American Journal of Respiratory and Critical Care Medicine

Airway inflammation is a central feature of asthma (1-3). The development, progression and/or resolution of airway inflammation in asthma appears critical to the presence and development of symptoms, pulmonary function abnormalities, including bronchial hyperresponsiveness, disease severity, and the target of present-day treatment. It has generally been assumed that abnormalities in airway physiology and immunopathology of asthma are reversible, either spontaneously or as the result of treatment. This assumption has recently undergone reevaluation.

Some patients with asthma have been shown to have irreversible changes in lung function, despite appropriate and aggressive anti-inflammatory therapy (4). In addition, studies have shown that some patients with asthma have an enhanced decline in lung function over time (5, 6). Furthermore, evaluation of airway tissues from patients with asthma has revealed abnormalities in the airway that may be irreversible such as smooth muscle hypertrophy, mucous gland hyperplasia, blood vessel proliferation, and sub-basement membrane collagen deposition (7-10). From these observations has come the application of the term “remodeling” to changes in the airway of some asthmatic patients. Whether airway remodeling is a separate process in asthma, a consequence of the inflammatory response in only some asthmatic patients, or the result of repetitive insults to the airway of the asthmatic patient, is not established but of considerable relevance and importance to the pathogenesis and treatment of asthma.

This workshop was convened to examine features of airway remodeling in asthma, discuss the lung tissues involved in this repair process, explore the model systems available for study, and identify the key questions and research areas of greatest promise and potential to establish the mechanisms and importance of remodeling to asthma.

Airways form in early fetal life, with branching being complete by 16 wk of gestation in humans. By 25 wk of gestation, airway epithelial cell differentiation has occurred, and smooth muscle, cartilage, and submucosal glands have formed. Smooth muscle is functional at birth, contracting spontaneously and responding to pharmacologic agents. The differentiation of airway epithelial cells and morphogenesis of submucosal glands have been explored in detail by fate-mapping techniques. These studies have shown that there are multiple progenitor cells within the epithelium with both fixed and pluripotential capacities for differentiation. They have also shown that all cells of a gland can derive from a clonal population of surface epithelial cells and that a specific transcription factor (Lef1) is expressed at the time of gland formation. Serous and mucous cells express a number of different genes and cell-specific patterns of mucin gene expression have been defined, although the functional consequences remain unclear. Postnatal growth primarily consists of enlargement of existing airway structures, although terminal bronchioles are altered as alveolar formation occurs. It is assumed that the same factors regulating formation of airway components in fetal life are operative during the postnatal period, although much work remains to be done in this area. The consequence of the complex inflammatory process of asthma added to this developmental process can have considerable effects; exactly how these interactions occur and their eventual consequence is not known.

The structural changes that occur in the airways in asthma are the result of inflammation and are variable, but they have common features, including vascular congestion, exudation of fluid and protein from blood vessels, and a migration of inflammatory cells out of the microvessels into the interstitial tissue. In addition, there can be increased secretion of mucus and a sloughing of epithelial cells from the surface into the lumen. When this inflammatory response becomes chronic, there is expansion of the vascular bed and an increase in the amount of collagen tissue within the impaired tissue, leading to remodeling of the structure of the conducting airways. These changes in structure likely lead to functional consequences.

It has been proposed that these structural changes in the airway may have little effect when the airway smooth muscle is relaxed. However, when the airway smooth muscle begins to shorten, even minimal increases in wall thickness will excessively reduce airway caliber. The degree to which the airway is narrowed will determine the intensity of airway caliber narrowing as smooth muscle contraction occurs. Furthermore, because the distal airways have smaller caliber, they are more vulnerable to these changes. This susceptibility to increased airway resistance as smooth muscle contracts occurs in the presence of a thickened airway wall and can also explain the intensity of airway hyperresponsiveness seen in asthma.

Longitudinal studies of asthma remain small in number and often do not take into account therapy issues that may confound results. For example, asthma may not be a single disease, but rather multiple different subtypes, each with different natural histories.

Many epidemiologic studies have tended to group all “asthma” together. Some of these studies suggest that there is a slow progressive fall in pulmonary function in asthma that actually approaches that of chronic obstructive pulmonary disease. However, these studies often do not contain adequate data to evaluate specific groups of individuals. The role of airway remodeling in each of these subtypes is likely to be different. Examination of the various subgroups, both epidemiologically and pathologically, will help determine the factors that lead to progressive disease, as opposed to the less progressive subtypes.

