Renal disease is a major global public health issue that affects 10% of the general population with premature morbidity and mortality related to cardiovascular disease and infection. Interstitial fibrosis is a common hallmark of progressive kidney dysfunction. There remains a stubborn discrepancy in identifying which patients suffer adverse events because of their disease or resulting treatment. Investigation in patient genome variation may explain this discrepancy. Caveolin-1 is the essential structural protein for caveolae that are ubiquitously distributed in fibroblasts, endothelial and epithelial cells. They act as molecular chaperones for transcellular signaling such as degradation of the activated TGFβ-1 receptor. In this thesis, caveolin-1 single nucleotide polymorphism rs4730751 CC genotype is shown to be associated with a better outcome in renal patients for arterial stiffness, and reduced mortality from cardiovascular disease, infection, malignancy in ANCA associated vasculitis. By inducing renal models of fibrosis in caveolin-1 knockout mice, deletion of caveolin-1 leads to increased fibrosis. In conclusion, this polymorphism could be used as a marker of disease risk either in isolation or as part of a clinical risk score to counsel patients on the likely prognosis of their condition. Manipulation of caveolin-1 expression may be a therapeutic strategy in reducing renal fibrosis.

Hepatitis C virus (HCV) is a major human pathogen and the leading cause of cirrhosis and liver cancer worldwide. HCV entry is clathrin- and pH-dependent, and requires CD81, Scavenger receptor BI (SR-BI), and the tight junction (TJ) proteins Claudin-1 and Occludin. Primary HCV strains cannot be efficiently cultured in vitro, making the evaluation of potential antiviral compounds in a biologically relevant context extremely difficult. Despite being suitable for high-throughput screening, most cell-based reporter assays rely on the secretion of serine alkaline phosphatase and thus do not allow the selection of HCV infected cells, or the screening of patient samples to identify cell culture infectious viral strains. We aimed to develop a cell-based reporter assay, which utilizes the viral NS3/4A protease to cleave and activate a fluorescent reporter protein constitutively expressed in Huh-7.5 hepatoma cells. HCV tropism is restricted to the liver, where hepatocytes are polarized and form TJ, which are indispensable for normal liver functionality. We demonstrate that in confluent cells, SR-BI and Claudin-1 expression is increased and that HCV entry is enhanced when cellular contact is established. Furthermore, cell junction formation and SR-BI overexpression, respectively, accelerated virus entry, suggesting a key role for SR-BI in HCV internalization. The mechanisms underlying HCV-associated hepatic injury are poorly understood, however, it is thought that HCV may disrupt TJ integrity, thus compromising hepatocyte polarity and function. We demonstrate that the HCV structural proteins modulate the expression and localization of TJ proteins, leading to their redistribution to cytoplasmic vesicles with possible consequences for TJ integrity in vivo.

Individuals or groups will form impressions of another based upon a series of traits, which may be relied upon when forming behaviour pattern towards others (Asch, 1946; Crocker and Major, 1989; Pinel, 1999; Albon, 2002; Corrigan and Wassel, 2008). These traits will depict the reception individuals receive within healthcare and may depend upon learnt and inherited ‘perceived’ ideals affecting the working and personal relationships experienced by positively diagnosed healthcare workers, predisposing stigma responses to others (Asch, 1946). A mixed method study investigating the potential existence of stigmatising values from student nurses towards positively diagnosed healthcare workers with Pulmonary Tuberculosis (PTB), Human Immunodeficiency Virus (HIV), Methicillin-resistant Staphylococcus Aureus (MRSA), Hepatitis C and Diabetes type 2, was undertaken. Analysis provided exploration of the stigmatising attitudes and values of 482 student nurses undertaking an education programme. Interpretation of the findings explored the participants views at course commencement, midpoint and completion considering variables of education (theoretical and clinical), personal and professional influences. Findings indicated that stigmatising values and attitudes return to those identified precourse and underpin the need for implementation of a change to nurse education within this area. The development of a longitudinal education model for healthcare workers considering disease processes and influencing factors psychologically, socially and physically, will provide opportunities to reduce the existence of stigmatisation for positively diagnosed healthcare workers.

In most patients with rheumatoid arthritis (RA), citrullinated autoantigens are targeted by autoantibodies (ACPA). However, the process leading to protein citrullination by peptidylarginine deiminases (PADs) in the joint remains unclear. In this thesis, I tested the hypothesis that generation of neutrophil extracellular traps (NETosis), can contribute to release of enzymatically active PADs and citrullinated autoantigens in inflamed joints. I have shown that in vitro induced NETosis leads to release of citrullinated proteins and enzymatically active PADs both attached to NETs and free in the supernatant. In the SF from RA patients DNA levels correlated with neutrophil concentrations, and DNA levels and PAD activity were found to be increased compared with OA patients. Finally, I demonstrated the antigenicity of in vitro generated NETs and identified citrullinated histone H3 as a NET-component recognised by ACPA and RA sera. Based on the findings in this thesis release of active PADs into SF by neutrophil cell death is a plausible explanation for the generation of citrullinated extracellular autoantigens. In ACPA positive RA patients the continuous production of these autoantigens combined with pre-existing ACPA may result in the formation of immune complexes and perpetuation of the inflammatory response.

Testing can be a substantial driver of health care costs. Increase in test use over recent decades has occurred despite disappointing results from test accuracy evaluations. Poor quality and reporting of primary test accuracy studies and difficulties with understanding and application of test accuracy information are purported to be important contributors to this observed evidence ‘gap’. The objectives of this thesis were to: * Systematically review evidence concerning the understanding and application of test accuracy metrics. * Undertake primary research building on the review of understanding and application. * Assess whether the contribution of test accuracy reviews to the test accuracy evidence base is compromised by deficiencies in their contextual fit, or of included primary studies. Existing research concerned with understanding and application of test accuracy information is not driven by the needs of decision makers. Contrary to the prevailing view in the literature, findings of original research from this thesis demonstrate that probability revision is not a feature of diagnostic decision making. Choice of test accuracy metric however was shown to have a profound influence on diagnostic decision making. Deficiencies in question formulation and contextualisation of test accuracy reviews are undermining their contribution to the test accuracy evidence base.

