Thesis (doctoral)--University of Tartu, Estonia, 1995. / Vita. Includes bibliographical references.

Thesis (Ph. D.)--University of Wisconsin--Madison, 1965. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

Idiopathic pulmonary fibrosis (IPF) is a fibrotic lung disease with no effective treatment. Myofibroblasts contribute to the pathology of IPF by secreting large amounts of extracellular matrix proteins such as alpha smooth muscle actin (α-SMA) and Collagen I (Col 1). Myofibroblasts have reduced Prostaglandin E2 (PGE2), a key anti-fibrotic mediator, due to diminished cyclooxygenase-2 (COX-2) expression. Primary fibroblasts isolated from lungs of IPF patients (F-IPF) expressed significantly less COX-2 in response to IL-1β and increased α-SMA and Col I compared with fibroblasts isolated from lungs of non-fibrotic patients (F-NL). COX-2 was gradually lost in F-NL treated with transforming growth factor-β (TGF-β1), a pro-fibrotic cytokine, whereas PGE2, and cAMP elevating agents increased IL-1β-induced COX-2 expression in F-IPF. Ras, a small G protein, has been shown to have a role in several fibrotic conditions. Farnesylthiosalicylic acid (FTS), a Ras inhibitor, increased IL-1β-induced COX-2 and prevented TGF-β1-induced reduction of COX-2. Previous studies suggest that COX-2 is epigenetically repressed. LBH589, a HDAC inhibitor, prevented TGF-β1-induced repressed COX-2 whereas BIX01294, a DNA lysine methyltransferase inhibitor, and RG108, a G9a histone methyltransferase inhibitor, both increased IL-1β-induced COX-2 in F-IPF. In conclusion, the gradual loss of PGE2/COX-2 anti-fibrotic mechanism during myofibroblast differentiation may contribute to the pathophysiology of pulmonary fibrosis and agents that increase cAMP levels, inhibit Ras or inhibit epigenetic repression of COX-2, may compensate for the lack of endogenous PGE2.

Background Airways diseases are increasingly recognised to be poorly defined phenomena with overlapping pathophysiology and symptoms. They are a significant and growing cause of morbidity, with increasing numbers of people affected globally and no improvement in key outcomes in the UK for the last decade despite ever increasing expenditure. The classification of airway diseases has changed little in the last 50 years, and may no longer be fit for purpose due to the growing appreciation of the complexity and heterogeneity of airways disease and the advent of molecular-based diagnostic techniques to target specific treatment. Aim To investigate whether strategies based on the measurement of selected phenotypic and biological characteristics of airways disease can help to improve the understanding of their pathogenesis and targeting of treatment. Methods Three characteristics of airways disease, namely (1) exhaled nitric oxide, (2) chronic productive cough of unknown cause and (3) the airway microbiota were described/measured in selected cohorts of patients in three clinical studies. Measurement of each of these characteristics was used to answer focused clinical questions regarding the pathogenesis and treatment of aspects of airways disease. Results (1) The baseline measurement of FENO in steroid naïve subjects with symptoms suggestive of asthma had a low diagnostic value for asthma but was an excellent predictor of inhaled steroid treatment response. (2) A cohort of subjects with chronic productive cough of unknown cause was described. These subjects tended to have radiological evidence of airway dilatation and chronic inflammatory changes but not significant bronchiectasis. Their cough responded well to treatment with azithromycin, with ongoing neutrophilic airway inflammation a particularly strong predictor of treatment response. (3) There were no significant differences in the abundance or community structure of the bacterial communities in the airways between subjects with mild (BTS 2) or severe (BTS 4) asthma or between severe (BTS 4) asthma patients taking inhaled fluticasone or budesonide. However a number of differences in relative abundance of certain species (including enrichment of Haemophilus parainfluenzae in the fluticasone group) were noted on comparison of the groups. Conclusions This thesis provides support for a new approach to the classification and treatment of airways disease. The recognition of pathologically important processes (treatable traits) which can be used to predict response to targeted treatment has the potential to revolutionise the management of airways disease and result in improved patient outcomes.