There are numerous aspects of airway remodeling that have been suggested as possible elements influencing progressive severity of the disease. Unfortunately, these studies must be discussed in the context of at least three limiting factors. For the most part, pathologic studies of asthmatics have been done cross-sectionally, as opposed to longitudinally, in a mild (often intermittent) group of asthmatic patients. Finally, the vast majority of studies have been done at the large airway (endobronchial) level, with little regard to the small airways.

A number of structural changes have been linked to chronic asthma and eventual effects of lung function.

1. Thickened lamina reticularis or subbasement membrane (SBM). The relevance of the SBM to severity or progression of disease is less than clear. Studies to suggest a correlation between the severity of the disease and the SBM thickness have generally been done in a surprisingly mild group of patients with “severe asthma,” i.e., patients who often can have their inhaled steroids stopped for a month or more, and patients whose FEV1 is between 70 and 80% predicted receiving no inhaled steroids (3). Studies of SBM thickness have been done at the level of the large airways with almost nothing described in the more distal airways. In a study of asthmatic patients receiving prolonged treatment with inhaled and oral steroids and a FEV1 < 60% predicted (with reversibility), no relationship of SBM thickness (at the large airway level) to severity of disease, level of current inflammation, or use of steroids was found, thus suggesting that prevention of this thickening by ICS may not influence the progression of the clinical symptoms and physiologic changes (11, 12).

2. Submucosal collagen. In the few studies that have evaluated submucosal collagen deposition in asthma, there has not been a consensus result in this area either (13).

3. Tenascin. Tenascin is a nonspecific protein produced in situations where injury and repair processes appear to be occurring. The report of Laitinin and colleagues (14) would suggest that the expression of this protein is increased in symptomatic asthmatics, of mild severity, is increased during times of clinical worsening (pollen season), and is inhibited by inhaled corticosteroids. The presence of this protein in asthma suggests that an ongoing injury and repair process is occurring; however, the relevance of this to long-term “remodeling” or progression to more fixed obstruction is not clear.

4. Elastin. Elastin fibrils have been reported qualitatively to be deranged in asthmatic airways, but there is little information regarding the quantitative or qualitative relationship to clinical or physiologic severity (15).

5. Smooth muscle. Perhaps one of the oldest recognized alterations in the airway structure of asthmatics has been the increase (either through hypertrophy or hyperplasia) of smooth muscle mass. This increase in smooth muscle remains difficult to quantify or qualify in endobronchial biopsy specimens.

6. Matrix metalloproteinases (MMPs). Although these proteinases are not specifically a structural element of the airways, the capacity of this wide range of enzymes to alter the collage, elastin, and other extracellular matrix proteins of the airways is enormous. There is little information on the relationship of MMP levels to severity of disease (16).

7. Blood vessels. Blood vessels have been reported to be increased in autopsy specimens from patients dying of severe acute asthma. This can clearly have implications regarding the propensity for edema in the airways. However, this is more difficult to evaluate in endobronchial biopsies and the relationship to severity again remains elusive.

8. Mucous glands. Another commonly recognized structural alteration which is poorly understood from the standpoint of severity of disease is the increase in mucous glands and goblet cells seen in asthmatic patients.

9. Small airway, parenchymal changes. Although there is quantitative data on inflammation in the lower airways from autopsies, there is almost no information on fibrosis, smooth muscle, or elastin from this portion of the lung (17).

10. “Airway inflammation.” The association between the classic eosinophil/lymphocyte response (and associated cytokines/growth factors) and or “other types” (neutrophilic) of inflammation and any of the above structural changes remains poorly understood.

