Onchocerciasis affects an estimated 37 million people in Sub-Saharan Africa and Latin America, causing debilitating skin and eye disease and accounting for a global loss of 1 million Disability-Adjusted Life-Years. The discovery of Wolbachia bacterial endosymbionts in filarial nematodes, including Onchocerca volvulus, has revolutionised the understanding of the parasite’s biology and immunopathogenic mechanisms of disease, and has offered a novel approach to its treatment and control with anti-Wolbachia antibiotics. Treatment with doxycycline is effective at sterilising and killing adult O. volvulus worms, proving superior to standard microfilaricidal treatment with ivermectin and of great potential as an alternative strategy for the control of onchocerciasis. Although the length of the required treatment has raised concerns about the use of doxycycline in Mass Drug Administration (MDA) strategies, a recent trial in Cameroon demonstrated the feasibility of a six week course of doxycycline MDA delivered with a community-directed approach. In the work presented here we found a significant reduction in microfilaridermia prevalence and loads four years after doxycycline MDA distribution, demonstrating its long-term effectiveness and supporting its implementation in existing control strategies. Wolbachia peptidoglycan-associated lipoprotein stimulates innate and adaptive immune responses, contributing to disease pathogenesis through the induction of pro-inflammatory cytokines and recruitment of neutrophils. These features together with the cytokine milieu induced by filarial nematodes could support the development of a pro-inflammatory Type-17 immune response. In this work, PBMC from patients with onchocerciasis were found to produce only minimal levels of IL-17 in response to filarial extracts. On the contrary, a rich IL-17+ cell infiltrate was found surrounding adult worms in Wolbachia positive onchocercomas using immunohistochemistry (IHC), which was depleted from onchocercomas following doxycycline treatment. Although a high percentage of Th17 cells were present in this infiltrate compared to other diseases, the majority of IL-17 producing cells in nodules were neutrophils, within an extracellular trap-like structure. This unexpected result was consistent with the reported IL-17 production by human neutrophils using IHC in the literature, but could not be confirmed at the protein or the transcription level in vitro in this work. Wolbachia is responsible for an abundant neutrophil infiltration in Onchocerca-infected tissues. However, the role of neutrophils in the immune response to filarial parasites and their interaction with Wolbachia is poorly known. The work presented here showed that Wolbachia lipoprotein activates human neutrophils in vitro, supporting previous studies demonstrating the link between Wolbachia and neutrophils in the host inflammatory responses to O. volvulus infection. These results also indentify Wolbachia lipoprotein as a key molecule driving human neutrophil recruitment and activation.

Background: Advanced heart failure is receiving increasing attention from clinicians and policy makers as a major chronic condition associated with poor quality of life in an ageing population. Aim: To explore how we could tailor health interventions to individual patients with advanced heart failure at the end of life. Design: (1) A systematic review of national and international chronic heart failure guidelines using the Appraisal of Guidelines for Research and Evaluation (AGREE II) tool and a data extraction framework based on the holistic needs assessment of the Gold Standards Framework (GSF). (2) A longitudinal qualitative study of 15 patients with New York Heart Association grade 3 or 4 heart failure recruited through two community based heart failure services. Semi-structured interviews were conducted with patients at 3 monthly intervals for 1 year (n= 52 interviews). A refined ‘case-based’ method as described by Griffiths et al. was adopted to identify the ideal type categories of adjustment and adaptation and assess how these categories change over time for each patient. This involved understanding individuals as complex systems, subject to internal and external influences, with the potential for transformation. The analysis drew on the theoretical concept of the emergent present - as developed by Adam - the current period of time when all domains of life have expression. Results: (1) A total of 19 guidelines were included in the review. Across all guidelines the lowest scoring domains were applicability and stakeholder involvement. Qualitative assessment showed that most guidelines adopt a disease-orientated approach to addressing need. In particular, domains on continuity of care and out of hours care were poorly covered. (2) Four distinct patterns of adjustment and adaptation were identified. The largest group was the Stuck and struggling category, which was characterised by participants wanting to move on but being unable to do so. Participants in the integrating group were able to accommodate the problems that they faced from moment to moment despite anticipating an uncertain future. Those in the submerged group were completely immersed in their illness and any expectation of a meaningful future had completely disappeared. The Past reminder group was characterised by a narrative based in the emergent present that was dominated by their experience of previous events. It was shown that some participants transformed from one category to another as a result of the care they received. For others, there was no change over the course of the study. Conclusion: This thesis identifies important differences between the ‘objective’ patient represented in clinical guidelines and the ‘subjective’ experience of the individual. The illness experiences of people living with advanced heart failure are diverse and do not lend themselves to standardised care. This raises important questions for the way knowledge is currently translated into clinical practice. Attending to the emergent present may be a clinically useful approach for supporting health care professionals to tailor care to needs of patients at the end of life.

