Thesis (doctoral)--Lund University, 1997. / Added t.p. with thesis statement inserted.

No description available.

Asthma is a chronic disease of airway hyper-responsiveness, airway inflammation and episodic bronchoconstriction. With asthma forecasted to increase by an additional 100 million cases by 2025, there is a critical and immediate need to address new asthma therapies. Guidelines for asthma treatment in the emergency department conditionally recommend intravenous magnesium sulfate (MgSO4). However, some investigations have failed to demonstrate beneficial effects. Ethical constraints limit evaluation of the bronchodilatory effects of MgSO4 alone in patients with acute asthma exacerbation, independent of other conventional therapeutics. To address this ethical dilemma, this study consisted of two phases: 1) quantification of the independent pulmonary effect of three doubling doses of MgSO4 in the spontaneous equine model of asthma during naturally occurring exacerbations of bronchoconstriction, and 2) evaluation of arterial blood gas parameters in response to administration of MgSO4 at a dose identified in phase 1 that yielded greatest efficacy without deleterious side effects.

Pulmonary surfactant is synthesized in the alveoli and lines the respiratory epithelium of the airways. Phospholipids, the main component of surfactant, confer it its ability to lower surface tension and to prevent alveolar collapse. Airway surfactant helps maintain smaller airway patency, improves muco-ciliary clearance, decreases bronchoconstriction, and modulates pulmonary immunity. Surfactant alterations in human asthma are therefore believed to contribute to the severity of airway obstruction. The goal of our first study was to characterize surfactant phospholipid composition and function in healthy horses, and to investigate the influence of age and bronchoalveolar lavage fluid (BALF) sample characteristics on surfactant. For that purpose, BALF was collected from 17 healthy horses and evaluated for BALF recovery percentage, cell count, and cell differential. BALF was separated into crude surfactant pellets (CSP) and supernatant and was analyzed for phospholipid content, protein content, phospholipid composition, and surface tension. Interestingly, phospholipid (surfactant) content in CSP significantly decreased with age. BALF recovery percentage, nucleated cell count, and cytological profile did not affect surfactant composition or function. The hypothesis of our second study was that surfactant alterations in RAO-affected horses are related to clinical stage of RAO. The objectives were 1) to compare surfactant phospholipid composition and function between Non-RAO and RAO horses at clinical stages and 2) to investigate relationships between surfactant alterations and variables assessing clinical stage of RAO. Seven horses with confirmed RAO and seven Non-RAO horses were evaluated in pairs (RAO/Non-RAO) at baseline, during exposure to hay, and post-exposure. Assessments included: clinical scoring, measure of maximal change in pleural pressure (ΔPplmax), airway endoscopy, and BALF cell counts and differentials. Samples were processed and analyzed as described above. Phospholipid levels in BALF were significantly lower in RAO versus Non-RAO horses, even in the absence of clinical signs. In the group of RAO horses, phospholipid content was significantly lower during exposure versus baseline. Furthermore, exposure to hay led to an increase in the protein versus phospholipid ratio in BALF from RAO horses. No significant differences were found in BALF protein content, phospholipid composition, or surface tension between or within groups of horses. Phosphatidylglycerol percentage had a tendency to be lower in RAO horses with higher clinical scores. Supernatant protein content was related to BALF neutrophilia in RAO crisis and overall ΔPplmax . In conclusion, our study demonstrated that surfactant alterations in RAO horses are present in remission and are exacerbated following exposure to hay. It is conceivable that a lower amount of surfactant in bronchioli of RAO horses may contribute to the horses' propensity to develop airway obstruction, mucous accumulation, and bronchial hyperresponsiveness. This may be exacerbated during crisis by a relatively higher protein versus phospholipid ratio. Furthermore, a progressive decrease of surfactant levels in older horses may contribute to a worsening of clinical signs in older RAO-affected horses. / Ph. D.

