Pharmacological regulation of mediator release from human mast cells

Asthma is a highly complex inflammatory disorder which involves the participation of a range of effector cells including macrophages, mast cells, neutrophils, lymphocytes and eosinophils. An understanding of the interactions between these cells and their mediators within the inflammatory environment is important for the development of novel therapeutic interventions. In order to dissect these complex pathways, a thorough knowledge of the biochemistry and pharmacology of the individual cells is essential. Mast cells are recognized as major players in immediate hypersensitivity reactions. Activation of these cells via crosslinking of the high affinity IgE receptor (FCεRI) leads to the release of both preformed (e.g. histamine and tryptase) and newly generated (e.g. sulfidoleukotrienes [LTs]) and prostaglandin D2 (PGD2). These mediators are responsible for the symptoms of the immediate response. Relatively recently it has been demonstrated that human mast cells have the ability to produce a number of cytokines including TNFα, IL-4, IL-5, IL-6 and IL-8. These findings suggest that the role of the mast cell extends beyond the immediate response, and that these cells contribute the late asthmatic response and are involved in the control of chronic inflammation. Understanding the mechanisms that modulate mast cell activity provides a rationale for drug therapy of human hypersensitivity. Prospective drugs for the treatment of human allergic diseases are traditionally assessed using animal models of anaphylaxis. Rat peritoneal mast cells are plentiful and can be purified easily. Thus, these cells are often used in assessing the pharmacological actions of novel compounds. However, the functional heterogeneity of mast cells from different species and from different tissues in the same species has called into question the usefulness of animal models. Some examples are given below. β2-adrenergic receptor agonists suppress immunologically induced release of histamine and eicosanoids from dispersed human lung, skin and tonsillar mast cells and inhibit TNFα release from human skin mast cells [ 1, 2]. In this issue of the journal, Shichijo et al. demonstrated that the release of histamine, LTs and PGD2 from cultured human mast cells is also inhibited by β2-agonists [ 3]. However, β2-agonists do not inhibit mast cell mediator release from rat peritoneal mast cells (RPMC) despite the identification of abundant β2-adrenergic receptors on these cells [ 4–6]. This suggests that functional heterogeneity extends beyond differential expression of cell surface receptors. The anti-allergic agents sodium cromoglycate and nedocromil sodium cause a dose-dependent inhibition of immunologically stimulated histamine release from RPMC. These agents show reduced activity on peritoneal mast cells from the hamster and are completely ineffective on these cells in the mouse [ 7]. Human mast cells from different tissue sites demonstrate a range of responses to the inhibitory action of sodium cromoglycate and nedocromil sodium. These anti-allergic agents inhibit IgE-mediated histamine and PGD2 release using mast cells dispersed from human lung, tonsillar and adenoidal tissue [ 8]. However, human skin mast cells are unresponsive to both these compounds [ 8]. Histamine, PGD2 and LT release induced by anti-human IgE from cultured mast cells is inhibited by sodium cromoglycate (1 mmol/L) as described by Shichijo et al. [ 3]. The long-term efficacy of sodium cromoglycate and nedocromil sodium is also dependant on the mast cell model used, with both agents exhibiting tachyphylaxis in lung and tonsillar mast cells but not in mast cells dispersed from adenoidal and intestinal tissue [ 8]. The non-selective phosphodiesterase inhibitor, theophylline, inhibits histamine release from human basophils, rat peritoneal mast cells and cultured murine mast cells [ 9–11]. However, at concentrations of up to 100 μmol/L, theophylline does not inhibit IgE-stimulated histamine release from either human lung fragments or cultured human mast cells [ 3, 12]. Rolipram, a phosphodiesterase 4 inhibitor, inhibits histamine release from cultured human mast cells at high concentrations (100 μmol/L) but does not cause significant inhibition of histamine release from human lung fragments at this concentration [ 3, 12]. This heterogeneity in the response of mast cells to anti-allergic agents demonstrates the problems involved in using animal models for the screening of mast cell-stabilizing compounds and emphasizes the necessity of conducting such studies on human preparations. Further, given the heterogeneity that exists between human mast cells from different anatomical sites, the model used must mimic the disorder targeted by the prospective therapeutic agent. However, there are significant practical difficulties involved in such an approach. Complicated processes are required to isolate even a small number of human mast cells, most often from surgical specimens. The use of mast cells from surgical specimens presents problems in itself. These specimens are often taken from macroscopically normal regions of diseased tissue and changes in mast cell responsiveness due to the disease state cannot be ruled out. Cultured human mast cells have