Pathophysiology and Airway Remodeling in Asthma NTPAA course:

1. Pathophysiology and Airway Remodeling in Asthma NTPAA course: Advanced Topics in Immunology (IMMU 7100) Research Topics in Physiology (PHGY 7190) Andrew J Halayko, PhD Office: RS321 Respiratory Hospital (810 Sherbrook), 787-2062 Lab: 547 John Buhler Research Centre, 789-3778 (November 2008)

2. Asthma – Definition • “Asthma is a disorder of the airways characterized by paroxysmal or persistent symptoms (dyspnea, chest tightness, wheeze and cough), with variable airflow limitation [and] airway hyperresponsiveness to a variety of stimuli” • The definition includes these concepts: • Asthma is a chronic inflammatory disease (thus without appropriate treatment is progressive) • There is airflow limitation that is reversible (though there is also an irreversible component) • A variety of stimuli can trigger airway’s response. • (National Asthma Task Force “Prevention and management of asthma in Canada”, 2000)

3. Airway Remodeling – the Progression of Asthma

4. Airway Hyperresponsiveness: Asthma as a Progressive Disease • Long-standing asthma  Recently diagnosed asthma Boulet et al. Am J Respir Crit Care Med 162: 1308-13, 2000

5. Branching of airways in the human tracheo-bronchial tree Conducting Airways: ciliated epithelia, mucous, smooth muscle Respiratory Zones: (air velocity ~ 0) phagocytic cells (alveolar macrophages), lymphatic channels drain into hilar lymph nodes

6. Anatomy of the Airways

7. The Anatomy of Airflow Limitation and Airway Hyperresponsiveness Baseline Antigen Atropine Tatalum bronchogram, allergen-sensitized canine, right lower lobe Kessler et al, 1975, JAP 38:96

8. Methacholine Cockroft, 1985 Airway Hyperresponsiveness: Provocative Challenge Test Common Stimuli:chemical (eg. histamine, Mch, -agonists, LT, adenosine); physical (eg. exercise, cold air); sensitizers (eg. allergen)

9. Methacholine (mg/mL) Airway Hyperresponsiveness: Provocative Challenge Test Common Stimuli:chemical (eg. histamine, Mch, -agonists, LT, adenosine); physical (eg. exercise, cold air); sensitizers (eg. allergen PC20 (mg/mL) Cockroft, 1985 & 1997

10. Measurement of Airway Function: Response to Bronchodilators Common Stimuli: chemical (eg. histamine, Mch, -agonists, LT, adenosine); physical (eg. exercise, cold air); sensitizers (eg. allergen

11. The Deep Breath Phenomenon: Structure and Function in the Lung During Mch challenge, if NORMAL subjects don’t take a deep inspiration, they respond to Mch the same way as asthmatics! “intrinsic impairment of the ability of inspiration to stretch airway smooth muscle is a major feature of asthma”(Skloot, Permutt, and Togias JCI, 96:2393, 1995) Why isn’t deep breath broncho-protective in asthma?

12. Lung Structure / Function Relationships Why isn’t deep breath broncho-protective in asthma? • Normally, there is interdependence of airway size and and lung volume – a primary determinant of airway calibre is lung volume (Mechanical coupling of the airways and lung parenchyma, thus the size and shape of one affects the other). Airways are hyperresponsive AND there are altered structure / function relationships of the airways in asthma ... the biological mechanism? Inflammation and remodelling!

13. Airways Remodelling: Important Concepts • Changes in airway wall thickness and organization, especially evident in severe and fatal asthmatics. • Remodelling in mild asthmatics & in children • Duration of disease determines effectiveness of therapy - remodelling in asthma is progressive. • No clear evidence for reversal of structural changes with asthma therapies. • The cause-effect relationship between the components of airways remodelling and altered lung function still not fully understood.

14. Moderate Asthmatic Asthma Pathology : Airway Inflammation and Remodeling Non-asthmatic Airway remodeling affects: Epithelium; submucosa (fibrosis & myofibroblasts); smooth muscle; adventitia (fibrosis); bronchial vasculature; mucus gland & goblet cell dysplasia)

15. Fatal asthma - airways blocked by mucus secretions admixed with inflammatory exudates Jeffery AJRCCM., 164:S28-38, 2001 Features of Airway Remodeling: Goblet cell and mucus gland hyperplasia Asthmatic Normal subject Ordonez et al AJRCCM 163, 517-23, 2002

16. Airways Remodelling in Asthma: Fibrosis • Thickening of the basement membrane and increase matrix deposition throughout the airway wall: • Coll I & III • fibronectin • laminin 2 • tenascin • elastin • versicam • hyaluronan Laminin Tenascin Jeffery AJRCCM., 164:S28-38, 2001 Laitinen et al. Asthma, pp. 209-23, 1997

17. Airways Remodeling: Submucosal (myo)fibroblasts Laitinen et al. Asthma, pp. 209-23, 1997

18. Atopic asthmatic – diluent Atopic asthmatic – allergen Myofibroblast Accumulation Occurs 24hrs After Allergen Challenge Atopic non-asthmatic (no challenge) Non-atopic, non-asthmatic (no challenge) 24 hrs !! Cell # per 0.1mm2 Myofibro MC Eos Gizycki et al. Am J Respir Cell Mol Biol. 1997