In healthy airways, mucus forms part of the innate immune response protecting the respiratory epithelium from damage by pathogens and environmental debris (Rose and Voynow, 2006). Conversely, in many respiratory diseases, mucus becomes part of the airway disease pathology. Mucus hypersecretion along with reduced clearance can cause blockage of the small airways, impairing gas exchange, promoting inflammation and becoming a culture medium for bacterial colonisation (Thornton et al., 2008). Recurrent airway obstruction (RAO) is a common yet poorly understood equine chronic respiratory disease where such altered mucus properties and clearance have been identified as major factors in the disease pathology (Davis and Rush, 2002; Gerber et al., 2000; Kaup et al., 1990; Robinson, 2001). The gel-forming mucins are largely responsible for the transport properties of mucus. The major equine airway gel-forming mucin in health is Muc5b and to a lesser extent Muc5ac; produced in specialised respiratory epithelial goblet cells and sub-mucosal glands (Rousseau et al., 2011b). Changes in mucin relative and net amounts and their macromolecular properties and interactions have been attributed to the altered physical properties of airway mucus in airways disease (Groneberg et al., 2002a; Jefcoat et al., 2001; Kirkham et al., 2002; Robinson et al., 2003; Sheehan et al., 1995).The project investigates the biochemical properties of mucins present in mucus from healthy horses and horses with RAO. This project identifies the anatomical presence of mucin-producing goblet cells and glands in fixed tissues from the respiratory tracts of healthy horses and subsequently examines mucin-production sites in respiratory tracts from horses with RAO. Finally the project investigates a methodology for the study of mucin production in airway cells harvested from live horses suffering from RAO.Our investigations confirmed that horses with RAO have more endotracheal mucus than healthy controls, and that Muc5b is the predominant mucin with Muc5ac also present in RAO horse mucus, both during symptomatic disease and when horses are asymptomatic. Mucins are produced in epithelial goblet cells and sub-mucosal glands dispersed throughout the length and circumference of the equine trachea and bronchi. Goblet cell hyperplasia occurs in symptomatic exposed RAO horse airways, although goblet cells are smaller than in asymptomatic RAO horse airways. Exposure to a dusty stable environment is associated with more goblet cells per length of bronchial compared to tracheal epithelium in all horses. RAO horses have larger sub-mucosal glands containing more mucin than control horses. Primary epithelial cell cultures grown at an air liquid interface are an alternative approach to study equine airway mucus, although the use of this culture system is in its early stages. We have developed novel ways to harvest equine airway epithelial cells (tracheal brushing) and shown it is possible to freeze cells collected via tracheal epithelial brushing in 20 % FBS and then culture to ALI at a later date.

