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Airway smooth muscle cells Andrew J. Halayko1–6 and Pawan Sharma1,5,6 Departments of 1Physiology, 2Internal Medicine, and 3Pediatrics and Child Health, 4 Section of Respiratory Disease, and 5CIHR National Training Program in Allergy and Asthma, University of Manitoba, Winnipeg, MB, Canada 6 Biology of Breathing Group, Manitoba Institute of Child Health, Winnipeg, MB, Canada

Introduction

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Airway smooth muscle (ASM) is a primary determinant of lung physiology because its ability to contract affords it dynamic control over airway diameter. Research for improved ways to treat asthma has long focused on factors that determine ASM contractility, including mechanical properties and responsiveness to spasmogens and relaxing factors. However, the scope of research on ASM biology and function has broadened greatly in the past decade and a half, embracing the now-recognized dynamic, multifunctional behavior that equips myocytes to be directly involved with the inflammation and fibroproliferative wound healing that underpins the development of airway remodeling. These cells can control airway diameter both acutely, via reversible contraction, and chronically, by driving fixed changes in structure and function properties of the airway wall. We review the spectrum of ASM phenotype and function in the context of pathobiology in obstructive airways disease and describe the impact of some emerging therapies on the properties of these cells.

Phenotype plasticity and functional diversity of smooth muscle Work over the past 40 years on arterial smooth muscle in vitro and in vivo led to the concept that a dynamic phenotypic spectrum of differentiated smooth muscle

cells exists, ranging between so-called “contractile” and “synthetic/proliferative” states [1,2] (Figure 12.1). Contractile myocytes exist in smooth muscle tissues throughout the body and are chiefly designed to stiffen, shorten, or relax in response to chemical and mechanical signals. Synthetic/proliferative cells exhibit what has also been called an immature phenotype that is characterized by a tendency to proliferate and synthesize extracellular matrix (ECM) and other biologically active proteins. Only after 1980 were primary cultured canine tracheal smooth muscle cells first described [3,4]. Nine years later [5,6] primary human ASM cell cultures were reported, leading to studies on the expression and function of physiologically relevant receptors, ion channels, and factors that regulate cell proliferation [7]. Though investigators recognized that airway myocytes likely modulated from a contractile phenotype when placed in culture, it was not until the late 1990s that systematic descriptions of phenotype and functional switching were published [8,9]. This provided impetus to examine the breadth of ASM function. Studies on biologic and molecular mechanisms that regulate phenotype and function have moved to the forefront, recognizing that ASM cells contribute broadly to the pathobiology of obstructive airways disease.

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Phenotype plasticity Phenotype plasticity is a feature of differentiated smooth muscle cells and is manifest as the reversible

Inflammation and Allergy Drug Design, First Edition. Edited by Kenji Izuhara, Stephen T. Holgate, Marsha Wills-Karp. © 2011 Blackwell Publishing Ltd. Published 2011 by Blackwell Publishing Ltd.

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“Synthetic/proliferative”

“Contractile”

Modulation

Maturation

• Cell division • Migration • ECM synthesis • Cytokine synthesis • Abundant synthetic organelles • Few caveolae • Few contractile proteins • Intermediate filaments are predominantly vimentin

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• Contracts • Cell hypertrophy • ECM synthesis (?) • Abundant contractile apparatus • Many caveolae • Abundant contractile proteins • Intermediate filaments are predominantly desmin

Figure 12.1 Reversible phenotypic plasticity of airway smooth muscle. Modulation of myocytes to a “synthetic/ proliferative” state occurs in primary culture and with exposure to mitogens and extracellular matrix (ECM) proteins such as fibronectin and collagen-1. Maturation to a “contractile” state occurs in cell culture at a high cell

density in response to mitogen withdrawal, exposure to insulin, and adherence to laminin-rich ECM. “Contractile” myocytes predominate in adult tissues and exhibit variable degrees of maturation based on expression of molecular markers. Key functional, ultrastructural, and biochemical features of the extreme phenotypic states are shown.

modulation and maturation of myocytes both in vitro and in vivo [10]. Cell culture studies have provided insight on the control of phenotype expression and its regulation by stimuli including growth factors, G protein-coupled receptor ligands, and ECM proteins. Depending on the stimulus, myocytes can be induced to modulate to a synthetic/proliferative phenotype or to undergo maturation to a functionally contractile state. In primary culture, contractile ASM cells undergo phenotype modulation [8], which promotes acquisition of a functional synthetic form marked by abundant organelles for protein and lipid synthesis and numerous mitochondria (Figure 12.1). The cells exhibit a high proliferative index but lose responsiveness to contractile agonists and contractile apparatus proteins [2,11]. The induction of primary cultured airway myocytes to a contractile phenotype is called maturation (Figure 12.1). Maturation is marked by the reaccumulation of contractile apparatus proteins, reacquisition of responsiveness to physiologic contractile agonists, and reduced numbers of synthetic organelles [9,12]. An additional ultrastructural

