Extracellular matrix, integrins, and mesenchymal cell function in the airways.

Subepithelial fibrosis is one of the characteristic features of asthmatic airways. The fibrotic response includes an increase in volume occupied by extracellular matrix (ECM) tissue, and a change in the ECM composition favouring wound type collagens, fibronectin and a number of glycoproteins and proteoglycans normally associated with development. The altered ECM is likely to be deposited by the mesenchymal cells (including (myo) fibroblasts and smooth muscle) that are increased in number in asthmatic airways. In turn, the altered asthmatic ECM is likely to influence the function of the resident airway cells, and may be directly responsible for increasing proliferation, migration, ECM synthesis, inflammatory mediator release, and survival of resident mesenchymal cells. Therefore, the deposited ECM may perpetuate the disease phenotype. The different components of the ECM bi-directionally communicate with cells through a family of transmembrane receptors called integrins. Current research has begun to characterize: 1) the particular ECM components altered in airways disease; 2) the breadth of activity of different ECM components on airway cell function; and 3) the particular integrins responsible for mediating these effects. Further understanding of the role of integrins in transmitting responses of ECM in healthy or diseased airways may lead to novel targets for anti-asthma therapy.

Regulation of human airway mesenchymal cell proliferation by glucocorticoids and beta2-adrenoceptor agonists.

Altered rates of cell proliferation play important roles in the pathogenesis of a variety of conditions, including cancer, inflammation and several airway and cardiovascular diseases. One of the most consistently observed changes in asthmatic airways is an increased volume of airway smooth muscle (ASM), that has been explained by proliferation, hypertrophy, extracellular matrix deposition within the smooth muscle bundles, and more recently, the migration of mesenchymal precursor cells to the airways. The best characterised of these is proliferation of ASM cells. In vitro studies suggest that the proliferation is driven by various mitogens, and ECM proteins found in asthma, such as collagen type I. Therefore, we compared the anti-mitogenic actions of two classes of anti-asthma agents, the glucocorticoids and the beta2-adrenoceptor agonists, in ASM cells grown on collagen type I. Culture on collagen type I prevented the anti-mitogenic actions of glucocorticoids, but not beta2-adrenoceptor agonists. In contrast, glucocorticoids are efficacious in regulating ASM production of GM-CSF, whereas beta2-adrenoceptor agonists are without effect. Therefore, combination therapy may have increased efficacy over glucocorticoids alone in controlling remodelling events due to complementary actions of the two classes of compounds.

Proliferative aspects of airway smooth muscle.

Increased airway smooth muscle (ASM) mass is perhaps the most important component of the airway wall remodeling process in asthma. Known mediators of ASM proliferation in cell culture models fall into 2 categories: those that activate receptors with intrinsic receptor tyrosine kinase activity and those that have their effects through receptors linked to heterotrimeric guanosine triphosphate-binding proteins. The major candidate signaling pathways activated by ASM mitogens are those dependent on extracellular signal-regulated kinase and phosphoinositide 3'-kinase. Increases in ASM mass may also involve ASM migration, and in culture, the key signaling mechanisms have been identified as the p38 mitogen-activated protein kinase and the p21-activated kinase 1 pathways. New evidence from an in vivo rat model indicates that primed CD4(+) T cells are sufficient to trigger ASM and epithelial remodeling after allergen challenge. Hyperplasia has been observed in an equine model of asthma and may account for the increase in ASM mass. Reduction in the rate of apoptosis may also play a role. beta(2)-Adrenergic receptor agonists and glucocorticoids have antiproliferative activity against a broad spectrum of mitogens, although it has become apparent that mitogens are differentially sensitive. Culture of ASM on collagen type I has been shown to enhance proliferative activity and prevent the inhibitory effect of glucocorticoids, whereas beta(2)-agonists are minimally affected. There is no evidence that long-acting beta(2)-agonists are more effective than short-acting agonists, but persistent stimulation of the beta(2)-adrenergic receptor probably helps suppress growth responses. The maximum response of fluticasone propionate against thrombin-induced proliferation is increased when it is combined with salmeterol.

Factors controlling airway smooth muscle proliferation in asthma.

Airway smooth muscle proliferation has been the focus of considerable attention, as it is a quantitatively important component of the airway wall remodeling response in asthma and has been suggested as a suitable target for the development of novel anti-asthma agents. Such agents are considered likely to reduce airway hyperresponsiveness and, consequently, airway obstruction, resulting in fewer symptoms and exacerbations. Identifying suitable drug targets has proved an elusive goal, as no dominant molecular mechanism for remodeling has emerged. Moreover, recent findings raise some doubt as to whether smooth muscle proliferation per se is the explanation of the increase in smooth muscle cell number in asthma, with alternative explanations including the proposal that cells migrate either from the interstitial compartment or from a circulating precursor stem cell population. Therefore, drug targeting of migration responses should be considered as an alternative approach to regulating the smooth muscle component of airway wall remodeling.