recently become available [ 13]. These cells are raised from cord blood mononuclear cells cultured in the presence of stem cell factor, interleukin-6 and prostaglandin E2. Such cells express FcεRI and, following stimulation with anti-IgE, release histamine, LTs, PGD2 and the inflammatory cytokine TNFα [ 13, 14]. The secretory responses of these cells to a range of stimuli resemble those of human lung mast cells [ 13]. In this issue of the journal, Shichijo et al. report the response of these cultured human mast cells to anti-asthmatic drugs [ 3]. Their findings suggest that the pharmacological response of the cultured mast cells to anti-asthma drugs is similar to that of the human lung mast cell. This study adds to the evidence that cultured human mast cells provide a useful and valid model for analysis of human lung mast cell pharmacology. The importance of this method in providing a ready supply of cells closely resembling the human lung mast cell, in surface receptor expression, pharmacological responses and mediator release characteristics, is clear. However, the value of these cells in assessing mast cell- directed therapies for asthma must be tempered by the fact that two distinct populations of lung mast cells are probably involved in the asthmatic response. One population, located beneath the basement membrane close to blood vessels and the fibrous stroma, may be obtained by enzymatic or mechanical dissociation of whole lung tissue and represent what are commonly termed human lung mast cells (HLMC). The second population of cells is situated between the basement membrane and the epithelium. These cells may be recovered by bronchoalveolar lavage (BAL) and are termed BAL mast cells. The location of BAL mast cells suggests that they come into immediate contact with inhaled allergen and thus mediate the initial stages of the asthmatic response, while HLMC most likely play a role following the development of chronic disease. The significant increase in BAL mast cell numbers in asthmatic subjects and the correlation of mast cell number to disease severity indicate that these cells do play an important role in asthma. Studies have indicated that these two types of pulmonary mast cells exhibit distinct differences in their functional properties. Nedocromil sodium is an order of magnitude more effective in suppressing histamine release from BAL mast cells than from HLMC [ 15]. Furthermore, tachyphylaxis to the inhibitory effects of nedocromil sodium and sodium cromoglycate occurs only in the HLMC population. The lack of tachyphylaxis to nedocromil sodium and sodium cromoglycate in BAL mast cells suggests they more accurately reflect the in vivo situation given the mode of use of these compounds in the management of asthma. More recently, additional functional differences have been demonstrated. The neuropeptides substance P, neurokinin A and calcitonin gene-related peptide (CGRP) induce histamine release from human BAL mast cells but not from HLMC [ 16, 17]. In this respect, the BAL mast cells are closer to skin mast cells than HLMC. Furthermore, BAL mast cells from patients with asthma and chronic non-productive cough have an increased response to substance P and CGRP, respectively [ 17, 18]. These data have identified a potential role for neuropeptide/mast cell interactions in the pathophysiology of these respiratory diseases, which would not have been revealed through the study of dispersed HLMC. Another example of the functional heterogeneity between these two cell populations is seen in the response to adenosine. Adenosine is well established as a modulator of mast cell activity in vitro. While alone adenosine does not elicit histamine release, it has been shown to either inhibit or potentiate anti-IgE-induced histamine release from HLMC when added prior to or following challenge, respectively [ 19]. However, adenosine alone can induce histamine release from human BAL mast cells [ 20]. Instillation of adenosine monophosphate directly into an airway segment of asthmatic subjects leads to a rapid reduction in airway calibre with concomitant increases in levels of histamine, PGD2 and tryptase in BAL fluid [ 21]. This again suggests that BAL mast cells more accurately reflect in vivo lung responses. The distinct functional difference between mast cell populations in the lung emphasizes the need for careful consideration of the model used for testing potential therapies. Human BAL mast cells also provide an example of the problems involved in using human cell models. In order to characterize fully the pharmacology of these cells a pure population is required. However, mast cells represent barely 0.1% of the total cell population in BAL fluid from normal human lungs. Although this percentage is significantly increased in BAL fluid from asthmatic subjects, the technical difficulties involved in obtaining isolated BAL mast cells are substantial and the purification of these cells has yet to be reported. However, given that phenotype of a mast cell is utterly dependent on its microenvironment it should be possible to raise mast cells of various phenotypes by altering the culture conditions described by Saito et al. [ 13]. Such an approach may help us gain insight into the underlying mechanisms for mast cell activation and inhibition in asthma.