The biochemical composition of the extracellular matrix provides lung cells with important clues that determine their patterns of response to injury. These clues are principally transmitted via membranes of the integrin family of adhesion receptors (18). For example, fibroblasts can contribute to tissue repair by migrating into sites of injury, by contracting newly formed extracellular matrix, and by assembling secreted matrix proteins into higher order structures such as fibrils. Specific extracellular matrix receptors on fibroblasts play critical roles in orchestrating each of these responses. Fibroblasts lacking the gene for the integrin β1 subunit have been used to determine that β1 integrins are required for optimal migration on a variety of extracellular matrix proteins, including vitronectin, fibronectin, or laminin, in response to serum, platelet-derived growth factor (PDGF), or lysophosphatidic acid. By expressing mutant forms of the β1 subunit lacking specific conserved tyrosine residues, it has been found that phosphorylation and dephosphorylation of specific tyrosines are required for optimal directed migration (19). Matrix contraction in response to serum is dependent on two different integrins, α2β1 and αvβ3. Surprisingly, distinct serum components differentially trigger contraction via each of these integrins. Both β1-integrins and αvβ3 also contribute to assembly of a fibronectin matrix, but, in this case, different integrins are involved in this function depending on the composition of the substrate on which fibroblasts are plated. Together, these findings indicate the complexity of cellular responses to injury and highlight the roles of specific serum components, extracellular matrix proteins, and integrins in directing fibroblasts into areas of injury, and in inducing structural remodeling and elaboration of new extracellular matrix. Clearly, much work is needed to determine how each of the steps used by other cells involved in airway injury and repair is modulated by input from the extracellular matrix.

Lung injury induces an inflammatory response followed by a repair process, which if successful, results in recovery of tissue architecture and function. Alternatively, a prolonged and abnormal repair process may result in progressive fibrosis and end-stage lung disease. Hence, understanding the repair process may provide key information about the pathogenesis of fibrosis, with its deleterious consequences on tissue architecture and function. The importance of inflammation in fibrosis is presumed to be due to the cytokines and other mediators produced by inflammatory cells. The target cells for these mediators have been traditionally assumed to be mesenchymal cells responsible for deposition of extracellular matrix in fibrotic lesions. Recently, a distinct fibroblast phenotype, the myofibroblast, has been identified as a key component of active fibrotic lesions that is largely responsible for collagen production (20). This cell is also an important source of cytokines, including C-C chemokines such as MIP-1α and MCP-1, important in mononuclear cell recruitment, and transforming growth factor-β (TGF-β), which stimulates extracellular matrix production. In animal models, these mediators are produced in eosinophils, macrophages, airway epithelial cells, and myofibroblasts, depending on the stage of development of fibrosis. In vivo and in vitro data suggest that TGF-β itself may be responsible for the differentiation of perivascular adventitial fibroblasts to myofibroblasts (21). Recent evidence also suggests that myofibroblasts are sensitive to apoptotic stimuli, suggesting that their disappearance by apoptosis may be key to resolution instead of progressive fibrosis. As a cell that expresses and responds to inflammatory cytokines and disappears by apoptosis in association with resolution of inflammation, the myofibroblast can be considered an inflammatory cell. Airway remodeling in response to injury and inflammation may involve similar pathogenic events, perhaps involving myofibroblasts derived from fibroblasts present in the adventitia of the airways.

The behavior of airway epithelial cells, the most populous cell in the airway wall, is dramatically affected by specific changes in the biochemical composition of the extracellular matrix. The largest and best characterized family of receptors that could be used to detect such changes is the integrin family, currently composed of 22 known members with diverse and overlapping ligand binding specificity (22). Of the eight integrins known to be expressed on airway epithelial cells, only three of them, α2β1, α3β1, and α6β4, are known to recognize as ligands proteins present in the normal basement membrane underlying these cells (23). Of these, α6β4 has been shown by inactivation of each subunit gene to be critical for maintenance of epithelial integrity, and α3β1 has been shown to be critical for branching morphogenesis and structural organization of the basement membrane. The precise functional role of α2β1 is unknown. The other five integrins expressed by these cells, α5β1, α9β1, αvβ5, αvβ6, and αvβ8, all recognize as ligands proteins that are not only absent from the normal basement membrane but are not present in contact with these cells under normal circumstances (24). The proteins known to be ligands for these integrins, including fibronectin, tenascin-C, osteopontin, and vitronectin, and all greatly enriched in the airway wall in association with injury and inflammation, suggesting that these integrins may actually serve as classic signaling receptors that are expressed on healthy cells so that they will be prepared to detect and respond to spatially restricted changes in the biochemistry of the matrix that occur in response to injury. Observations in mice lacking the integrin αvβ6 support such a role in directing response to injury and inflammation since these mice have dysregulated inflammatory and fibrotic responses, most prominent in the airways and skin (25, 26).