Motivational based approaches have been shown to be effective in reducing problematic behaviours in the general population. The present study reviewed 31 studies that aimed to reduce a number of problematic behaviours amongst individuals with severe mental health problems. Evidence supporting the efficacy of motivational based approaches in increasing adherence to medication was inconsistent. In the minority of studies where an increase in medication adherence was reported, the duration of effect appeared to decline over time. The evidence from the studies that aimed to reduce substance use was more consistent. A number of other positive outcomes were reported reductions in: dependence, general functioning and mental health symptomatology. The empirical paper presents a quantitative study that aimed to look at the reasons for cannabis use amongst individuals with and without severe mental health problems. It was found that the motivations for using cannabis did not differ between the two groups; both using cannabis to cope with negative affect, for pleasure and for social reasons. However, individuals with severe mental health problems differed in that they expected cannabis to be more ‘socially and sexually facilitative’. Irrespective of mental health status, participants who used cannabis more problematically endorsed more coping and pleasure motives.

Childhood obesity is a major global concern but there is little evidence for effective childhood obesity prevention strategies. People of South Asian origin are a specific target group for obesity prevention as they are vulnerable to cardiovascular health effects of obesity. This thesis explores the contextual influences on the development of childhood obesity by presenting two studies; a quantitative analysis of routine datasets explores the association between the school physical activity environment and obesity in children, and a qualitative study explores the beliefs of South Asian community members and other stakeholders about the causes of childhood obesity and potential interventions to prevent it. A process of development of a childhood obesity prevention programme tailored to South Asian communities, guided by the MRC framework for complex interventions is described. The contextual information from the stakeholder focus groups is central to this process. Finally, this thesis examines the potential consequences of obesity in South Asian children by exploring its association with body image. Understanding the psychosocial consequences of obesity in target communities will enable future interventions to be appropriately designed. The findings of this thesis highlight the importance of understanding the cultural context with respect to childhood obesity causes, consequences and intervention.

Women’s views of antenatal screening have been widely researched, but men’s remain under-explored. The original contribution of this research was to conduct a mixed-methods study about men’s experiences specifically. In-depth interviews were firstly conducted with twelve men. Six women were interviewed about their views on men’s involvement. A grounded theory was developed, which was that men began developing a prenatal paternal identity and a schema of their child. These conceptualisations were reinforced or distorted by screening, causing their ideas and feelings about their growing family to be in a state of flux. To explore this theory with a more diverse group, a questionnaire was designed, pretested (n=30), piloted (n=53) and administered to 200 men. Exploratory factor analysis showed prenatal paternal identity and child-schema consisted of ‘bonding and closeness’, ‘genetic relationship’ and ‘imagined interactions’. Regression analyses showed investing time in screening, seeing more ultrasound scans, and making screening decisions, predicted higher scores on ‘bonding and closeness’. Investing time and being younger predicted higher scores on ‘genetic relationship’. Investing time, feeling fetal movements, being of a higher socioeconomic status and being younger predicted higher scores on ‘imagined interactions’. Longitudinal research is now required to identify implications for men, women and children.

Events in utero have been linked with diseases throughout life, however there is a lack of consensus regarding the ability of neonatal tests to predict these outcomes. Systematic reviews and meta-analyses were performed, assessing umbilical cord pH and base excess at birth, standards of low birth weight, and the Apgar score, including a total of 218 papers and 26704980 individuals. The prognostic association and predictive accuracy of these tests for adverse outcomes, including neonatal mortality and morbidity, childhood morbidity including cerebral palsy, and adult outcomes, were determined. A decision-analytic model based analysis assessed the cost-effectiveness of varying the umbilical cord pH threshold, and treatment with neonatal hypothermia. This thesis determined that all of the tests examined had a strong association with neonatal mortality, and a significant but smaller association with neonatal morbidity and childhood cerebral palsy. In general, where the association was strong, tests had a high specificity and positive likelihood ratio for adverse outcome, but poor sensitivity and negative likelihood ratio, indicating that negative tests do not reduce the risk. The cost effectiveness analysis showed that the threshold of pH used in current practice to recommend neonatal hypothermia is more effective and less costly than a higher threshold.

Measles is a leading cause of mortality in infants in countries with suboptimal vaccination coverage. This disease is caused by a negative-strand RNA virus, measles virus (MeV). Wild-type strains of the virus use two cellular receptors to invade cells and establish infection: the signalling lymphocyte activation molecule f1 (SLAMF1), which is present on certain immune cells, and nectin-4, which is expressed in the lung epithelium. During infection, MeV can spread through the release of virions or by inducing cell-cell fusion. The aim of this thesis is to determine the molecular mechanism underlying viral entry and exit. Herein, I observed that, upon attachment to SLAMF1+ cells, MeV particles induce extensive but transient membrane blebbing and cytoskeleton contraction. MeV entry occurred simultaneously with fluid-uptake and was sensitive to inhibitors of macropinocytosis and cytoskeleton dynamics. In contrast, the cortical actin network restricted the early stages of MeV-induced cell-cell fusion, in RhoGTPases, ezrin and moesin dependent manner. By resolving the proteome of infected cells, conserved phosphorylated residues in the viral haemagglutinin were also shown to impact on dimerization and cell-cell fusion. These results suggest the manipulation of several cellular components and pathways during entry and exit of MeV.