Dexamethasone, a glucocorticoid, is used extensively in clinical medicine in the treatment of respiratory diseases, notably asthma. This medical effect is probably due in part to a down-regulation of many cytokines, e.g. IL-8. However, the side effects of steroids often may outweigh the beneficial effects, especially if the disease in question is of a chronic nature. This combined with steroid resistance has led to an increase in the use of non-steroidal anti-inflammatory drugs. These tend to be aspirin derivatives, and many have side-effects. I have investigated a factor that is naturally produced by the body which may alleviate the symptoms of inflammation without many of the side- effects, with an aim to providing a viable alternative. A novel steroid induced monocyte derived factor was described twenty years ago by Stevenson as part of an MD thesis submitted to this university. This remains an unusual phenomenon, as most of the effects of steroids are due to a down-regulation of release; here is one of the few examples of the reverse. Although it raised many interesting points about dispersive motility of populations of polymorphonuclear leukocytes (PMN) it did not look at individual cells, nor at many of the interesting properties of neutrophils. One of the known effects of some steroids (though not Dexamethasone), and many NSAIDs, is a reduction of adhesion of PMN to venous endothelium. Human umbilical vein endothelial cells (and bovine aorta endothelium) were used as an ex vivo model for this, and my results show there is a marked decrease in this adhesion. This effect was observed not only on resting endothelium, but also when the endothelial cells had been pre-treated with ILip or thrombin. This decreased adhesion was due to an interaction with the PMN and the factor, as shown by adhesion being reduced when protein coated coverslips were used as the substrate for adhesion. It is suspected that the reasons for this may be due to an inactivation of integrins such as Mac-1, although a role for selectins cannot be ruled out. Recent work by Diaz-Gonzalez suggests that some NSAIDs act by inducing PMN to shed L-selectin. Other effects of this steroid induced monocyte derived factor on human PMN were determined using biological and biochemical techniques. Previous work has shown a novel dispersive effect of this factor on PMN when used in a uniform concentration, and so I decided to look at the moiphology of treated PMN. Using scanning electron microscopy I observed a polarisation of the PMN, but without any raffling of the membrane, a feature that is normally observed with polarisation. This morphology was not observed with control supernatants taken from monocytes cultured in the absence of Dexamethasone. This moiphology was conserved using cells in suspension and adhered to bovine aorta endothelium. The underlying actin cytoskeleton was examined using confocal microscopy, and the microfilamentous airay was noted as being devoid of spikes, observed with activation, for example with fMLP. Late cytoskeletal controlled effects, the release of granule contents, were also investigated, and it was noted that the release of primary granule contents could be inhibited by this factor in a dose dependent manner. This fusion of granules with the plasma membrane is controlled by the activation of numerous tyrosine kinases, and follows a strict order. Secondary granule release was shown to be inhibited also, as assayed by the cleavage of type I collagen, and analysis of SDS gels. The effects on the metabolic burst showed conflicting results with inhibition being present, again in a dose dependent manner, but much of this activity was removed with increasing purification, leaving only very slight inhibition.

No description available.

No description available.

This thesis is focussed on developing new methods and outcomes to assess respiratory function that require little or no volitional effort on behalf of the participants being tested. Specifically to attempt to detach the behaviour of the patient from the accuracy of the test of respiratory function, resulting in techniques that are simpler and easier to administer and undertake for both assessor and participant. It aims to develop methods that reduce the involvement of the participant during assessment of respiratory function. The human body’s way of controlling respiration has evolved into a sophisticated system that optimises breathing pattern to maintain the most efficient homeostatic action of the respiratory system. Eliciting and assessing this automatic response is the key to removing the action of participation from respiratory functiontesting. The focus must therefore be on developing non-invasive, sub-maximal techniques that allow participants to enter into a steady state of respiration and how this can be assessed. Two techniques were investigated; Respiratory Endurance (as the inspiratory work of breathing) and Tidal Breathing Flow Profile, and these were successfully applied in 99 adult participants (68 healthy controls and 31 COPD patients) and 75 children (48 clinical group and 27 healthy controls) who completed 467 respiratory endurance trials whilst seated and exercising, and 249 relaxed tidal breathing trials. The difficulties with lung function assessment are well established and have been described in this thesis. Much recent emphasis has been put on developing existing devices and protocols rather than developing new techniques and approaching these difficulties from alternative viewpoints. This thesis has described the development of innovative techniques to assess the function of the respiratory systems that aim to overcome the issues associated with maximal testing. It was shown that these techniques are easy to undertake for a range of participants, simple to analyse and are able to reliably differentiate between health and disease, suggesting that they could become a useful adjunct to existing methods of respiratory assessment.