Tissue repair requires an orderly progression of events to reestablish the integrity of damaged tissue. The initial injury sets off a programmed series of interdependent yet separate responses such as reepithelialization and epithelial proliferation, inflammation, angiogenesis, fibroplasia, matrix accumulation, and eventual resolution. During each stage in the process, proteinases act to remove and remodel extracellular matrix components in both epithelial and interstitial compartments, in part to accommodate cell migration and other ongoing events (27). In human lung, a specific MMP, matrilysin (MMP-7), is constitutively expressed by ductal and serous cells of submucosal glands and by ciliated cells. This expression of an MMP in intact, noninflamed mature tissue is unusual. On the basis of in vivo and in vitro observations, it appears that matrilysin is released into the airway lumen where it is likely to play a role in innate host defense. In murine intestine, matrilysin is responsible for activation of prodefensins, explaining the inability of matrilysin null mice to kill pathogenic Escherichia coli and their susceptibility to usually nonlethal doses of Salmonella. In response to lung injury, matrilysin is markedly induced in upper airway epithelial cells, and expression is also induced in alveolar type II cells in cystic fibrosis and diffuse alveolar damage. Other MMPs, specifically collagenase-1, stromelysin-1, and the 92 kD gelatinase, are not produced in healthy or injured respiratory epithelia, suggesting a prominent role for matrilysin in the injury-mediated responses of the lung. Interestingly, in response to tracheal injury, migrating epithelial cells reverse the polarity of matrilysin expression and direct it toward the basal surface. Reepithelialization of the injured trachea is essentially blocked in matrilysin null mice. Together, these findings suggest that matrilysin has diverse and important functions in the lung epithelium in response to injury, one of which is to facilitate cell migration.

On the basis of studies of airway epithelial cells and immune cells in vitro and in vivo in mice and humans, it is apparent that mucosal immunity depends on the level of epithelial expression of a subset of immune-response genes typified by the cell adhesion molecule ICAM-1 and the chemokine RANTES (28). In the case of ICAM-1, gene expression is governed by the formation of a transcriptional enhanceosome that includes the Stat1 and Sp1 transcription factors and the CBP/p300 coactivator (29). For RANTES, expression is also critically controlled at the post-transcriptional level based on a distinct mRNA turnover element and mRNA-binding factor. Some respiratory viruses selectively trigger these systems for transcriptional initiation and mRNA stabilization (either directly or via production of interferons) and, thereby, are eliminated through the programmed death of the host cell. In other cases, however, viruses have engineered proteins (analogous to dominant negative mutations) to interrupt signal transduction and evade the immune response leading to epithelial apoptosis. Interestingly, the same systems appear to be selectively activated in epithelial cells in asthma even without an apparent viral infection. This finding raises the possibility that viral remnants are left in epithelial cells of asthmatic subjects, and these remnants no longer cause infection but still modify the immune response. A goal of future research is to define the genetic building blocks that allow epithelial immune-response genes to successfully mediate host defense and to determine how alterations in these same genetic components lead to inflammatory airway diseases.

Two phenomena are critical to the development of the mucus hypersecretion characteristic of chronic airway diseases. First, the occurrence of mucous cell metaplasia by which (pleiomorphic) cells of the airway surface epithelium differentiate to become mucous cells and, second, the occurrence of collagen degradation beneath the epithelium, permitting epithelial cells to migrate downward to form new or enlarge existing mucous glands. The understanding of such events in molecular terms requires the use of biochemical endpoints. Mucin and MMPs were chosen as genes whose activation is arguably rate-limiting for mucous metaplasia and gland enlargement, respectively. Pseudomonas aeruginosa (a pathogen of cystic fibrosis) directly upregulates mucin expression in epithelial cells. On the basis of results using a reporter driven by a 1.4 kb 5′ flanking region of the MUC-2 promoter, this response is mediated by a signaling pathway the activates NFkB via a Scr-dependent shc I, ras/raf/mekk/erk and pp90rsk pathway. In addition to bacterial stimuli, tobacco smoke has been shown to be capable of directly activating transcription of an additional mucin gene (MUC 5AC) through both a Src- and epidermal growth factor (EGF) receptor kinase-dependent pathway (30). Identification of the rate-limiting signaling molecules in these pathways could be clinically important, identifying new targets for drug intervention to block mucus hypersecretion. In relation to events necessary for epithelial downmigration, an inducer of metalloproteinase production (Emmprin) has been identified in bronchial epithelial cells and lymphocytes in inflamed airways that upregulates production of several MMPs in fibroblasts. Current work is examining the signaling pathways leading to stimulation of Emmprin transcription, again with the hope of developing new drug targets to prevent or reverse mucous gland hypertrophy and hyperplasia.