feature of contractile myocytes is the presence of abundant caveolae and their unique structural proteins, caveolin-1 and -2, which modulate signal transduction [13,14]. Molecular markers for the contractile phenotype include smooth muscle (SM) α-actin, SM-myosin heavy chain, calponin, h-caldesmon, SM22, desmin, caveolin-1, β-dystroglycan, and integrin α7 [8,9,15– 19]. Regulation of the expression of these proteins requires coordinated control of gene transcription and translation [10,20]. Intracellular signaling cascades, including Rho/Rho kinase and phosphatidylinositide 3 kinase (PI3K)-dependent pathways play a central role in smooth muscle specific gene expression and protein synthesis [21–25]. In culture, growth factors, such as TGF-β1 and insulin, and ECM proteins, such as laminins, support the expression of a contractile ASM phenotype, whereas mitogens such as platelet-derived growth factor (PDGF) or fetal serum and extracellular fibronectin matrices support proliferative function [16,26–28]. Maturation requires endogenously expressed laminin-2 and is reliant on cells expressing a unique repertoire of laminin-binding α-integrin subu-

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Airway smooth muscle cells

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nits [15,16]. Notably, in primary culture only a subset of airway myocytes mature appear able to reacquire a functionally contractile phenotype, suggesting—and supported by in situ evidence [29]—the existence of intrinsic heterogeneity between myocytes and perhaps suggesting that divergent mesenchymal sublineages may seed the ASM bed during development [9,11]. Notably, the maturation of contractile myocytes parallels the process of cellular hypertrophy, involving an increase in cell size and reliance on PI3K-associated signaling pathways [21,25,30]. Increased smooth muscle mass is a feature of airways remodeling and thus evaluation of myocyte maturation in culture contributes to the understanding of the pathogenesis of obstructive airway disease. Functional diversity Although there are molecular and ultrastructural markers for contractile phenotype myocytes, there also exists a broad diversity in the sensitivity and reactivity of smooth muscle factors that trigger contraction and relaxation. Diversity in functional responses to proliferative and prosynthetic mediators is also seen in cells expressing an immature phenotype [31,32]. Thus, although myocytes may be classified as “contractile” or “immature”, there are mechanisms that fine-tune cell function. Moreover, airway myocytes in a contractile phenotype retain a variable capacity to synthesize and express cytokines, chemokines, and ECM proteins [15,16,33]. This indicates that strict designation of airway myocytes as being contractile or synthetic/proliferative is not appropriate, as cells of any phenotype retain diversity in functional responses (Figure 12.2). One element of functional diversity that is dynamic over relatively short durations is termed mechanical plasticity, which stems from the remodeling of actomyosin filaments in response to changes in muscle length during stretch or contraction [34,35]. A functional capacity of this nature meets the physiologic burden of constantly variable forces, for example in breathing. Mechanical adaptation confers the capacity to retain maximum force generation but can alter shortening velocity, which may contribute to airway hyperresponsiveness [36,37]. An additional mechanism for the functional diversity of mature myocytes is a variation in the expression of proteins that mediate responses to extra-

cellular cues. For example, arterial resistance vessels contract after mechanical stretch owing to the activation of inward cation channels, whereas healthy bronchial smooth muscle exhibits little or no myogenic response [38,39]. Indeed, the repertoire of proteins expressed as receptors, receptor regulators, signaling effectors, and ion channels contributes significantly to the functional fine-tuning of smooth muscle cells across the range of phenotypic states [12,13, 40–42].

Functional plasticity airway smooth muscle: role in asthma pathogenesis Phenotypic plasticity of airway myocytes confers a multifunctional capacity—contraction, proliferation, hypertrophy, migration, and synthesis of inflammation and fibrosis-modulating biomolecules (Figure 12.3) [20]. Thus, myocytes contribute both to bronchial spasm and to structural changes associated with fixed airway obstruction. That ASM cells can synthesize inflammatory biomolecules and are a rich source of cytokines, chemokines, and growth factors is well documented [43,44]. Cultured myocytes express a number of cytokines (Th1 type: interleukin 2 [IL-2], interferon γ [IFN-γ], IL-12; Th2 type: IL-5, IL-6, granulocyte–macrophage colony-stimulating factor [GM-CSF]) and high levels of RANTES (regulated on activation, normal T cellexpressed, and secreted), eotaxin, IL-8, and IL-11 [45]. This indicates that ASM cells can determine local inflammation and establish a self-regulating mechanism for phenotype plasticity as a determinant of asthma pathogenesis and symptoms. Cultured ASM cells from asthmatics produce an altered composition of ECM [46,47]. The ECM affects all aspects of the functional repertoire of smooth muscle cells. For instance, seeding ASM onto fibronectin or collagen type I promotes a proliferative phenotype, whereas laminin-rich matrices are needed for maturation of a contractile phenotype [16,26,48]. Laminin-2 and its selective receptor, α7β1 integrin, are required for airway myocyte maturation and hypertrophy [15]. Moreover, laminin-2 is increased in the asthmatic airway [16]. Focus on the role of ECM receptor expression by ASM is warranted as it could yield viable therapeutic targets to combat, and even reverse, airway remodeling. 165