EPLIN (Epithelial Protein Lost In Neoplasm) is a cytoskeletal associated protein whose expression is often reduced in cancer cells. It may function as a tumour suppressor through its effects on cancer cell migration and invasion. To date, its role in wound healing has not been elucidated. We examine the impact of EPLIN on keratinocyte migration and its implications in wound healing. A mammalian expression construct containing the full EPLIN coding sequence was used to overexpress EPLIN in human keratinocyte cell (HaCaT). Following overexpression verification, the impact of EPLIN on HaCaT cell migration was assessed using a conventional scratch wounding assay and an electric cell-substrate impedance sensing (ECIS) system-based assay. Protein expression was examined using western blot, ICC and IFC analysis. Transfection of HaCaT cells with the EPLIN expression construct successfully resulted in enhanced HaCaT EPLIN expression. Enhanced EPLIN levels were seen to negatively impact on cell migration as determined by both the scratch wound assay and the ECIS model system with migration rates of HaCaT cells overexpressing EPLIN being substantially less than the control HaCaT cells. Overexpression of EPLIN was found to slow keratinocyte migration rates using two independent assays as well as show convincing association and interaction with two NWASP and E-Cadherin. These important findings suggest novel routes to positively manipulate the wound healing process and has significance in further translational research.

There is a growing demand worldwide to increase the quality and productivity of healthcare services thereby increasing the value of the healthcare services delivered. To deal with these demands, increasingly importance is being placed on analysing and reducing unwarranted variations in healthcare services to achieve significant savings in healthcare expenditure. Unwarranted variations are defined as the variations in the utilisation of healthcare services that cannot be explained by variation in patient illness or patient preferences. Current modelling and simulation approaches for healthcare service efficiency and effectiveness improvements in hospitals do not utilise multiple types of heterogeneous service data such as qualitative information about hospital services and quantitative data such as historic system data, electronic patient records (EPR), and real time tracking data for analysing unwarranted variations in hospital. Consequently, due to the presence of large amount of unwarranted variations in the service delivery systems, service improvement efforts are often inadequate or ineffective. Therefore, there is urgent need to: (i) accurately and efficiently model complex care delivery services provided in hospital; (ii) develop integrated simulation model to analyse unwarranted variations on a care pathway of a hospitals; and, (iii) develop analytical and simulation models to analyse unwarranted variations from a care pathway. Current process modelling methods to represent healthcare services rely on simplified flowchart of patient flow obtained based on on-site observations and clinician workshops. However, gathering and documenting qualitative data from workshops is challenging. Furthermore, resulting models are insufficient in modelling important service interactions and hence the resulting models are often inaccurate. Therefore, a detailed and accurate process modelling methodology is proposed together with a systematic knowledge acquisition approach based on staff interviews. Traditional simulation models utilised simplified flow diagrams as an input together with the historic system data for analysing unwarranted variations on a care pathway. The resulting simulation models are often incomplete leading to oversimplified outputs from the conducted simulations. Therefore, an integrated simulation modelling approach is presented together with the capability to systematically use heterogeneous data to analyse unwarranted variations on service delivery process of a hospital. Maintaining and using care services pathway within hospitals to provide complex care to patients have challenges related to unwarranted variations from a care pathway. These variations from care pathway predominantly occur due ineffective decision making processes, unclear process steps, their interactions, conflicting performance measures for speciality units, and availability of resources. These variations from care pathway are largely unnecessary and lead to longer waiting times, delays, and lower productivity of care pathways. Therefore, methodologies for analysing unwarranted variations from a care pathway such as: (i) system variations (decision makers (roles) and decision making process); (ii) patient variations (patient diversion from care pathway); are discussed in this thesis. A system variations modelling methodology to model system variations in radiology based on real time tracking data is proposed. The methodology employs generalised concepts from graph theory to identify and represent system variations. In particular, edge coloured directed multi-graphs (ECDMs) are used to model system variations which are reflected in paths adopted by staff, i.e., sequence of rooms/areas traversed while delivering services. A pathway variations analysis (PVA) methodology is proposed which simulates patient diversions from the care pathway by modelling hospital operational parameters, assessing the accuracy of clinical decisions, and performance measures of speciality units involved in care pathway to suggest set-based solutions for reducing variations from care pathway. PVA employs the detailed service model of care pathway together with the electronic patient records (EPRs) and historic data. The main steps of the methodology are: (i) generate sample of patients for analysis; (ii) simulate patient diversions from care pathway; and, (iii) simulation analysis to suggest set-based solutions. The aforementioned unwarranted variations analysis approaches have been applied to Magnetic Resonance (MR) scanning process of radiology and stroke care pathway of a large UK hospital as a case study. Proposed improvement options contributed to achieve the performance target of stroke services.