Chronic inflammation and subepithelial fibrosis are hallmarks of the asthmatic airway (31). Various lines of evidence suggest that these processes are intimately linked with inflammatory mediators inducing repair responses designed to restore tissue integrity. When this repair response is dysregulated, these mediators are believed to generate the features of the remodeled airway. A variety of mediators have been implicated in the generation of tissue fibrotic responses. Among the best studied is TGF-β1, a multifunctional pleiotropic peptide produced by alveolar macrophages and other cells, which regulates the growth and differentiation of a wide variety of tissues, including epithelial cells, mesenchymal cells, and cells of hematopoietic origin. In the lung, TGF-β1 regulates branching morphogenesis, the accumulation of extracellular matrix, and the generation of tissue fibrosis. The latter is the result of direct activation of transcription of target genes, including collagens, fibronectin, and plasminogen activator inhibitor-1, by TGF-β1. Signaling pathways from TGF-β involves the activation of novel cytoplasmic proteins called Smads by cell membrane receptor serine-theronine kinases. Phosphorylated Smad2 and Smad3 associate with a common mediator, Smad4, and are translocated to the nucleus where they participate in transcription complexes (32, 33). A variety of other growth factors appear to signal through receptor tyrosine kinases, which can also activate the Smad signaling pathway. Additional investigations are required to define the relative importance of and interactions between these and other mediators in the generation of the fibrotic, metaplastic, proliferative, and other features of the repairing and remodeled airway. The receptors and signal transduction pathways that are involved in mediating the effects of these mediators need to be identified, as well as delineating the benefits and possible adverse consequences that result from the inhibition of these responses. Lastly, the relationship between normal and pathologic remodeling needs to be elucidated and the factors that determine which response predominates need to be defined.

Investigators in the field of airway remodeling are faced with a variety of challenges. One stems from the fact that the majority of investigations performed to date have focused on models of acute airway injury and repair. Although these studies have proven useful in the characterization of the early events that mediate airway inflammation and physiologic dysregulation, they have not appropriately modeled the consequences of chronic airway inflammation and repair. Another challenge relates to the complexity of the remodeling response and our inadequate knowledge of the in vivo effector functions of mediators that have been implicated in the many facets of the asthmatic diathesis. The latter is being compounded by the flood of knowledge that has and continues to be provided by the genome project(s) and DNA-based approaches such as differential display and EST-based gene chip and array analyses. These approaches have identified genes that may be involved in the generation of the asthmatic diathesis without providing adequate insight into the roles that they play in in vivo responses.

Relevant antigen- and/or virus-driven models of chronic airway remodeling are required if appropriate scientific progress is to be made in this area. In addition, transgenic approaches have, and should continue, to be useful in attacking these issues. The utilization of mice with targeted genetic mutations is now routine in studies of biologic homeostasis and has been used successfully in models of acute airway inflammation (34, 35) and will likely be equally useful when combined with appropriate models of airway remodeling. A number of laboratories have demonstrated that the Clara cell 10 kD protein promoter (CC10; also called Clara cell secretory protein [CCSP]) can be used to overexpress known or newly discovered genes in the murine airway. This allows investigators to define and correlate pathologic, immunologic, and physiologic in vivo effector function of a given transgene. This approach successfully highlighted the ability of IL-4 to generate airway mucus metaplasia (36), IL-5 to generate eosinophilic inflammation and airway remodeling (37), IL-9 to induce airway mast cell hyperplasia (38), and IL-11 to induce airway fibrosis, myocyte, and myofibroblast hyperplasia and physiologic dysregulation (39).