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Phenotype plasticity

Functional diversity

• Proliferation • Migration

• Contractility • Responsiveness to spasmogens & dilators

Maturation Modulation

• ECM synthesis • Cytokine synthesis

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Mechanical Plasticity

• ECM synthesis • Cytokine synthesis

Figure 12.2 Schematic representation of phenotype plasticity and functional diversity of smooth muscle. Phenotype plasticity results from reversible modulation and maturation of myocytes. Mechanical plasticity, a form of functional diversity, occurs in contractile phenotype myocytes as the result of subcellular reorganization of the contractile filaments in response to changes in muscle length during stretch or contraction. Functional diversity

exists cells of any phenotype owing to differences in expression of (i) receptor subtypes for growth factors, hormones, neurotransmitters, and extracellular matrix (ECM), (ii) regulators of receptors and receptor-induced signaling proteins (e.g., caveolins and GRKs), (iii) receptorinduced signaling proteins, and (iv) ion channels that determine membrane electrophysiology.

Increased ASM mass in human asthmatics results from cellular hyperplasia and hypertrophy [49,50]. ASM cells cultured from endobronchial biopsies from asthmatics proliferate at a faster rate than those from healthy individuals, and this difference appears to be directly linked to changes in the profile of ECM proteins synthesized by “asthmatic” myocytes [31,47,51]. This suggests the existence of an intrinsic, stable abnormality in the phenotype of myocytes in asthmatics that is associated with altered functional responses to asthma-associated mitogens. Prohypertrophic factors that are increased in asthmatic airways include TGF-β1 and endothelin-1, which induce ASM cell hypertrophy in culture [30,52]. Increased ASM mass, even in mild-to-moderate asthma, is accompanied by

increased proteins associated with contraction, such as myosin light chain kinase, which underpins the increased contractility of ASM tissue in asthma and in asthma models [50,53,54]. Additional mechanisms that could account for increased ASM mass in asthma also stem from the multifunctional capacity of these cells. Migration of fibroblasts from the submucosa to the muscle layer with subsequent maturation to a contractile phenotype could promote the accumulation of ASM tissue. A mechanism in which circulating mesenchymal stem cells home and migrate into damaged airways and then undergo differentiation to smooth muscle has also been postulated [55]. Moreover, to account for increases in airway myofibroblasts, ASM cells could

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Inflammation and fibraisis Epi MF

Migration ASM

Hypertrophy

Proliferation

Figure 12.3 Phenotypic and functional plasticity of mesenchymal cells in airway remodeling. Airway smooth muscle (ASM) cells (gray) and myofibroblasts (MF) (white) regulate the local tissue microenvironment by releasing inflammatory mediators and increasing expression of cell adhesion molecules and extracellular matrix (ECM) components. Increased ASM mass results from myocyte proliferation and hypertrophy. Myofibroblasts migrating toward the ASM layer may also contribute to muscle thickening—upon reaching the ASM compartment, mesenchymal cells undergo maturation. The process is likely modulated by local cytokines, chemokines, and ECM. Migration of phenotypically modulated ASM to the submucosal compartment leads to an increased number of myofibroblasts.

undergo phenotype modulation and migrate toward the epithelium [56].

Emerging therapies and their impact on airway smooth muscle Phosphodiesterase inhibitors Cyclic nucleotide phosphodiesterases (PDE) degrade the phosphodiester bond in cAMP and cGMP, imparting important signal transduction control [57]. Theophylline, a nonspecific PDE inhibitor, has been used to treat airway diseases for over 50 years and has bronchodilating, anti-inflammatory, and antifibrotic effects [58,59]. Subtype-specific PDE inhibitors that minimize unwanted side-effects now exist. PDE-4 is the predominant PDE subtype expressed in the airways [60]. First- and second-generation PDE-4 inhibitors like rolipram, piclamilast, ciclamilast,

cilomilast (Ariflo®), and roflumilast (Daxas®) show promise for reducing features of airway remodeling [61,62]; largely by raising cAMP in inflammatory, immune, and ASM cells. PDE-4 inhibitors appear to have effects on the breadth of ASM cell function. Roflumilast directly inhibits TGF-β1-induced ECM protein synthesis by asthmatic and nonasthmatic cultured ASM cells and in tracheal rings in vitro [63]. It also reduces subepithelial fibrosis and tracheal epithelial activation in mice that are chronically challenged with allergen [64]. Cilomilast reduces both basal- and PDGF-stimulated migration of human ASM cells in vitro [65]. Both rolipram and NIS-62949 reduce airway hyperresponsiveness (AHR) and bronchoconstriction in animal models of allergic asthma AHR [66,67]. Thus, there are compelling data emerging to suggest that PDE-4 inhibitors prevent AHR, airway remodeling, and inflammation via pathways involving ASM.