Dans ma thèse, j'étudie trois questions importantes dans l'économie industrielle. Chapitre 1 étudie l'impact de l'intégration verticale sur l'innovation dans une industrie où les entreprises doivent entreprendre des R&D investissements risqués à des étapes de production et de distribution. L'intégration verticale permet une meilleure coordination au sein de l'entreprise intégrée, qui renforce son incitation à l'investissement aux niveaux amont et en aval. Cependant, ce n'est que bénéfique que pour les entreprises d'intégrer quand innovations à la fois en amont et en aval sont importantes. Quand l'innovation compte qu'à un seul niveau, les entreprises favorisent la séparation verticale. L'analyse donne un aperçu de la vague de fusions et R&D sous-traitance observées dans l'industrie pharmaceutique et d'autres industries verticale. Chapitre 2 étudie l'effet de la discrimination de la qualité sur la conception des produits. Dans le contexte de l'Internet, les fournisseurs de contenu sont l'objet de discrimination de la qualité des fournisseurs de services Internet. Nous montrons que les fournisseurs de contenu sont biaisées à choisir un design plus larges. Cela réduit la différenciation des produits sur le marché, et l'intervention est nécessaire pour atteindre l'efficacité dans le marché du contenu. Le résultat apporte un nouvel éclairage sur le débat sur la neutralité du net, qui impose l'égalité d'accès à tous les participants sur Internet. Chapitre 3 étudie le rôle de la publicité pour attirer et manipuler l'attention des consommateurs. Quand un produit est caractérisé par plusieurs attributs, les entreprises utilisent aussi stratégique annoncés de manipuler l'attention des consommateurs. Un monopole a tendance à annoncer trop peu d'attributs, et la concurrence n'améliore pas nécessairement la situation. En outre, dans une économie de l'attention-rares, la concurrence pour l'attention des consommateurs conduit les entreprises à annoncer moins d'attributs et réduit l'information à la disposition des consommateurs. / In my thesis, I study three important issues in industrial organization. Chapter 1 studies the impact of vertical integration on innovation in an industry where firms need to undertake risky R&D investments at both production and distribution stages. Vertical integration brings better coordination within the integrated firm, which boosts its investment incentive at both upstream and downstream levels. However, it is only mutually beneficial for firms to integrate when both upstream and downstream innovations are important. When innovation only matters at one level, firms favour instead vertical separation. The analysis provides insights for the wave of mergers and R&D outsourcing observed in the pharmaceutical industry and other vertically related industries. Chapter 2 studies the effect of quality discrimination on product designs. In the context of Internet, content providers are subject to quality discrimination from the Internet Service Providers. We show that content providers are biased to choose broader designs. This reduces product differentiation in the market, and intervention is necessary to achieve efficiency in the content market. The result brings new insights into the discussion about net neutrality, which mandates equal access to every participant on the Internet. Chapter 3 studies the role of advertisements in attracting and manipulating attention from consumers. When a product is characterized by several attributes, firms also strategically use advertisements to manipulate the attention of consumers. A monopolist tends to advertise too few attributes, and competition does not necessarily improve the situation. Moreover, in an attention scarce economy, competition for consumers attention leads firms to advertise fewer attributes and reduces information available to consumers.