The limitations of transgenic modeling with conventional overexpression and knockout approaches are very evident. Targeted gene disruption cannot be employed to study the importance of a protein in an adult phenotype if the same protein plays a critical role in development. The redundancy of biologic processes can also allow compensatory mechanisms to appear in knockout animals that may not be relevant to the wild-type phenotype. In addition, in overexpression modeling, transgene expression can be initiated in utero and proceed chronically thereafter. This makes the differentiation of developmental and adult phenotypic features difficult and makes the modeling of waxing and waning diseases such as asthma problematic. Recent studies, however, have demonstrated that mice can be produced in which a gene can be disrupted after birth or after the lung is fully developed in an adult animal (40). In addition, overexpression transgenic technology has now reached a point where genes can be selectively expressed in murine lungs at different points in time during development (41), and inducible dominant-negative approaches are a reality. These new approaches will provide impressive insights into the genes that are involved and the importance of development in the generation of the remodeling response. Maximal insight into pathogenesis will be obtained when appropriate chronic models of remodeling and externally modulatable transgenic approaches are combined and state-of-the-art immunologic, pathologic, and physiologic assessment techniques are utilized to address crucial issues in airway biology.

The asthmatic airway is characterized by eosinophil infiltration and activation with evidence of degranulation. Eosinophils are recruited to the airway after allergen exposure and during asthma exacerbations. Many factors contribute to eosinophil recruitment to the airway, including cytokines (i.e., IL-3, IL-5, and GM-CSF), chemokines (eotaxin and RANTES), and adhesion molecules (ICAM-1 and VCAM-1). Moreover, the eosinophil contributes to the development of acute tissue inflammation and injury through release of granular enzymes, toxic oxygen products, and cytokines. Eosinophil survival is an important variable in this cell's capability and capacity to cause airway injury. Recent evidence has shown that the interaction of eosinophils with matrix proteins (i.e., fibronectin and laminin) prolongs eosinophil survival and that this effect is due, in part, to the generation of and autocrine/paracrine effects of GM-CSF produced by local eosinophils (42). At the present, there is little conclusive evidence as to how the eosinophil participates in the repair or remodeling process in the airway. However, since eosinophils are known to produce metalloproteinases, collagenase, and growth factors (i.e., TGF-β and PDGF), which regulate the proliferation and matrix production of fibroblasts and other stromal tissues, there is reason to believe that they play an important role. Collectively, these isolated data suggest that eosinophils can contribute to acute and persistent inflammation. Additional information is required about the pathways of eosinophil activation in the airway, the processes that are responsible for chronic eosinophilic inflammation, the interactions between eosinophils and various matrix tissues, the role of eosinophils in the generation of the remodeling phenotype, and the mechanisms of these effector functions.

Mast cell hyperplasia is a characteristic feature of the asthmatic airway and is a frequent pathologic finding in tissue fibrotic and remodeling responses (43). In vitro and in vivo studies have also demonstrated that mast cells and their mediators can play an important role in chronic inflammatory and remodeling disorders. The contribution of mast cells to the airway remodeling response in asthma, however, is incompletely understood. What is known is that mast cell components, in particular β-tryptase, the major protein component of the mast cell secretory granule, are present at high levels in airway fluid and presumably in airway tissues. β-Tryptase is also poorly inhibited by the classic serine protease inhibitors present in human fluids and tissues and, thus once secreted, may result in persistent enzymatic activity. In vitro biologic activities of tryptase include metabolism of fibrinogen and certain neural peptides, the generation of C3a, activation of prostromolysin, activation of PAR-2 on cell surfaces, and the stimulation of fibroblast proliferation and collagen production (44). Interestingly, certain proteolytic activities of tryptase are optimal at acidic pH such as that encountered at sites of inflammation and wound healing. Additional studies are required to characterize the in vivo role of mast cells, the mast cell products that mediate these in vivo effects, the stimuli that induce mast cell activation and degranulation during the remodeling response and the signal transduction pathways that mediate these important remodeling effector functions.

Growth and remodeling of the microvasculature occurs in many chronic inflammatory diseases and tumors. However, little is known about these processes in the airways. The sparse literature on vascular remodeling in the airway wall seems not to be a sign of the infrequency of this process in respiratory pathology but instead of the relatively recent recognition that it is part of the pathophysiology of airway disease. It has long been recognized that, in fatal asthma, the airway mucosa is edematous and contains dilated, congested blood vessels. More recent morphometric studies have confirmed these early observations and have shown that enlarged, congested mucosal blood vessels contribute to the increased airway wall thickness in asthma. Changes in the number or caliber of mucosal blood vessels are functionally important because even modest increases in airway wall thickness can amplify the effacement of the airway lumen produced by a fixed amount of bronchial muscle contraction.

