology of airway constriction has some similarities, including eosinophil and T helper type 2 (TH2) lymphocyte infiltration, the presence of Fc[epsilon]RI+ cells, and cells expressing IgE mRNA. [4] In a series of biopsies f rom atopic or non-atopic patients with asthma, the main difference was a stronger macrophage infiltration in non-atopic asthma, although there were more similarities than differences between these subgroups of patients. Both forms of the disease could be IgE mediated, although in non-atopic patients the putative antigen is unknown, and IgE production appears to be local rather than generalised as in atopy. [4] Alternative mechanisms of mast cell degranulation (for example, by locally produced neurotransmitters) may also underlie this disease. However, the origin of non-atopic asthma remains highly controversial, and mast cell independent mechanisms could play a major role in a subgroup of patients.

Work over the past 10 years led to the recognition that chronic inflammation underlies the clinical syndrome of asthma. [5 6] At necropsy, asthmatic lungs typically show hyperinflation, mucus plugging in the airways, clusters of sloughed epithelial cells, and crystalline precipitates of eosinophil derived proteins. Bronchial mucosae are oedematous, the number of goblet cells is increased, the basement membrane is thickened, and the smooth muscle is hypertrophied. T cells, mast cells, eosinophils, and macrophages infiltrate the subepithelium, and the bronchi contain an inflammatory exudate in the bronchus itself. [7]

Although peribronchial inflammation and exaggerated bronchospastic responses are the pathological and physiological cornerstones of the asthmatic syndrome, the mechanisms underlying the initiation and maintenance of these processes remain poorly understood. It is most likely that different immunological processes mediate different aspects of asthma, and that various types of inflammatory responses contribute differentially to the multiple features manifested by patients with asthma. As a result, it has been difficult to design specific inflammation based therapeutic interventions for this disease.

Nevertheless, there is overwhelming evidence that CD4+ T helper cells are responsible for the orchestration of this complex immune reaction. In patients with asthma, CD4+ T cells producing interleukin 4 (IL-4), IL-5, IL-9, and IL-13 (so called TH2 cytokines) have been identified in bronchoalveolar lavages (BAL) and airways biopsies. Because TH2 cytokines are required for the development of airway eosinophilia and IgE in mouse models, it has been proposed that TH2 cells stimulate an inflammatory response that results in asthma. [8] In this review, we will focus on the evidence supporting the role of cytokines in general and the TH2 pathway in particular for asthma. Basically, two different approaches have contributed to highlight the involvement of cutokines. Experimental animal models have enabled the expression and function of individual cytokines to be assessed in vivo and have also allowed the efficacy of cytokine antagonism in asthma-like situations to be studied. In addition, human genetic studies have m ainly helped shed some light on a series of candidate target genes encoding cytokines.

Pathophysiology of asthma

IgE AND MAST CELLS IN THE ACUTE PHASE OF ASTHMA

Similar to allergic rhinitis and atopic dermatitis, asthma is often accompanied by increased concentrations of circulating IgE. Genetic analyses of families have shown that BHR and IgE values are linked. [9] Mast cells are thought to be the main link between IgE and BHR. Crosslinking of IgE bound to mast cells by Fc[epsilon]RI triggers the release of preformed vasoactive mediators such as histamine, the synthesis of prostaglandins and leukotrienes, and the transcription of cytokines. In the bronchial mucosa, these mediators of immediate hypersensitivity reactions rapidly induce mucosal oedema, mucus production, and smooth muscle constriction, and eventually elicit an inflammatory infiltrate.

Quite surprisingly, it has been difficult to demonstrate a precise role for IgE in the pathogenesis of asthma using murine models of the disease. Inflammation of the bronchial mucosa and induction of BHR are elicited to the same extend in wild-type and IgE-/- mice subjected to repeated inhalations of allergen extracts of Aspergillus fumigatus. [10] Even the syndrome of active anaphylaxis, with mast cell activation and mediator release, can be displayed by both ovalbumin (OVA) sensitised IgE-/- and Fc[epsilon]RI-deficient mice after intravenous challenge with OVA. [3 11] Although these findings point to the existence of parallel pathways of allergic reactions, they do not exclude an important role for IgE in allergic diseases in humans. The strong expression of hypersensitivity reactions in the absence of IgE might be species specific; in mice, the IgG1 isotype effectively sensitises mast cells and can passively confer hypersensitivity. Furthermore, in asthma, most animal analyses focused primarily on aspects of the disease that may be essentially cell driven, including eosinophil recruitment and BHR. It is possible that IgE plays a greater role in acute responses to inhaled allergen induced bronchospasm and late phase responses of the airways. [2 3]

Mast cells express approximately 300 000 high affinity IgE receptors/cell, but aggregation of only 100 receptors is required for detectable responses. [12] Histamine, the best studied of mast cell products, accounts for 5-10% of mast cell granule content, and is stored in association with proteoglycans. Histamine receptor stimulation results in smooth muscle contraction, increased vascular permeability, and prostaglandin generation. Chemotactic factors and neutral proteases (mainly tryptase) are the main other preformed mediators found in mast cell granules. Arachidonic acid metabolites, including prostaglandins ([PGD.sub.2]) and leukotrienes ([LTC.sub.4]), are another important group of mast cell derived mediators that are not stored but produced de novo after mast cell degranulation. [PGD.sub.2] and [LTC.sub.4] are potent bronchoconstrictors for human airways in vitro. [12]

Mast cell derived mediators have been found in lavage fluid from patients with asthma, supporting the role of these cells in the immediate or early allergic reaction in asthma. When allergen challenge preceded BAL, increases were documented for histamine, [LTD.sub.4] [PGD.sub.2] and tryptase. The role of mast cells in the late allergic response has been more difficult to resolve. However, they are thought to play a key role in the development of the chronic inflammatory phase through their production of cytokines and chemotactic factors that lead to the recruitment of other cell types such as eosinophils. [3]

EOSINOPHILS AS EFFECTORS OF THE LATE PHASE

Even if the role of the eosinophil remains somewhat enigmatic, the current view is that it is a proinflammatory cell with a substantial tissue destructive potency. The biological activities exerted by the eosinophil are related to the products released from its granules, including the eosinophil cationic protein (ECP) and the major basic protein (MBP). These two potent cytotoxic proteins have the capacity to kill both mammalian and non-mammalian cells, such as parasites, by making pores in cell membranes, which leads to osmotic lysis. The accumulation and activation of eosinophils in the lungs are governed by the upregulation of adhesion molecules on lung endothelial cells and the production of various cytokines and chemotactic molecules by mast cells and T cells. Of these cytokines, IL-5 seems to play a central role, because it regulates most aspects of eosinophil behaviour, such as growth, apoptosis, adhesion, and secretion. [13] Activation of the endothelium by cytokines such as IL-4 favours their migratio n to the lungs by upregulating the expression of vascular cell adhesion molecule 1 on endothelial cells. [14] Finally, chemokines are responsible for tissue recruitment (see below).

EPITHELIAL CELLS AS GATEKEEPERS

Traditionally, the main function of airway epithelial cells was thought to be preventing the entry of noxious inhaled substances into the body and clearing particulates out of the airways. However, recent …