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Rho kinase inhibitors Rho kinases are effectors of RhoA, a monomeric GTP-binding protein, that cause Ca2+ sensitization of ASM by inactivating myosin light chain phosphatase, the enzyme chiefly responsible for tempering contraction and causing relaxation. Two isoforms of Rho kinase exist (ROCK-1 and ROCK-2 [68]), there are no isoform selective inhibitors, and both isoforms are expressed in lung and inflammatory cells. The major roles of Rho kinase in cell physiology include contraction, cell attachment, migration, proliferation, and cell survival. A widely studied Rho kinase inhibitor, Y-27632, is a potent cell-permeable competitive inhibitor with bronchodilatory effects when delivered to the airways in allergen exposed animals [69,70]. In animal models of asthma, Rho kinase inhibitors suppress acute AHR and airway remodeling arising from allergic inflammation [70,71]. Analogs of Y-27632 also promote smooth muscle relaxation [72,73]. Rho kinase is also involved with “activation” of airway fibroblasts to myofibroblasts that contribute collagen deposition [74]. Based on the links between Rho A and Rho kinase activity with bronchoconstriction, inflammatory cell recruitment, AHR, smooth muscle growth, and myofibroblast activation, Rho kinase inhibitors may be useful for treatment, and abundant research in this area is expected. 167


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Statins Statins inhibit mevalonate synthesis, the proximal step in cholesterol biosynthesis. They reduce serum cholesterol but emerging evidence indicates that they have an even greater health benefit, thus spawning a growing number of clinical trials of their impact on lung health. The positive effects of statins in asthma control are equivocal [75–77]; however, studies to date are limited because they only included mild asthmatics, statin use was of relatively short duration, and in some cases steroid use stopped. Reports indicate a direct impact for statins on ASM cell growth, fibrotic function, and cell survival [78–80]. Statins impact cell responses via the indirect inhibition of small GTPase proteins (e.g., Ras, Rac, Rho), mitogenactivated protein (MAP) kinases, and nuclear factor κB; this suppresses airway inflammation in allergenchallenged mice and improves lung physiology [81,82]. Although the investigation into the use of statins to treat asthma is in its infancy, studies to date suggest that these compounds could open doors for new therapeutic approaches as mechanisms for their pleiotropic effects are unraveled. Bronchial thermoplasty Bronchial thermoplasty uses radiofrequency energy to reduce ASM mass and attenuate bronchoconstriction [83]. Initial studies in canines show that the application of radiofrequency to the airways, at temperatures of 65°C and 75°C, reduced airway responsiveness to methacholine by ∼50% [84]. These effects were maintained for up to 3 years, and histologic analysis revealed complete ablation of ASM with emergence of a thin layer of collagen [84]. The procedure is in clinical trial; to date, patients report decreased numbers of mild and severe exacerbations and increased symptom-free days [85]. The changes to the airway wall and the effectiveness of treatment on long-term asthma control are currently under investigation, and more insight is expected in the foreseeable future.

Concluding remarks The broad functional behavior of ASM may be at the root of the biologic mechanisms that lead to changes in airway function and structure in obstruc-

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tive airways disease. Increased bronchial spasm with altered airway structure, which manifests as AHR, are likely linked to the plastic multifunctional behavior of airway myocytes. For instance, increased myocyte contraction due to elevated myosin light chain kinase leads to greater airway narrowing. Similarly, changes in pharmacologic responsiveness of myocytes occur in the face of asthma-associated mediators and thus underpin increased sensitivity to inhaled allergic and nonallergic contractile agonists. Notably, airway myocytes are also rich sources of inflammatory mediators, making them determinants of the local inflammatory environment, and the accumulation of airway ECM is largely the result of altered function of ASM and (myo)fibroblasts. Thus, the role of mesenchymal cells in the pathogenesis of obstructive airways disease is at the forefront of current research in airway biology. Advances expected in the next decade will likely yield significant insight that may dictate the direction for the development of new and more effective pharmacologic interventions.

Acknowledgment PS holds a Frederick Banting and Charles Best Canadian Graduate Scholarship from Canadian Institute of Health Research (CIHR) and National Training Program in Allergy and Asthma. AJH is supported by the Canada Research Chair program, Canada Foundation for Innovation, and the Manitoba Institute of Child Health.