by Martin Li. / Year shown on spine: 1997. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1996. / Includes bibliographical references (leaves 150-157). / DECLARATION --- p.II / ABSTRACT --- p.III / ACKNOWLEDGEMENTS --- p.VII / CONTENTS --- p.VIII / LIST OF ABBREVIATIONS --- p.XIV / LIST OF TABLES --- p.XVI / LIST OF CHART --- p.XXIII / LIST OF FIGURES --- p.XXIV / Chapter CHAPTER 1 --- OBSTRUCTIVE AIRWAY DISEASE: PUBLIC HEALTH AND CLINICAL ASPECTS --- p.1 / Chapter 1.1. --- Background --- p.1 / Chapter 1.2. --- Magnitude of the problem --- p.2 / Chapter 1.2.1. --- Asthma --- p.2 / Chapter 1.2.2. --- Chronic obstructive pulmonary disease --- p.3 / Chapter 1.2.3. --- Prevalence of osteoporosis in Hong Kong --- p.4 / Chapter 1.2.4. --- History of asthma care --- p.5 / Chapter 1.2.5. --- Treatment of OAD --- p.5 / Chapter 1.3. --- Side effects of Glucocorticoid in OAD patients --- p.6 / Chapter 1.4. --- Side effccts of inhaled corticosteroids in OAD patients --- p.7 / Chapter 1.5. --- Trend of asthma therapy in Hong Kong --- p.8 / Chapter CHAPTER 2: --- OSTEOPOROSIS: PUBLIC HEALTH AND CLINICAL ASPECTS --- p.11 / Chapter 2.1. --- Bone Biology --- p.11 / Chapter 2.2. --- Skeletal Organisation --- p.11 / Chapter 2.3. --- Bone remodelling --- p.12 / Chapter 2.4. --- Effect of corticosteroids on bone remodelling --- p.13 / Chapter 2.5. --- Corticosteroids induccs osteoporosis --- p.13 / Chapter 2.6. --- Factors affecting BMD --- p.14 / Chapter 2.6.1. --- Peak bone mass --- p.14 / Chapter 2.6.2. --- Ethnic factors --- p.14 / Chapter 2.6.3. --- Aging --- p.15 / Chapter 2.6.4. --- Calcium intake --- p.15 / Chapter 2.6.5. --- Oestrogen --- p.16 / Chapter 2.6.6. --- Alcohol consumption --- p.17 / Chapter 2.6.7. --- Cigarette smoking --- p.17 / Chapter 2.7. --- Physical activity and BMD --- p.17 / Chapter 2.8. --- Body composition in Chinese subjects --- p.18 / Chapter CHAPTER 3 --- "PHASE I: BODY COMPOSITION AND BONE MINERAL DENSITY IN OBSTRUCTIVE AIRWAY DISEASE PATIENT AND NORMAL CONTROL SUBJECTS: OBJECTIVES, SUBJECTS AND METHODS" --- p.20 / Chapter 3.1. --- Objectives --- p.20 / Chapter 3.2. --- Subjects and methods --- p.21 / Chapter 3.2.1 --- OAD patients --- p.21 / Chapter 3.2.1.1 --- Disease definition and selection criteria --- p.21 / Chapter 3.2.1.2. --- Normal Control subjects --- p.21 / Chapter 3.3. --- Power of estimation --- p.22 / Chapter 3.4. --- Survey methods --- p.22 / Chapter 3.5. --- Questionnaire --- p.23 / Chapter 3.6. --- Body composition and bone mineral density measurement --- p.23 / Chapter 3.6.1. --- Body composition analysis --- p.24 / Chapter 3.6.2. --- Lumbar spine and proximal hip bone mineral density analysis --- p.24 / Chapter 3.6.3. --- Routine quality control of measurements --- p.24 / Chapter 3.6.4. --- Precision on patient repositioning --- p.25 / Chapter 3.7. --- Statistical methods --- p.25 / Chapter 3.8. --- Bone mineral density of normal control subjects --- p.25 / Chapter CHAPTER 4 --- PHASE II: FLUORIDE IN THE TREATMENT OF OSTEOPOROSIS --- p.27 / Chapter 4.1. --- Introduction --- p.27 / Chapter 4.2. --- Mechanisms of action --- p.28 / Chapter 4.2.1. --- Antiresorptive effect of fluoride --- p.28 / Chapter 4.2.2. --- Force-oriented osteogenic effect of fluoride --- p.28 / Chapter 4.2.3. --- Biochemical osteogenic effect --- p.29 / Chapter 4.3. --- Effect of fluoride salts on BMD: results of clinical trials --- p.29 / Chapter 4.4. --- Effcct of fluoride on bone histomorphology --- p.30 / Chapter 4.5. --- Compliance with sodium fluoride therapy --- p.31 / Chapter 4.6. --- Contradiction of fluoride treatment --- p.31 / Chapter 4.7. --- Sodium monofluorophosphate preparation --- p.32 / Chapter CHAPTER 5 --- PHASE II: THE EFFECTS OF FLUORIDE ON BONE MINERAL DENSITY OF OAD PATIENTS ON STEROID TREATMENT --- p.37 / Chapter 5.1. --- Objectives --- p.37 / Chapter 5.2. --- Subjects and methods --- p.37 / Chapter 5.2.1. --- Power of the study --- p.37 / Chapter 5.2.2. --- Subjects --- p.37 / Chapter 5.2.3. --- Method of randomisation --- p.38 / Chapter 5.2.4. --- Treatment modalities --- p.39 / Chapter 5.2.4.1. --- Treatment group --- p.39 / Chapter 5.2.4.2. --- Control group --- p.39 / Chapter 5.2.5. --- Bone mineral density measurements --- p.39 / Chapter 5.2.6. --- Routine quality control of measurement and precision on patient repositioning --- p.40 / Chapter 5.2.7. --- Methods of monitoring drug compliance --- p.40 / Chapter 5.2.8 --- Statistical methods --- p.40 / Chapter CHAPTER 6 --- RESULTS FOR PHASE I --- p.42 / Chapter 6.1. --- Statistical power of this phase of the study --- p.42 / Chapter 6.2. --- Clinical features of OAD subjects on inhaled steroid --- p.42 / Chapter 6.3. --- Anthropometric measurements and bone mineral density --- p.45 / Chapter 6.4. --- Analysis of covariance for BMDs differences --- p.48 / Chapter 6.5. --- Multiple regression --- p.50 / Chapter 6.6 --- Correlation --- p.51 / Chapter CHAPTER 7 --- RESULTS FOR PHASE II: FLUORIDE AND CALCIUM TRIAL --- p.81 / Chapter 7.1. --- Factors affects the power of studies --- p.81 / Chapter 7.2. --- Clinical findings --- p.82 / Chapter 7.3. --- Body measurements and bone mineral densitometry --- p.85 / Chapter CHAPTER 8: --- DISCUSSION FOR PHASE I --- p.117 / Chapter CHAPTER 9: --- DISCUSSION FOR PHASE II: TRIDIN AND CALCIUM TRIAL --- p.124 / APPENDIX 1: QUESTIONNAIRE FOR OAD BONE MINERAL DENSITY STUDY --- p.132 / APPENDIX 2: BONE SCANS FROM HOLOGIC QDR2000 --- p.137 / APPENDIX 3. TABLES AND REFERENCE CURVES FOR NORMAL HONG KONG CHINESE FEMALE OR MALE BMD --- p.142 / REFERENCE --- p.150