PPAR gamma ligands, 15-deoxy-delta12,14-prostaglandin J2 and rosiglitazone regulate human cultured airway smooth muscle proliferation through different mechanisms.

The influence of two peroxisome proliferator-activated receptor gamma (PPARgamma) ligands, a thiazolidinedione, rosiglitazone (RG) and the prostaglandin D2 metabolite 15-deoxy-Delta(12,14)-prostaglandin J2 (15d-PGJ2) on the proliferation of human cultured airway smooth muscle (HASM) was examined. The increases in HASM cell number in response to basic fibroblast growth factor (bFGF, 300 pm) or thrombin (0.3 U ml-1) were significantly inhibited by either RG (1-10 microM) or 15d-PGJ2 (1-10 microM). The effects of RG, but not 15d-PGJ2, were reversed by the selective PPARgamma antagonist GW9662 (1 microM). Neither RG nor 15d-PGJ2 (10 microM) decreased cell viability, or induced apoptosis, suggesting that the regulation of cell number was due to inhibition of proliferation, rather than increased cell death. Flow-cytometric analysis of HASM cell cycle distribution 24 h after bFGF addition showed that RG prevented the progression of cells from G1 to S phase. In contrast, 15d-PGJ2 caused an increase in the proportion of cells in S phase, and a decrease in G2/M, compared to bFGF alone. Neither RG nor 15d-PGJ2 inhibited ERK phosphorylation measured 6 h post mitogen addition. The bFGF-mediated increase in cyclin D1 protein levels after 8 h was reduced in the presence of 15d-PGJ2, but not RG. Although both RG and 15d-PGJ2 can inhibit proliferation of HASM irrespective of the mitogen used, only the antiproliferative effects of RG appear to be PPARgamma-dependent. The different antimitogenic mechanisms of 15d-PGJ2 and synthetic ligands for PPARgamma may be exploited to optimise the potential for these compounds to inhibit airway remodelling in asthma. British Journal of Pharmacology (2004) 141, 517-525. doi:10.1038/sj.bjp.0705630

Impact of extracellular matrix and strain on proliferation of bovine airway smooth muscle.

1. The proliferation of airway smooth muscle (ASM) and collagen deposition are major contributors to airway remodelling in asthma that may be an important component of airway hyperresponsiveness. The ratio of collagen to laminin in asthma is increased as a result of fibrosis. 2. We investigated the effects of the extracellular matrix proteins laminin and collagen type I, as well as airway cell elongation (mechanical strain), on the proliferative responses of cultured bovine ASM cells and the impact of these biomechanical influences on glucocorticoid antimitogenic actions. 3. Bovine ASM cells were cultured onto a flexible silastic membrane coated with laminin or collagen. Cells were pretreated with dexamethasone (100 nmol/L) prior to incubation with the mitogen basic fibroblast growth factor (bFGF) for 72 h, at which time cells were enumerated. Cells were either subjected to mechanical strain (cell elongation) by stretching the flexible silastic membrane or were grown under static conditions. 4. Culture on collagen without elongation enhanced ASM proliferation compared with cells grown on laminin. Cells grown on laminin and subjected to 4% elongation did not respond to the mitogenic actions of bFGF. 5. Dexamethasone-mediated inhibition of bFGF-induced proliferation was unaffected by the extracellular matrix on which cells were seeded, or the degree of cell elongation. 6. These data support the hypothesis that the asthmatic airway wall microenvironment (collagen, reduced airway strain) enhances the proliferation of ASM cells.

Collagen-induced resistance to glucocorticoid anti-mitogenic actions: a potential explanation of smooth muscle hyperplasia in the asthmatic remodelled airway.

Glucocorticoids (GCS) inhibit mitogenesis of airway smooth muscle (ASM) cells grown on plastic. We have now evaluated the effects of GCS on proliferation of ASM grown on extracellular matrix proteins (ECM) abundant in noninflamed airways (laminin) and in fibrotic asthmatic airways (collagen type I). Dexamethasone inhibited basic fibroblast growth factor (bFGF)-induced proliferation in cells maintained on laminin, but not collagen. Cells grown on collagen were resistant to the anti-mitogenic actions of fluticasone propionate. In addition, dexamethasone did not inhibit thrombin-induced proliferation. Thus, resistance induced by collagen is not dependent on the mitogen and appears to be a class effect on GCS. The inhibition of bFGF-induced granulocyte-macrophage colony-stimulating factor production was unaffected by the ECM type on which cells were grown. The impaired anti-mitogenic activity of GCS in cells maintained on collagen may be due to a lack of efficacy against the collagen-amplified mitogenesis, rather than any defect in responsiveness that is specific to glucocorticoid receptor mechanisms.