Mast cell tryptase and its role in tissue remodelling

Tissue remodelling is a normal physiological response to injury and constitutes a vital facet of the healing process. In most instances the injury is repaired with complete resolution, but in certain disorders this repair process is associated with the development of fibrosis. Fibrosis is a characteristic feature of chronically inflamed tissue and has, as its hallmark, an excessive deposition of extracellular matrix (ECM) collagen [ 1, 2]. The collagens play vital roles in maintaining the structural integrity, in addition to contributing to the mechanical properties of the lung and are thus critically involved in the remodelling process. In the normal lung, the amount of collagen deposited is tightly regulated to ensure a strict balance between the rate of biosynthesis and degradation. Thus, an inappropriate control of this balance can lead to enhanced collagen deposition with resultant fibrosis. The fibrotic response to injury has been observed in several disorders including acute respiratory distress syndrome [ 3], cryptogenic fibrosing alveolitis [ 4] and sarcoidosis [ 5], where an increase in collagen deposition has been observed. In this issue of Clinical and Experimental Allergy, Abe et al. [ 6] show that the mast cell-specific serine protease, tryptase, stimulates not only the proliferation of dermal fibroblasts but also the production of type I collagen. These findings confirm previous reports that tryptase can enhance collagen synthesis in lung fibroblasts [ 7] and that this effect is mediated via transcriptional activation of the gene for type I collagen [ 8]. Consistent with these findings is the observation that lung tissue from patients with fibrotic lung disease show increased numbers of mast cells, many in a state of partial degranulation [ 9]. Most of the mast cells are closely apposed to proliferating fibroblasts [ 9, 10], pointing to a close interaction between mast cells and fibroblasts, which may be pivotal in the pathogenesis of fibrosis. Elevated concentrations of tryptase and other mast cell products have been found in bronchoalveolar lavage (BAL) fluid from patients with fibrotic lung disease [ 11, 12]. In animal models, fibrotic lesions induced by bleomycin [ 13], asbestos [ 14] and ionizing radiation [ 15], show evidence of mast cell hyperplasia, indicating that the mast cell–fibroblast axis is important in the pathogenesis of not just lung disease but in scleroderma, keloid formation, chronic graft vs host disease and other fibrotic conditions [ 2, 16, 17]. The potential role of mast cell products in the development of fibrosis is further supported by evidence derived from in vitro experiments. Ingestion of mast cell granules by fibroblasts can stimulate collagenase secretion [ 18, 19] while other studies have demonstrated that histamine can stimulate fibroblast proliferation and collagen synthesis [ 20, 21]. Whilst an alteration in the amounts of collagen deposited is characteristic of fibrosis, it is now recognized that the type of collagen synthesized may contribute equally to cellular abnormalities in the lung. Abe et al's paper shows a specific increase of type I collagen in response to tryptase, again confirming a previous report [ 6] where a similar increase in this collagen type was observed in lung fibroblasts. The normal human lung is composed of 65% type I and 30% type III collagen. A specific increase in the amounts of type I collagen, with a concomitant decrease in type III collagen has been reported in idiopathic chronic pulmonary fibrosis [ 22], and in fibrosis associated with atherosclerosis [ 23] and liver cirrhosis [ 24]. Cells derived from belomycin-induced fibrotic animals show a similar increase in type I compared with type III collagen [ 25], in agreement with changes observed in fibroblasts treated with this drug in vitro. The preferential stimulation of type I collagen by tryptase suggests a role for tryptase in chronic fibrosis resulting from ongoing mast cell activation since type I collagen is preferentially found in established fibrotic lesions while type III is characteristic of early fibrosis [ 26]. In general, compliant tissues have a low collagen I:III ratio while less compliant tissues such as that found in pulmonary fibrosis have a higher type I:III ratio. The increased number of fibroblasts observed after tryptase stimulation, initially reported by Ruoss et al. [ 27] and subsequently by Cairns and Walls [ 7], may reflect increased recruitment of these cells by tryptase [ 8] from the surrounding areas, similar to that documented for eosinophils [ 28]. Airway biopsies from asthmatic patients reveal increased numbers of degranulated eosinophils proximal to epithelial cells and just beneath the basement membrane. An increased level of ECP in BAL fluid from patients with fibrotic disease [ 29], together with the enhanced release of ECP from eosinophils challenged with tryptase in vitro [ 28], have confirmed that these cells can be activated by tryptase. Consistent with this observation is the presence of numerous mast cells in close proximity to eosinophils, many in a state of degranulation. Eosinophil products have the potential to contribute to fibrogenesis. For example, neurotoxin derived from human eosinophils can stimulate fibroblast growth and proliferation [ 30] and may lead indirectly to enhanced collagen synthesis. In certain fibrotic disorders, tissue eosinophilia and inflammatory fibrosis co-exist [ 31], suggesting that eosinophil-derived mediators may modulate fibroblast function. In addition, MMP-9, another product of