References 1 Wissler RW. The arterial medial cell, smooth muscle, or multifunctional mesenchyme? Circulation 1967; 36: 1–4. 2 Chamley-Campbell J, Campbell GR, Ross R. The smooth muscle cell in culture. Physiol Rev 1979; 59: 1–61. 3 Avner BP, Delongo J, Wilson S, Ladman AJ. A method for culturing canine tracheal smooth muscle cells in vitro: morphologic and pharmacologic observations. Anat Rec 1981; 200: 357–70. 4 Tom-Moy M, Madison JM, Jones CA, de Lanerolle P, Brown JK. Morphologic characterization of cultured smooth muscle cells isolated from the tracheas of adult dogs. Anat Rec 1987; 218: 313–28.

168


3/16/2011 8:27:16 PM


5 Twort Ch, van Breemen C. Human airway smooth muscle in cell culture: control of the intracellular calcium store. Pulm Pharmacol 1989; 2: 45–53. 6 Panettieri RA, Murray RK, DePalo LR, Yadvish PA, Kotlikoff MI. A human airway smooth muscle cell line that retains physiological responsiveness. Am J Physiol 1989; 256: C329–35. 7 Hall IP, Kotlikoff M. Use of cultured airway myocytes for study of airway smooth muscle. Am J Physiol 1995; 268: L1–11. 8 Halayko AJ, Salari H, Ma X, Stephens NL. Markers of airway smooth muscle cell phenotype. Am J Physiol 1996; 270: L1040–51. 9 Halayko AJ, Camoretti-Mercado B, Forsythe SM, et al. Divergent differentiation paths in airway smooth muscle culture: induction of functionally contractile myocytes. Am J Physiol Lung Cell Mol Physiol 1999; 276: L197–206. 10 Halayko AJ, Solway J. Molecular mechanisms of phenotypic plasticity in smooth muscle cells. J Appl Physiol 2001; 90: 358–68. 11 Mitchell RW, Halayko AJ, Kahraman S, Solway J, Wylam ME. Selective restoration of calcium coupling to muscarinic M(3) receptors in contractile cultured airway myocytes. Am J Physiol Lung Cell Mol Physiol 2000; 278: L1091–100. 12 Mitchell RW, Halayko AJ, Kahraman S, Solway J, Wylam ME. Selective restoration of calcium coupling to muscarinic M(3) receptors in contractile cultured airway myocytes. Am J Physiol Lung Cell Mol Physiol 2000; 278: L1091–100. 13 Gosens R, Stelmack GL, Dueck G, et al. Caveolae facilitate muscarinic receptor-mediated intracellular Ca2+ mobilization and contraction in airway smooth muscle. Am J Physiol Lung Cell Mol Physiol 2007; 293: L1406–18. 14 Halayko AJ, Tran T, Gosens R. Phenotype and functional plasticity of airway smooth muscle: role of caveolae and caveolins. Proc Am Thorac Soc 2008; 5: 80–8. 15 Tran T, Ens-Blackie K, Rector ES, et al. Laminin-binding Integrin {alpha}7 is required for contractile phenotype expression by human airway myocyte. Am J Respir Cell Mol Biol 2007; 37: 668–80. 16 Tran T, McNeill KD, Gerthoffer WT, Unruh H, Halayko AJ. Endogenous laminin is required for human airway smooth muscle cell maturation. Respir Res 2006; 7: 117. 17 Solway J, Seltzer J, Samaha FF, et al., Structure and expression of a smooth muscle cell-specific gene, SM22 alpha. J Biol Chem 1995; 270: 13460–9. 18 Gosens R, Stelmack GL, Dueck G, et al. Role of caveolin-1 in p42/p44 MAP kinase activation and proliferation of human airway smooth muscle. Am J Physiol Lung Cell Mol Physiol 2006; 291: L523–34.