Idiopathic pulmonary fibrosis (IPF) is a fatal form of fibrotic lung disease. The pathogenesis of IPF is unclear. An aberrant alveolar epithelial wound repair is likely to be involved in the disease process. Alveolar bronchiolisation, a process where bronchiolar Clara cells migrate into the affected alveoli, is a manifestation of abnormal alveolar wound repair. The role of Clara cells during alveolar injury repair in IPF is controversial. This study was undertaken to investigate the role of Clara cells in alveolar epithelial wound repair and pulmonary fibrosis. Currently, there is no curative treatment for IPF; therefore, stem-cell mediated regenerative therapy has been suggested. In this study, the paracrine role hMSC and hESC on pulmonary epithelial wound repair has also been evaluated. A direct-contact co-culture in vitro model was utilised to evaluate the role of Clara cells on alveolar epithelial cell wound repair. Immunohistochemistry was conducted on IPF lung tissue samples to replicate the in vitro findings ex vivo. The paracrine role of hMSC and hESC on pulmonary epithelial cells was evaluated by utilising the in vitro wound repair system. This study demonstrates that Clara cells induce apoptosis in AEC through a TRAILdependent mechanism, resulting in significant inhibition of wound repair. Furthermore in the IPF lungs, TRAIL-expressing Clara cells were detected within the fibrotic alveoli, together with widespread AEC apoptosis. This study also demonstrates that hMSC enhance AEC and SAEC wound repair via a paracrine mechanism through stimulation of cell migration; whereas, secretory factors of differentiated hESC promote AEC wound repair through stimulation of both cell proliferation and migration. Through this study I propose a novel hypothesis which implies that the extensive profibrotic remodelling associated with IPF could be driven by TRAIL-expressing Clara cells inducing AEC apoptosis through a TRAIL-dependent mechanism. My study also supports the notion of clinical application of hMSC and hESC or their secretory products as regenerative therapeutic modality for IPF.

Non-specific or medically unexplained physical symptoms (MUPS) are common among children, persist in considerable proportions of those affected, and can lead to primary care consultations. A systematic review in this thesis has provided limited evidence of an association between MUPS in parents and children. This thesis has investigated the association between GP consultation for MUPS in 5417 parent-child pairs registered with 12 GP practices, and examined whether this is related to persistent GP consultations for MUPS in children. One descriptive study, two case-control studies, and one prospective cohort study were conducted using GP electronic medical records. In children, the annual GP consultation prevalence for MUPS was 21%, and 12% of all consultations were for MUPS. A significant association was found between consultations for MUPS in mothers and children (adjusted OR 1.42, 95% CI 1.24, 1.63). No association was found between fathers and children, but the association was stronger when both parents consulted for MUPS (adjusted OR 1.52, 95% CI 1.19, 1.93). Significant dose-response relationships were found between numbers of consultations for MUPS and numbers of MUPS in mothers and children. These associations were clearest in maternal-child consultations for painful MUPS and MUPS in specific bodily systems including gastrointestinal, musculoskeletal and neurologic MUPS. Over a quarter (27%) of children who consulted for MUPS at baseline had persistent GP consultations for MUPS at one-year follow-up. Exposure to maternal consultations for MUPS was associated with persistent consultations for similar symptoms in children (adjusted RR 1.29, 95% CI 1.05, 1.58). Exposure to maternal consultations for painful, gastrointestinal, and neurologic MUPS was associated with persistence consultations for similar MUPS in the child. This thesis provides important information about the impact of parental health on child health and consulting behaviour. The implications for primary care and future research are highlighted.

Vascular access via the radial artery has recently been shown to reduce access site related vascular complications but is associated with a significant learning curve. Radial artery spasm, arterial puncture failure, vascular anomalies, failure to reach the ascending aorta and concern regarding higher radiation exposure with the transradial are some obstacles that impede widespread uptake of this technique. This study was performed to assess some of these learning curve issues and to explore the use of transradial access in high-risk patient subgroups. Six interlinked projects were setup for this study and a total of 3125 patients evaluated. Access site vascular complications remain unacceptably high in contemporary practice as discussed in Chapter 2. The transradial approach could minimise such complications. Radial artery anomalies are relatively common and are a common cause of transradial procedure failure as detailed in Chapter 3. Forearm arterial diameter variations and the effect of sublingual GTN were discussed in Chapter 4. The radial artery is bigger than the ulnar artery and GTN increases their diameters by an average of 15-22%. The issues with radiation exposure were studied as detailed in Chapter 5. With strict control of various variables and optimal radiation protection, we demonstrated that there is no difference in radiation exposure between transradial and transfemoral diagnostic angiography when performed by an experienced operator. The application of transradial technique in 2 high-risk patient subgroups was analysed as detailed in Chapter 6. Transradial rescue angioplasty for failed reperfusion and percutaneous right and left heart catheterisation via the arm approach without interruption to Warfarin therapy are found to be safe and effective. These findings have important clinical implication and may help shorten the learning curve and optimise procedure technique including high-risk patient subgroups, thereby help to further drive the adoption of transradial approach.