19. Subepithelial Myofibroblasts from Asthmatics Express a Pro-fibrotic Phenotype IL-4 effects on collagen, MMP-2 & TIMP-2 expression BNF = bronchial fibroblast (non-asthmatic subjects) BAF = bronchial fibroblasts (mild/moderate asthmatic subjects) MMP-2 TIMP-2 Baseline + IL-4 (6hr) Pro-collagen I (1) mRNA BNF BAF BNF BAF BNF BAF Bergeron et al, Clin Exp Allergy  33: 1389-1397, 2003

20. Increased ECM Production by Asthmatic ASM Potentiates Proliferative Responses Production of ECM proteins Proliferation of ASM Cells ECM from Non-asthmatic Non-asthmatic ECM from Asthmatic Asthmatic % ASM Proliferation Johnson et al, J Allergy Clin Immunol 113: 690-696, 2004

21. Airways Remodelling: Airway Smooth Muscle • Inflammation & fibrosis • ASM Thickening • - cellular hyperplasia and hypertrophy • -Type I and Type II asthma • Hyperresponsiveness • - due to altered sensitivity to spasmogens & dilators, and from increased ASM Non-asthmatic (SEM of mucosa) Fatal asthma (SEM of mucosa) Jeffery AJRCCM., 164:S28-38, 2001

22. Increased ASM Mass In Mild-to-Moderate and Severe Asthma • Mild-to-Moderate Asthma: • Myocyte hyperplasia (>2x vs. control) • (Woodruff et al., Am J Resp Crit Care Med 169:1001-1006, 2004) • Myocyte hypertrophy (<2x vs. control) • (Benayoun et al., Am J Resp Crit Care Med 167:1360-1368, 2004) • Severe Asthma: • Type I (myocyte hyperplasia in proximal airways) • Type II (proximal myocyte hyperplasia & hypertrophy along distal airways) • (Ebina et al., Am Rev Resp Dis 148:720-726, 1993) • Myocyte hypertrophy (>3x vs. control) • (Benayoun et al., Am J Resp Crit Care Med 167:1360-1368, 2004)

23. Can Airway Remodelling and Associated Hyperresponsiveness be Reversed? HDM naive 5x/wk x 7 wks BALB/c HDM extract (25g intranasal) A,B - mucin production in epithelial goblet cells (Alcian blue (pH 2.5)) C,D - subepithelial collagen (Picro Sirius red) E,F – sm--actin Johnson et al 2004 Am J Respir Crit Care 169:378-385

24. Can Airway Remodelling and Associated Hyperresponsiveness be Reversed? 5x/wk x 7 wks 9 wks resolution BALB/c HDM extract (25g intranasal) % Area Collagen % Area sm--actin BALF Eos (x106/ml) Naive Naive Naive Resolution 3wk HDM 5wk HDM Resolution 3wk HDM 5wk HDM Resolution 3wk HDM 5wk HDM Johnson et al 2004 Am J Respir Crit Care 169:378-385

25. Can Airway Remodelling and Associated Hyperresponsiveness be Reversed? 5x/wk x 7 wks 9 wks resolution BALB/c HDM extract (25g intranasal)  naive mice  5 wk HDM  resolution Airway response to methacholine Johnson et al 2004 Am J Respir Crit Care 169:378-385

26. Airways Remodelling: Neovascularization Jeffery AJRCCM., 164:S28-38, 2001

27. Airways Remodelling: Neovascularization Control COPD High-magnification bronchovideoscope • measures vessel length density & vessel area density Asthma: Newly diagnosed, steroid-naive Asthma: 5 Yrs, + ICS Tanaka et al. 2004 Am J Resp Crit Care Med 168:1495-1499

28. # MC / mm2 airway e MC Norm. Control Asthma Eosinophilic bronchitis asm # MC / mm2 smooth muscle MC asm Norm. Control Asthma Eosinophilic bronchitis Airways Remodelling: Mast Cells Asthmatic subject, airway biopsy • atopic nonatopic Brightling et al. N Engl J Med 346 1699-1705, 2002

29. TAA, 400µg bid 42 µg bid * ICS = TAA (triamcinolone acetonide) Airway Mast Cells in Persistent Asthma Kraft et al., Chest, 124:42-50, 2003 The Asthma Clinical Research Network • N=45 with clinically stable, persistent asthma • 35 completed treatment phase; 10 randomized among groups had treatment failure • RX failure criteria (exacerbations): • - need >1 prednisone • - > 1 ER visit • - hospitalization for asthma • BAL tryptase shows the same trend as does MC # Median MC #/mm2)

30. Asthma Pathophysiology – Take Home Messages • Lung is the “end organ” – bronchi, bronchioles, alveloi • Multi-factorial - allergy; genes; environment; phenotype • Asthma is a progressive disease (if untreated – perhaps also if treated?) • Pulmonary function - variable airflow obstruction (PEF); airway hyperresponsiveness (FEV1) • Airway inflammation / airway remodelling - fibrosis; edema; inflammatory cells; hypertrophy/hyperplasia • epithelium; (myo)fibroblasts; airway smooth muscle; mucous glands; nerves; vasculature; eosiniphils, neutrophils, lymphocytes (TH1/TH2); mast cells