19 Sharma P, Tran T, Stelmack GL, et al., Expression of the dystrophin–glycoprotein complex is a marker for human airway smooth muscle phenotype maturation. Am J Physiol Lung Cell Mol Physiol 2008; 294: L57–68. 20 Halayko AJ, Tran T, Ji SY, Yamasaki A, Gosens R. Airway smooth muscle phenotype and function: interactions with current asthma therapies. Curr Drug Targets 2006; 7: 525–40. 21 Halayko AJ, Kartha S, Stelmack GL, et al. Phophatidylinositol-3 kinase/mammalian target of rapamycin/p70S6K regulates contractile protein accumulation in airway myocyte differentiation. Am J Respir Cell Mol Biol 2004; 31: 266–75. 22 Camoretti-Mercado B, Fernandes DJ, et al., Inhibition of transforming growth factor beta-enhanced serum response factor-dependent transcription by SMAD7. J Biol Chem 2006; 281: 20383–92. 23 Camoretti-Mercado B, Liu HW, Halayko AJ, et al. Physiological control of smooth muscle-specific gene expression through regulated nuclear translocation of serum response factor. J Biol Chem 2000; 275: 30387–93. 24 Liu HW, Halayko AJ, Fernandes DJ, et al., The RhoA/ Rho kinase pathway regulates nuclear localization of serum response factor. Am J Respir Cell Mol Biol 2003; 29: 39–47. 25 Zhou L, Goldsmith AM, Bentley JK, et al. 4E-binding protein phosphorylation and eukaryotic initiation factor4E release are required for airway smooth muscle hypertrophy. Am J Respir Cell Mol Biol 2005; 33: 195–202. 26 Hirst SJ, Twort Ch, Lee Th. Differential effects of extracellular matrix proteins on human airway smooth muscle cell proliferation and phenotype. Am J Respir Cell Mol Biol 2000; 23: 335–44. 27 Schaafsma D, McNeill KD, Stelmack GL, et al. Insulin increases the expression of contractile phenotypic markers in airway smooth muscle. Am J Physiol Cell Physiol 2007; 293: C429–39. 28 Dekkers BG, Schaafsma D, Nelemans SA, Zaagsma J, Meurs H. Extracellular matrix proteins differentially regulate airway smooth muscle phenotype and function. Am J Physiol Lung Cell Mol Physiol 2007; 292: L1405–13. 29 Halayko AJ, Stelmack GL, Yamasaki A, McNeill K, Unruh H, Rector E. Distribution of phenotypically disparate myocyte subpopulations in airway smooth muscle. Can J Physiol Pharmacol 2005; 83: 104–16. 30 Goldsmith AM, Bentley JK, Zhou L, et al. Transforming growth factor-beta induces airway smooth muscle hypertrophy. Am J Respir Cell Mol Biol 2006; 34: 247–54. 31 Johnson PR, Roth M, Tamm M, et al. Airway smooth muscle cell proliferation is increased in asthma. Am J Respir Crit Care Med 2001; 164: 474–7. 32 Burgess JK, Ge Q, Boustany S, Black JL, Johnson PR. Increased sensitivity of asthmatic airway smooth

169


3/16/2011 8:27:17 PM

Ay

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muscle cells to prostaglandin E2 might be mediated by increased numbers of E-prostanoid receptors. J Allergy Clin Immunol 2004; 113: 876–81. 33 Redhu NS, Saleh A, Shan L, et al. Proinflammatory and Th2 cytokines regulate the high affinity IgE receptor (FcepsilonRI) and IgE-dependent activation of human airway smooth muscle cells. PLoS One 2009; 4: e6153. 34 Gunst SJ, Tang DD, Opazo Saez A. Cytoskeletal remodeling of the airway smooth muscle cell: a mechanism for adaptation to mechanical forces in the lung. Respir Physiol Neurobiol 2003; 137: 151–68. 35 Ford LE, Seow CY, Pratusevich VR. Plasticity in smooth muscle, a hypothesis. Can J Physiol Pharmacol 1994; 72: 1320–4. 36 Wang L, Pare PD, Seow CY. Changes in force-velocity properties of trachealis due to oscillatory strains. J Appl Physiol 2002; 92: 1865–72. 37 King GG, Pare PD, Seow CY. The mechanics of exaggerated airway narrowing in asthma: the role of smooth muscle. Respir Physiol 1999; 118: 1–13. 38 McGregor E, Gosling M, Beattie DK, Ribbons DM, Davies AH, Powell JT. Circumferential stretching of saphenous vein smooth muscle enhances vasoconstrictor responses by Rho kinase-dependent pathways. Cardiovasc Res 2002; 53: 219–26. 39 Wu X, Davis MJ. Characterization of stretch-activated cation current in coronary smooth muscle cells. Am J Physiol Heart Circ Physiol 2001; 280: H1751–61. 40 Janssen LJ. Ionic mechanisms and Ca(2+) regulation in airway smooth muscle contraction: do the data contradict dogma? Am J Physiol Lung Cell Mol Physiol 2002; 282: L1161–78. 41 Borchers MT, Biechele T, Justice JP, et al. Methacholineinduced airway hyperresponsiveness is dependent on Galphaq signaling. Am J Physiol Lung Cell Mol Physiol 2003; 285: L114–20. 42 Walker JK, Gainetdinov RR, Feldman DS, et al. G protein-coupled receptor kinase 5 regulates airway responses induced by muscarinic receptor activation. Am J Physiol Lung Cell Mol Physiol 2004; 286: L312–19. 43 Howarth PH, Knox AJ, Amrani Y, Tliba O, Panettieri RA, Jr., Johnson M. Synthetic responses in airway smooth muscle. J Allergy Clin Immunol 2004; 114: S32–50. 44 Halayko AJ, Solway J. Molecular mechanisms of phenotypic plasticity in smooth muscle cells. J Appl Physiol 2001; 90: 358–68. 45 Halayko AJ, Amrani Y. Mechanisms of inflammationmediated airway smooth muscle plasticity and airways remodeling in asthma. Respir Physiol Neurobiol 2003; 137: 209–22. 46 Johnson PR, Black JL, Carlin S, Ge Q, Underwood PA. The production of extracellular matrix proteins by human passively sensitized airway smooth-muscle cells