The urokinase plasminogen activator receptor (uPAR, PLAUR) is a cell surface receptor actively involved in the regulation of cell homeostasis. Expression is elevated in the bronchial epithelium in vivo and also in serum and sputum in asthma and elevated expression often indicates poor prognosis in a number of human diseases. The relative contribution of uPAR to asthma disease mechanisms is not fully understood and the functional roles of uPAR isoforms remains to be resolved. The key aims of this thesis were to i) investigate how the uPAR pathway may influence bronchial epithelial barrier properties; ii) investigate the gene expression patterns in the bronchial epithelium in asthma; iii) identify functions of different forms of uPAR in human bronchial epithelial cells (HBEC) and to iv) investigate the association between genetic polymorphisms spanning the PLAUR gene with clinical features and the presentation of asthma in moderate to severe asthma. Using two cell based approaches we identified an inverse relationship between soluble-cleaved uPAR expression and epithelial barrier properties. Importantly, we demonstrated that blocking uPAR-integrin interactions provides a potential therapeutic opportunity to improve epithelial barrier function. Using whole transcriptome analysis genes differentially expressed between cultured asthma and control subjects were identified which were related to cell growth, repair and immune regulation. Furthermore, uPAR expression was elevated in epithelial cells in asthma subjects compared to healthy controls, suggesting expression is inherently altered in the bronchial epithelium in asthma. Transcriptomics was used to provide novel insight into the specific and overlapping functions of uPAR isoforms and to determine the effects of elevated uPAR expression on HBEC function. Finally, the contribution of PLAUR genetic variants to clinical and immunological traits within asthma were investigated and found that PLAUR single nucleotide polymorphisms (SNPs) did not show an association with the traits measured in a severe asthma population. Overall this work has provided new insight into the function of uPAR as a regulator of the bronchial epithelium in asthma.

Lung cancer is the most commonly diagnosed cancer and the most common cause of cancer related death worldwide. Population-based lung cancer screening programmes have been initiated in the USA and could soon be implemented in other countries. The overarching purpose of this thesis was to explore the emotional impact of lung cancer screening. The research was conducted as part of a clinical trial that was investigating the effectiveness of a blood autoantibody test, EarlyCDT®-Lung, in identifying individuals at the risk of lung cancer. A systematic review was conducted that aimed to identify factors associated with the emotional impact of screening for lung cancer. Participants with indeterminate test results, current smokers and females were more likely to experience negative non-specific and specific emotional outcomes. In addition to highlighting several key factors associated with higher levels of emotional distress following screening, factors that warranted further research were also identified. Such factors included age, education level, marital status, ethnic origin, and perceived risk of developing lung cancer. Finally, important methodological and theoretical limitations in the literature were identified. One key methodological limitation was that no studies measured positive emotional outcomes. A longitudinal study was conducted exploring the impact of lung cancer screening on positive affect, negative affect, lung cancer worry and distress specific to screening for lung cancer. Participants from each of the EarlyCDT®-positive, EarlyCDT®-negative, and control groups completed questionnaires containing emotional outcome measures at pre-randomisation and then at one, three, six and 12 months post randomisation. Scores for each outcome measure were described by groups over time and multilevel regression modelling was used to compare scores over time within and between groups. Results were reassuring as screening was found to have no clinically important impact on positive affect, negative affect, frequency of lung cancer worry or impact of lung cancer worry on mood and ability to perform daily activities. Although screening specific distress in the EarlyCDT®-positive group was significantly higher than that of the EarlyCDT®-negative group, it did reduce over time. Statistically significant and clinically important increases in the proportion of participants reporting anxiety about the results of future tests/treatments were identified. As a result of this finding, a further study was carried out to identify factors that could influence an individual’s level of anxiety about the results of future tests/treatment. Participants more at risk of reporting anxiety about the results of future tests/treatment were younger participants, non-white participants, current smokers and participants who did not own or have a mortgage on their home. Psychological variables associated with increased anxiety were: higher general anxiety scores, higher depression scores, higher negative affect scores, participants who reported that they were upset when they thought about their risk of lung cancer, participants who were worried about getting lung cancer, and those who reported the highest impact of lung cancer worry on mood and ability to perform daily activities. The final chapter of this thesis presents the results of a randomised controlled trial embedded within the emotional outcomes study (described above), which evaluated the effect of timing of monetary incentives (£5 voucher sent with questionnaire vs. £5 voucher sent on receipt of questionnaire) on the following outcomes: study participation rates, questionnaire response rates over time, the number of reminders sent and the completeness of returned questionnaires over time. Previous research had found that monetary incentives were useful in increasing response rates in clinical trials. Results from this trial extended the evidence base by showing that the timing of monetary incentives makes no difference to the above outcomes. In each chapter the findings of this thesis are discussed in terms of their contribution to knowledge. Recommendations for future research and clinical practice are also made within each chapter.