activated eosinophils, may play a crucial role in tissue remodelling in vivo [ 32] by degrading proteoglycan of the ECM and in so doing, facilitate migration of eosinophils through the ECM. Earlier reports in the literature have already highlighted the involvement of tryptase in connective tissue catabolism. This enzyme can degrade type VI collagen microfibrils [ 33], inactivate fibrinogen [ 34], and activate latent collagenase in rheumatoid synovium via an indirect mechanism involving activation of MMP-3 [ 35]. An increase in collagenolytic activity has been observed in BAL fluid from patients with diffuse interstitial fibrosis (DIF) [ 36], where an elevation in levels of tryptase has also been reported [ 11, 37]. Thus, tryptase has the potential to modulate both collagen synthesis and degradation, two processes that are crucial in tissue remodelling and it is the net result of these two processes that determines the outcome. In asthma, the airways are characterized by an increased thickening of the basement membrane, which has been attributed to subepithelial fibrosis [ 38]. Immunohistochemically, the thickened basement membrane has been shown to consist of collagen, together with fibronectin. In fibrotic lungs from human patients and in animal models of pulmonary fibrosis, myofibroblasts are observed in great numbers in the interstitial areas [ 39]. These cells have morphological characteristics of both fibroblasts and smooth muscle cells and are regarded as the key cells involved in wound healing and tissue remodelling. An increased proliferation of cells such as myofibroblasts, with the potential to contract is likely to contribute to further impairment of lung function, which is already severely compromised in the end stages of the disease. In subscribing to this view, it is noteworthy that tryptase has been shown to induce smooth muscle cell proliferation [ 40], which will not only contribute to airway narrowing but also to the development of fibrotic lesions in diseases such as emphysema, bronchiestasis and DIF, where smooth muscle cell hyperplasia is a prominent feature [ 39]. Tryptase has also been shown to modulate epithelial cell function in terms of proliferation and cytokine release [ 41], adding to an ever-increasing repertoire of actions with which this protease has been associated. Enhanced epithelial cell proliferation in response to tryptase is indicative of a potential role in the repair process, but one where an apparent benevolent effect may lead to an over-expansion of the epithelial cell population, particularly in the presence of ongoing mast cell activation. Observations that tryptase can act in an autocrine fashion to degranulate mast cells in vitro [ 42] has provided supportive evidence that persistent mast cell activation may occur in vivo. The unlimited expansion of epithelial cells could result in the sloughing of ‘excess’ epithelial cells into the BAL fluid, characteristic of epithelial desquamation. Accordingly, in areas of epithelium denuded by mechanical trauma, there is evidence of directed migration by epithelial cells to cover the defect and this is often accompanied by cell proliferation. Studies in mice have shown that injury to the epithelium and basement membrane can be positively correlated with the fibrotic process [ 43]. Thus, tryptase-mediated enhancement of ICAM-1 expression and IL-8 release by tryptase could serve to promote the recruitment of leucocytes to the epithelium and the ensuing damage caused by these cells could initiate and contribute to a relentless cycle of injury and fibrosis. Elsewhere, a role for mast cell products has been strongly implicated in the development of liver fibrosis. In particular, tryptase has been demonstrated to stimulate the proliferation of hepatic stellate cells which are important in the fibrotic response in the liver [ 44], whilst an elevation in levels of histamine has been similarly implicated in the pathogenesis of hepatic fibrosis [ 45]. Taken together, the actions of tryptase on a variety of cell types suggest that this enzyme is able to initiate and drive a fibrotic response either by acting directly on fibroblasts or by modulating connective tissue catabolism, both of which are important in tissue remodelling. These findings are in accord with observation of high levels of tryptase and other mast cell products in biological fluids of patients with conditions characterized by abnormal collagen deposition. In fibroblasts, the overall increase in collagen production by tryptase may lead to mechanical impairment in the lung, while the selective increase in type I could contribute to the overall rigidity of the fibrotic lung. All the known actions of tryptase on cells appear to be mediated via its catalytic activity. Indeed, the proteinase-activated receptor-2 (PAR-2) has now been shown to mediate some of the actions of tryptase [ 46], and has reinforced the view that tryptase acts specifically on target cells that express this receptor. PAR-2 is the second member of a new class of G-protein-coupled receptors, with an unusual mechanism of activation. Cleavage of the amino terminus of this receptor by tryptase unmasks a new amino terminus to reveal a ‘tethered’ ligand which is able to activate itself by binding to a different domain on the receptor and thus initiate signal transduction. Thus, there is mounting evidence to propose a new role for mast cell tryptase in tissue remodelling and this suggests that the development of typtase inhibitors may represent a new class of drugs for the treatment of chronic conditions associated with mast cell activation.