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in culture: the effect of beclomethasone. Am J Respir Crit Care Med 2000; 162: 2145–51. 47 Johnson PR. Role of human airway smooth muscle in altered extracellular matrix production in asthma. Clin Exp Pharmacol Physiol 2001; 28: 233–6. 48 Tran T, Gosens R, Halayko AJ. Effects of extracellular matrix and integrin interactions in airway smooth muscle phenotype and function: it takes two to tango! Curr Respir Med Rev 2007; In Press. 49 Ebina M, Takahashi T, Chiba T, Motomiya M. Cellular hypertrophy and hyperplasia of airway smooth muscles underlying bronchial asthma. A 3-D morphometric study. Am Rev Respir Dis 1993; 148: 720–6. 50 Benayoun L, Druilhe A, Dombret MC, Aubier M, Pretolani M. Airway structural alterations selectively associated with severe asthma. Am J Respir Crit Care Med 2003; 167: 1360–8. 51 Johnson PR, Burgess JK, Underwood PA, et al. Extracellular matrix proteins modulate asthmatic airway smooth muscle cell proliferation via an autocrine mechanism. J Allergy Clin Immunol 2004; 113: 690–6. 52 McWhinnie R, Pechkovsky DV, Zhou D, et al. Endothelin-1 induces hypertrophy and inhibits apoptosis in human airway smooth muscle cells. Am J Physiol Lung Cell Mol Physiol 2007; 292: L278–86. 53 Jiang H, Rao K, Halayko AJ, Liu X, Stephens NL. Ragweed sensitization-induced increase of myosin light chain kinase content in canine airway smooth muscle. Am J Respir Cell Mol Biol 1992; 7: 567–73. 54 Ammit AJ, Armour CL, Black JL. Smooth-muscle myosin light-chain kinase content is increased in human sensitized airways. Am J Respir Crit Care Med 2000; 161: 257–63. 55 Stewart AG. Emigration and immigration of mesenchymal cells: a multicultural airway wall. Eur Respir J 2004; 24: 515–17. 56 Kelly MM, O’Connor TM, Leigh R, et al. Effects of budesonide and formoterol on allergen-induced airway responses, inflammation, and airway remodeling in asthma. J Allergy Clin Immunol 2010; 125: 349–56, e13. 57 Weiss B, Hait WN. Selective cyclic nucleotide phosphodiesterase inhibitors as potential therapeutic agents. Annu Rev Pharmacol Toxicol 1977; 17: 441–77. 58 Finney MJ, Karlsson JA, Persson CG. Effects of bronchoconstrictors and bronchodilators on a novel human small airway preparation. Br J Pharmacol 1985; 85: 29–36. 59 Yano Y, Yoshida M, Hoshino S, et al. Anti-fibrotic effects of theophylline on lung fibroblasts. Biochem Biophys Res Commun 2006; 341: 684–90. 60 Kroegel C, Foerster M. Phosphodiesterase-4 inhibitors as a novel approach for the treatment of respiratory disease: cilomilast. Expert Opin Investig Drugs 2007; 16: 109–24.

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170


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61 Bundschuh DS, Eltze M, Barsig J, Wollin L, Hatzelmann A, Beume R. In vivo efficacy in airway disease models of roflumilast, a novel orally active PDE4 inhibitor. J Pharmacol Exp Ther 2001; 297: 280–90. 62 Mata M, Sarria B, Buenestado A, Cortijo J, Cerda M, Morcillo EJ. Phosphodiesterase 4 inhibition decreases MUC5AC, expression induced by epidermal growth factor in human airway epithelial cells. Thorax 2005; 60: 144–52. 63 Burgess JK, Oliver BG, Poniris MH, et al. A phosphodiesterase 4 inhibitor inhibits matrix protein deposition in airways in vitro. J Allergy Clin Immunol 2006; 118: 649–57. 64 Kumar RK, Herbert C, Thomas PS, et al. Inhibition of inflammation and remodeling by roflumilast and dexamethasone in murine chronic asthma. J Pharmacol Exp Ther 2003; 307: 349–55. 65 Goncharova EA, Billington CK, Irani C, et al. Cyclic AMP-mobilizing agents and glucocorticoids modulate human smooth muscle cell migration. Am J Respir Cell Mol Biol 2003; 29: 19–27. 66 Singh SP, Mishra NC, Rir-Sima-Ah J, et al. Maternal exposure to secondhand cigarette smoke primes the lung for induction of phosphodiesterase-4D5 isozyme and exacerbated Th2 responses: rolipram attenuates the airway hyperreactivity and muscarinic receptor expression but not lung inflammation and atopy. J Immunol 2009; 183: 2115–21. 67 Dastidar SG, Ray A, Shirumalla R, et al., Pharmacology of a novel, orally active PDE4 inhibitor. Pharmacology 2009; 83: 275–86. 68 Noma K, Oyama N, Liao JK. Physiological role of ROCKs in the cardiovascular system. Am J Physiol Cell Physiol 2006; 290: C661–8. 69 Iizuka K, Shimizu Y, Tsukagoshi H, et al. Evaluation of Y-27632: a rho-kinase inhibitor, as a bronchodilator in guinea pigs. Eur J Pharmacol 2000; 406: 273–9. 70 Henry PJ, Mann TS, Goldie RG. A rho kinase inhibitor, Y-27632; inhibits pulmonary eosinophilia, bronchoconstriction and airways hyperresponsiveness in allergic mice. Pulm Pharmacol Ther 2005; 18: 67–74. 71 Schaafsma D, Bos IS, Zuidhof AB, Zaagsma J, Meurs H. Inhalation of the Rho-kinase inhibitor Y-27632; reverses allergen-induced airway hyperresponsiveness after the early and late asthmatic reaction. Respir Res 2006; 7: 121. 72 Uehata M, Ishizaki T, Satoh H, et al. Calcium sensitization of smooth muscle mediated by a Rho-associated protein kinase in hypertension. Nature 1997; 389: 990–4.