All the events of egg activation in mammalian eggs are triggered physiologically by transient increases in cytosolic free Ca²⁺ referred to as Ca²⁺ioscillations. These oscillations are initiated by the sperm derived PLC isoform, PLCζ. PLCζ releases Ca²⁺ by hydrolysing its substrate PI(4,5)P₂ to produce IP₃, however, many of the mechanisms by which PLCζ elicits Ca²⁺release in eggs are poorly understood. The results of this thesis confirm that whilstPLCζ cRNA and recombinant protein is able to cause Ca²⁺ioscillations in mouse eggs the sperm derived protein PAWP does not cause any Ca²⁺ release in any circumstances. It is shown that EF hand domain and XY linker of PLCζ are important in determining its Ca²⁺i releasing ability by enabling PLCζ binding to its substrate PI(4,5)P₂ through electrostatic interactions. The C2 domain of PLCζ was also found to play a crucial role in the Ca²⁺ releasing ability of PLCζ, possibly by binding to lipids or proteins in the target membrane. The Ca²⁺releasing ability of eggs is acquired during oocyte maturation and a dramatic increase in PLCζ sensitivity of oocytes occurs after germinal vesicle breakdown. A variety of markers for PLCζ’s substrate PI(4,5)P₂ including fluorescent PI(4,5)P₂ and gelsolin based fluorescent probes suggests that this PI(4,5)P₂ is localised to intracellular vesicles that could derive from Golgi apparatus. Attempts are made to measure PI turnover in these intracellular compartments of eggs during PLCζ induced Ca²⁺i oscillations using several probes. The results of this thesis suggest that PLCζ releases Ca²⁺ by a novel IP₃ based signalling pathway that involves an intracellular source of PI(4,5)P₂.

Chronic neuropathic pain (CNP) occurs as a consequence of injury to the nervous system. Despite recent advances, CNP lacks objective diagnostic criteria, is often unrelenting and refractory to treatment. The primary aims of this thesis are twofold; the identification of CNP biomarkers using both human cohorts and an animal model (spinal nerve ligation; SNL) of neuropathic pain, and to provide clarity on the role of GTP cylcohydrolase I (GCH1) in CNP. Analysis of GCH1 and related genes and metabolites was conducted. As biomarkers, nitrite/nitrate and neopterin did not differentiate controls from CNP patients. However, significant differences were observed with biopterins, whilst correlations were observed between GCH1, nitrite/nitrate and neopterin, which were notably stronger in patients than controls. Analysis in human cohorts and in the SNL model also inferred that downregulation of GCHFR may contribute to BH4 synthesis. In order to provide clarity on the role of the GCH1 pain protective haplotype, reporter gene assays were used. This demonstrated a potential regulatory role for the GCH1 5’ SNP (rs8007267). In silico prediction of transcription factor binding sites suggested that this may be mediated by the aryl hydrocarbon nuclear translocator. The use of electrophoretic mobility shift assays showed strong specific binding with probe pertaining to the major allele. Further analysis is required to elucidate transcription factor binding, potentially facilitated by 2D-PAGE and mass spectrometry. In order to further elucidate potential CNP biomarkers, microarray analysis and qRT-PCR were performed using blood obtained from CNP patients. Data refinement led to the isolation of 27 potential CNP biomarkers, of which several cross-validated between cohorts. Microarray data, literature evidence, and correlations with previous microarrays provided evidence suggestive of a role for TIMP1. Multiple other genes, including CASP5, TLR4, TLR5, MC1R and CX3CR1, were differentially regulated in CNP. Genes surviving microarray data refinement were subsequently analysed in the dorsal horn of Sprague Dawley and Wistar Kyoto rats after SNL. Several genes, including Dpp3, Mc1r and Timp1, were similarly differentially expressed in the rodent SNL model, which suggests that these genes may be involved in the pathophysiological mechanisms of CNP, and may also function as potential translational biomarkers of CNP. This work provides multiple avenues for expansion and further investigation. Clearly, the challenges associated with biomarker discovery in CNP states are considerable, though it is hoped that this thesis provides valuable insight and the necessary foundation for future work.