Objetivos: Determinar o percentual de asmáticos com má percepção da dispnéia e sua correlação com a intensidade da broncoconstrição aguda, hiper-responsividade brônquica, uso de medicação de manutenção e controle da asma. Métodos: Ensaio clínico não controlado com pacientes asmáticos do ambulatório de asma do Hospital São Lucas da PUCRS. Foram realizados testes de broncoprovocação com metacolina com protocolo dosimetrado e avaliada a percepção da dispnéia após cada dose administrada, utilizando a escala de Borg. Foram coletados dados demográficos e questionado quanto ao controle da asma, medicação em uso e necessidade de broncodilatador de curta ação. Para análise estatística foram utilizados teste de Chi-Quadrado e Teste t de Student e correlação de Spearman. Resultados: Foram estudados 65 pacientes com asma, dos quais 53 tiveram sua avaliação completa. Trinta e dois pacientes apresentaram percepção da dispnéia (60,5%) quando ocorreu broncoconstrição induzida pela metacolina. Entretanto, 21 pacientes (39,5%) não apresentaram alteração em relação aos sintomas de dispnéia, mesmo com queda de 20% do VEF1. Os grupos dos percebedores e não percebedores não apresentavam diferenças quanto ao VEF1 basal, percentagem de queda do VEF1 e dose de metacolina necessária para broncoprovocação. Não houve correlação significativa entre percepção da dispnéia com idade (p=0,247), sexo (p=0,329), uso de medicação de manutenção (p=0,152), controle da asma (p=0,562), hiper-responsividade brônquica (p=0,082) e gravidade da broncoconstrição aguda (p=0,749). Conclusões: Percentagem significativa dos asmáticos apresenta baixo grau de percepção da broncoconstrição. Os fatores relacionados com a incapacidade de identificação da modificação da função pulmonar não estão bem definidos. Diagnóstico e orientação dos maus percebedores é fundamental para redução de morbidade e mortalidade por asma. / Objective: To assess the percentage of poor perception of dyspnea in asthmatics and its correlation with acute bronchoconstriction severity, airway hyperresponsiveness, medication use and asthma control. Methods: Uncontrolled clinical trial of asthmatics from outpatient department HSL-PUCRS. Methacholine challenge testing was performed with five-breath dosimeter protocol. The perception of airway narrowing after 20% fall in FEV1 was evaluated using the Borg scale. Data concerning demographic information, asthma control, long-term management medication and rescue medication consumption were recorded. Chi-square test and Student´s T test and Spearman’s correlation were applied for the statistical analysis. Results: 65 patients were included and 53 completed the evaluation. 32 patients presented dyspnea (60,5%) when methacholine induced bronchoconstriction occurred but 21 patients (39,5%) did not show any difference related to dyspnea symptoms, even with 20% fall in FEV1. There were no differences between the two groups in terms of the baseline FEV1, % of fall FEV1, and methacoline dose that promoted a positive test. There was no significant association between airway obstruction and age (p= 0.247), sex (p=0.329), long term management medication use (p=0.152), asthma control (p=0.562), airway hyperresponsiveness (p=0.082), and acute bronchoconstriction severity (p=0.749). Conclusion: Significant percentage of astmatics presents poor perception of bronchoconstriction. The identification and orientation of this group of patients is essential to make plans of interventions and eventually reduce asthma morbidity and mortality.