73 Ishizaki T, Uehata M, Tamechika I, et al., Pharmacological properties of Y-27632: a specific inhibitor of rhoassociated kinases. Mol Pharmacol 2000; 57: 976–83. 74 Urata Y, Nishimura Y, Hirase T, Yokoyama M. Sphingosine 1-phosphate induces alpha-smooth muscle actin expression in lung fibroblasts via Rho-kinase. Kobe J Med Sci 2005; 51: 17–27. 75 Menzies D, Nair A, Meldrum KT, Fleming D, Barnes M, Lipworth BJ. Simvastatin does not exhibit therapeutic anti-inflammatory effects in asthma. J Allergy Clin Immunol 2007; 119: 328–35. 76 Hothersall EJ, Chaudhuri R, McSharry C, et al. Effects of atorvastatin added to inhaled corticosteroids on lung function and sputum cell counts in atopic asthma. Thorax 2008; 63: 1070–5. 77 Ostroukhova M, Kouides RW, Friedman E. The effect of statin therapy on allergic patients with asthma. Ann Allergy Asthma Immunol 2009; 103: 463–8. 78 Ghavami S, Mutawe MM, Hauff K, et al. Statintriggered cell death in primary human lung mesenchymal cells involves p53-PUMA and release of Smac and Omi but not cytochrome c. Biochim Biophys Acta 2010; 1803: 452–67. 79 Schaafsma D, Dueck G, Ghavami S, et al. The mevalonate cascade as a target to suppress extracellular matrix synthesis by human airway smooth muscle. Am J Respir Cell Mol Biol 2010. 80 Takeda N, Kondo M, Ito S, Ito Y, Shimokata K, Kume H. Role of RhoA inactivation in reduced cell proliferation of human airway smooth muscle by simvastatin. Am J Respir Cell Mol Biol 2006; 35: 722–9. 81 Zeki AA, Franzi L, Last J, Kenyon NJ. Simvastatin inhibits airway hyperreactivity: implications for the mevalonate pathway and beyond. Am J Respir Crit Care Med 2009; 180: 731–40. 82 Kim DY, Joo JK, Ryu SY, Park YK, Kim YJ, Kim SK. Clinicopathological characteristics and prognosis of carcinoma of the gastric cardia. Dig Surg 2006; 23: 313–18. 83 Cox G, Miller JD, McWilliams A, Fitzgerald JM, Lam S. Bronchial thermoplasty for asthma. Am J Respir Crit Care Med 2006; 173: 965–9. 84 Danek CJ, Lombard CM, Dungworth DL, et al. Reduction in airway hyperresponsiveness to methacholine by the application of RF energy in dogs. J Appl Physiol 2004; 97: 1946–53. 85 Cox G, Thomson NC, Rubin AS, et al. Asthma control during the year after bronchial thermoplasty. N Engl J Med 2007; 356: 1327–37.

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AUTHOR QUERY FORM Dear Author, During the preparation of your manuscript for publication, the questions listed below have arisen. Please attend to these matters and return this form with your proof. Many thanks for your assistance. Query References

Query

Remarks

1

AU: Edit to sentence beginning ‘Airway smooth muscle is a primary…’ OK?

2

AU: Sentence beginning ‘Work over [the past] 40 years…’ OK to insert ‘…the past…’?

3

AU: Sentence beginning ‘Notably, in primary culture only a subset of airway…’ Is ‘…airway myocytes mature appear…’ correct? Can ‘mature’ be deleted? Or should it be ‘…mature airway myocytes…’?

4

AU: Please provide trade names and manufacturer names and addresses for rolipram, piclamilast and ciclamilast and provide manufacterers for ciclomilast and roflumilast.

5

AU: Please provide updated details for reference 48 (Tran).

6

AU: Please provide volume number and page range for reference 79 (Schaafsma) if available.

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