There are at least two types of vascular remodeling, sprouting angiogenesis and microvascular enlargement. In sprouting angiogenesis, endothelial cells proliferate and migrate to form new vessels. In microvascular enlargement, which is sometimes called nonsprouting angiogenesis, vessels can enlarge circumferentially as a result of the proliferation of endothelial cells and other elements of the vessel wall. The new vessels are larger than the original vessels, and thus may appear to be dilated or congested, but they may not be more numerous. This type of vascular remodeling may occur in asthma, where vessel enlargement and endothelial cell proliferation may occur in the absence of an apparent increase in vessel number. The characteristics of the vascular remodeling in the asthmatic airway are poorly understood. It is clear, however, that both sprouting angiogenesis and microvascular enlargement can be seen in airway pathologies. This is nicely illustrated in studies of mycroplasma infection in strains of mice and other rodents that are susceptible or resistant to the infection (45). Airway vascularity is increased in susceptible and resistant strains after infection. However, the type of vascular remodeling is strikingly different. In the airways of resistant mice, the numbers of vessels are increased. In contrast, neither the number nor the length of vessels change in susceptible mice. Instead, vessel diameter and endothelial cell number increase and the proportion of venules increases with a corresponding decrease in the number of capillaries. Thus, the same stimulus can result in blood vessel proliferation or enlargement in the airway mucosa depending on the genetic determinants of the host inflammatory response. Additional investigation is required to define the vascular abnormalities in the asthmatic airway, the factors that contribute to the pathogenesis of these alterations, and the factors that regulate vascular permeability at sites of airway inflammation. In addition, the contribution that vascular abnormalities make to the perpetuation of asthmatic airway inflammation and the generation of asthmatic symptomatology and physiologic abnormalities needs to be clarified.

An increase in airway smooth muscle mass caused by varying degrees of hypertrophy and hyperplasia is well described in the asthmatic airway (46, 47). The importance of airway smooth muscle cells in the regulation of airway tone is well described. The ability of smooth muscle hyperplasia and hypertrophy to cause exaggerated physiologic responses to contractile agonists and play a role in the generation of airway hyperresponsiveness is being increasingly appreciated (46, 48). Recent studies have also demonstrated that airway smooth muscle cells contribute to the orchestration and perpetuation of tissue inflammation and are important sources of extracellular matrix in the asthmatic airway. In keeping with the importance of the smooth muscle abnormalities in the generation of air flow limitation in asthma, studies have focused on the pathways that are responsible for smooth muscle proliferation. These studies demonstrated that smooth muscle mitogens fall into two broad categories, those that activate receptor tyrosine kinases and those that are coupled to heterotrimeric GTP binding proteins (49). PDGF and EGF activate receptors with intrinsic kinase activity and have been shown to induce myocyte proliferation (50). Contractile agonists such as thrombin, which mediates its effects through G-protein-coupled receptors, could also induce smooth muscle cell mitogenesis (51). Interestingly, endothelin-1, thromboxane-A2, and leukotriene D4 alone have little effect on smooth muscle cell proliferation but do increase the growth responses induced by thrombin and EGF. Recent evidence suggests the phosphoinositol-3-kinase (PI3 kinase) plays a role in the regulation of airway smooth muscle cell proliferation and that the ribosomal S6 kinase family (pp70s6k) may be critical in downstream signaling events for mitogen-induced activation of P13 kinase and smooth muscle cell proliferation.

Additional investigations are required to define stimuli that induce smooth muscle cell proliferation and hypertrophy, the signal transduction pathways that mediate the effects of these stimuli and the sites at which interventions can be targeted.

On the basis of the presentations and general discussion of the topics reviewed above, the following recommendations for future research directions were made.

1. Natural History of Airway Remodeling in Human Asthma

A number of critical issues need to be addressed. For example, when is airway remodeling seen in human asthma? Is the remodeling response similar in all asthmatics? Do they all manifest the same degree of mucous metaplasia, airway fibrosis, etc? Alternatively, do different patients manifest qualitatively and quantitatively different responses? Do some get fibrosis without metaplasia, others get metaplasia without fibrosis, and others never manifest remodeling at any point in time during the course of their disease? In addition, can subpopulations that do and do not develop specific features of the remodeling response be isolated and studied to determine what correlates with the development of the remodeling response, pathologically and physiologically, and what are the processes that induce this response in vivo at the cellular and molecular level? What pathologic processes correlate with partial and complete irreversibility of airways obstruction? To what degree can fixed airway obstruction be attributed to refractory inflammation versus airway fibrosis or other pathologic processes? Can better tools be developed for assessing the airway remodeling response at the population level (for example, enhanced imaging techniques of he airway wall including CT-scan, MRI, etc.)? Can in vivo correlates of the remodeling response be established as valid biomarkers with the ability to predict and/or correlate with disease progression or therapeutic responsiveness (for example, BAL hyaluronic acid levels, collagen fragments, elastin degradation product, etc.)? What is the role of infection, either in early life or repeated infections in later life, in the development of the various features in the remodel airway?