ABSTRACT Heaves is triggered by exposure to dust and its components, such as endotoxin, and is characterized by clinical signs such as coughing, decreased exercise tolerance, difficulty breathing and abnormal lung sounds which are due to bronchoconstriction and accumulation of neutrophils in the airways. Pulmonary intravascular macrophages (PIMs) are believed to increase horses sensitivity to endotoxemia-induced lung inflammation. The first objective of this study was to investigate a hitherto unknown role of PIMs in equine heaves. I used mouldy hay (MH) to induce heaves and gadolinium chloride (GC) to deplete PIMs in order to compare responses between non-treated and GC-treated heaves horses. A modified randomized crossover study (2X2 factorial) was conducted in which mares (N=9) were exposed to 4 different treatments: alfalfa cubes (Cb), alfalfa cubes + GC (Cb-GC), mouldy hay (MH) and MH + GC (MH-GC). Each treatment was followed by broncholaveolar lavage (BAL). MH was fed for 7 days to induce heaves followed by Cb for 21 days to achieve remission, whereas the treatments in which heaves was not induced (Cb; Cb-GC), the cubes were fed prior to the BAL and for 14 days after the BAL to allow recovery from the BAL procedure. BAL fluids were processed to investigate total cell, neutrophil and alveolar macrophage concentrations. In addition, TNFá protein levels as well as TNFá, IL-8, and TLR4 mRNA expression in BAL cells were assessed in order to infer on their activation state.<p> Data showed higher concentration of dust (3X), endotoxin (20X), and endotoxin per milligram of dust (7X) in MH compared to the Cb environment. Clinical scores and neutrophil concentrations in BAL were higher when mares received MH compared to MH and GC (MH-GC). Real time reverse transcriptase PCR revealed a significant lower expression of IL-8 and TLR4 mRNA in BAL cells from MH-GC mares compared to MH. TNFá mRNA expression as well as protein concentration were not affected by the different treatments. In vitro secondary LPS challenge significantly increased IL-8 mRNA expression in cells from MH treatment compared to without LPS, but not in the MH-GC treatment. TLR4 expression was not affected by the secondary challenge. Although secondary LPS challenge increased expression of TNFá mRNA and protein, the differences among treatment groups were not meaningful. In conclusion, PIM depletion attenuates clinical scores, migration of inflammatory cells into the alveolar space and expression of pro-inflammatory molecules in BAL cells of heaves horses.<p> The observations on the role of PIMs in heaves in horses prompted me to examine the occurrence of PIMs in human lungs. I found a trend for higher numbers of septal macrophages in autopsied lungs from human patients who died of non-pulmonary pathologies compared to those having either COPD or asthma. If these septal macrophages indeed represent the PIMs, this finding is contrary to existing belief that humans, unlike horses, do not have PIMs.

ABSTRACT Heaves is triggered by exposure to dust and its components, such as endotoxin, and is characterized by clinical signs such as coughing, decreased exercise tolerance, difficulty breathing and abnormal lung sounds which are due to bronchoconstriction and accumulation of neutrophils in the airways. Pulmonary intravascular macrophages (PIMs) are believed to increase horses sensitivity to endotoxemia-induced lung inflammation. The first objective of this study was to investigate a hitherto unknown role of PIMs in equine heaves. I used mouldy hay (MH) to induce heaves and gadolinium chloride (GC) to deplete PIMs in order to compare responses between non-treated and GC-treated heaves horses. A modified randomized crossover study (2X2 factorial) was conducted in which mares (N=9) were exposed to 4 different treatments: alfalfa cubes (Cb), alfalfa cubes + GC (Cb-GC), mouldy hay (MH) and MH + GC (MH-GC). Each treatment was followed by broncholaveolar lavage (BAL). MH was fed for 7 days to induce heaves followed by Cb for 21 days to achieve remission, whereas the treatments in which heaves was not induced (Cb; Cb-GC), the cubes were fed prior to the BAL and for 14 days after the BAL to allow recovery from the BAL procedure. BAL fluids were processed to investigate total cell, neutrophil and alveolar macrophage concentrations. In addition, TNFá protein levels as well as TNFá, IL-8, and TLR4 mRNA expression in BAL cells were assessed in order to infer on their activation state.<p> Data showed higher concentration of dust (3X), endotoxin (20X), and endotoxin per milligram of dust (7X) in MH compared to the Cb environment. Clinical scores and neutrophil concentrations in BAL were higher when mares received MH compared to MH and GC (MH-GC). Real time reverse transcriptase PCR revealed a significant lower expression of IL-8 and TLR4 mRNA in BAL cells from MH-GC mares compared to MH. TNFá mRNA expression as well as protein concentration were not affected by the different treatments. In vitro secondary LPS challenge significantly increased IL-8 mRNA expression in cells from MH treatment compared to without LPS, but not in the MH-GC treatment. TLR4 expression was not affected by the secondary challenge. Although secondary LPS challenge increased expression of TNFá mRNA and protein, the differences among treatment groups were not meaningful. In conclusion, PIM depletion attenuates clinical scores, migration of inflammatory cells into the alveolar space and expression of pro-inflammatory molecules in BAL cells of heaves horses.<p> The observations on the role of PIMs in heaves in horses prompted me to examine the occurrence of PIMs in human lungs. I found a trend for higher numbers of septal macrophages in autopsied lungs from human patients who died of non-pulmonary pathologies compared to those having either COPD or asthma. If these septal macrophages indeed represent the PIMs, this finding is contrary to existing belief that humans, unlike horses, do not have PIMs.

Thesis (Ph. D.)--University of Oklahoma. / Bibliography: leaves 116-150.