2. Biology of the Remodeled Airway

Deeper insights into the biology of the remodeled airway are required. In particular, what are the mechanisms responsible for the chronicity of airway inflammation? How does normal inflammation resolve in the airway and what role do anti- inflammatory cytokines and processes like apoptosis play in this process? Is the integrity of these pathways intact in the asthmatic airway? Is the chronic inflammation characteristic of the asthmatic airway the result of chronic antigen exposure or inadequate/incomplete inflammation resolution or a combination of both? What processes regulate airway mucus and mucin production and mucous and serous cell hyperplasia and differentiation? The vascular responses to airway injury and inflammation need to be defined. What processes regulate airway vascular permeability and what is the role of vascular abnormalities in airway remodeling and in the generation of asthmatic symptoms and physiologic of dysregulation?

3. Pathogenetic Events involved in Airway Remodeling

Additional insight is required into the pathogenesis of the unique features of the remodeled airways, including subepithelial fibrosis, mucous metaplasia, secretory cell hyperplasia, hyperplasia and hypertrophy of airway smooth muscle cells, hyperplasia of myofibroblasts, chronic inflammation, and the vascular abnormalities seen in asthmatic tissues. In addition, what mediators, mediator-mediator interactions, cell-cell interactions, signal transduction pathways, and effector mechanisms are responsible for these biologic processes? Are local neural mechanisms altered in remodeled airway structures? What are the consequences of augmenting and/or inhibiting each of these responses (beneficial and/or detrimental) and the physiologic consequences of these responses? Particular attention needs to be directed at comparisons of modeling systems and patient populations that manifest or do not manifest the different phenotypic features of remodeled airway. Deeper insight is required into effector mechanisms and cell-cell interactions that play a prominent role in the asthmatic airway. In particular, what are the mechanisms by which eosinophils alter fibroblast, myofibroblast, blood vessel and mucous cell function and local matrix accumulation? Are similar interactions occurring between mast cells and inflammatory and stromal cell populations? What are the processes that regulate myofibroblast proliferation and activation and to what extent do myofibroblasts versus fibroblasts contribute to local fibrotic responses?

4. Better Animal Modeling Systems

Our knowledge of the cellular and molecular events that are involved in airway remodeling is limited, in a major way, by our lack of appropriate, relevant models of chronic airway inflammation and tissue alteration. Better antigen and/or virus-driven models and more sophisticated transgenic approaches are required. Optimal scientific insights will be obtained when appropriate chronic models are combined with sophisticated transgenic approaches and state-of-the-art techniques involving pathology, immunology, and physiology to address crucial issues in airway biology. Computerized models of asthma that explore the impact of various structural alterations on airway function need to be developed.

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Correspondence and requests for reprints should be addressed to Susan Banks-Schlegel, Ph.D., Division of Lung Diseases, National Heart, Lung, and Blood Institute, Two Rockledge Center, Suite 10018, 6701 Rockledge Drive, MSC 7952, Bethesda, MD 20892-7952.

List of participants: Susan Banks-Schlegel, Ph.D., Carol Basbaum, Ph.D., Jerome Brody, M.D., William Busse, M.D. (co-chair), Jack Elias, M.D. (co-chair), James Hogg, M.D., Michael Holtzman, M.D., Donald McDonald, M.D., John McDonald, M.D., Ph.D., Deane Mosher, M.D., Patricia Noel, Ph.D., Reynold Panettieri, Jr., M.D., Peter Pare, M.D., William Parks, Ph.D., Sem Hin Phan, M.D., Anita Roberts, Ph.D., Lawrence Schwartz, M.D., Dean Sheppard, M.D. (co-chair), Julian Solway, M.D., and Sally Wenzel, M.D.

Workshop supported by the Division of Lung Diseases, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland, September 14–15, 1998.


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