Human embryonic stem cells (hESCs) have the potential to differentiate into all cell types of the three germ layers. However, various limitations hinder their use in the clinic, including possibilities of teratoma formation, xenogenic exposure through the use of Matrigel™ and feeder layers, along with poor attachment and expansion rates and inability to transport hESCs into an in vivo site. This thesis has aimed to overcome the above limitations. Electrospun nanofibrous substrates from a purely synthetic FDA approved material have been developed and investigated for the novel use in the expansion of undifferentiated hESCs. Synergistic effects between the oxygen environment and nanofibre technology were revealed which demonstrated the expansion of pluripotent hESCs in physiological normoxia (2% O2) on these substrates, with retention of differentiation capacity. However, in hyperoxia (21% O2), hESCs cultured on these substrates dictated embryoid body formation. A range of polymers (PCL, PLLA and PLGA) were tested (aligned and random conformations) where the optimal polymer (PCL) was further investigated at 2% O2 at various fibre diameters to reveal its impact on hESC clonogenicity.

This thesis sets out to test the central hypothesis that the paradigm for clinical decision support needs to shift from a technology centred paradigm to a coherent ontological-epistemological paradigm. This was achieved by formalising a coherent ontological and epistemological framework, and then applying it practically to clinical decision support. Initially the thesis reviews the need for a coherent philosophy in clinical decision support. It then goes on to describe the systematic analysis of established fundamental principles of philosophy, and the formalism of the ontological and epistemological framework. Following this the framework is applied to an analysis of clinical decision making and clinical decision support. The models derived from the analysis are then applied practically to the modelling of the management of acute renal failure patients in the intensive care setting. The results of this modelling are then combined with the decision models as the basis for the structure of a model of the decision making which controls the patient's renal replacement therapy. Finally the models representing the clinical problem and the clinical decision making process are used in the design and development of a prototype renal replacement therapy management system. The thesis concludes that a coherent ontological and epistemological framework provides clarity and insight during the analysis for and design of clinical decision support tools. The contributions of the thesis relate to the derivation and application of the framework, and the development of the renal replacement therapy management system. Thus the thesis is a foundation for future research in these two areas.

Men who have sex with men (MSM) remain disproportionately affected by HIV and sexual infections, which are acquired predominately through condomless anal sex, known as ‘barebacking’. This thesis is concerned with the experiences of HIV-negative or unknown status gay men who have recently engaged in bareback sex. Using data obtained through interpretative phenomenological analysis (IPA), this thesis makes a unique and holistic contribution to the barebacking discourse by detailing the factors that influence HIV-negative and unknown status MSM to engage in bareback sex through the analytical lens of sexual position. MSM in London were targeted via gay press, e-mail broadcasts and leafleting, and asked to take part in in-depth qualitative interviews. The interviews were digitally recorded and transcribed verbatim, and the data were managed using NVivo9™. A total of 13 MSM were interviewed; the average age of participants was 39 years (range 29-55) and all had engaged in bareback sex between 0-90 days prior to the interview. The findings are organised around a pragmatic analytical framework generated from the mens’ narratives and comprise three main themes: ‘How participants set the scene to their barebacking encounters’; ‘The act of bareback sex’ and ‘The meanings men ascribe to bareback sex’. By examining how participants locate their barebacking encounters, how bareback sex is communicated and negotiated during an encounter, and how men ascribe meaning to bareback sex, I demonstrate how participation in bareback sex is the result of a dynamic process involving different combinations of factors. These findings are presented in three separate chapters. In addition, this thesis provides new insights regarding sexual position and bareback sex. The thesis concludes with a discussion about the implications of the findings for those who work with MSM and also considers areas of possible future research.

Aims were (1) to review the literature on clinical audit; its history, methodology and role in the current National Health Service (NHS) Clinical Effectiveness initiative; and (2) to describe and discuss one national clinical audit exercise, the Gynaecology Audit Project in Scotland (GAPS) audit of the investigation and initial management of infertility. Care by general practitioners: The review of referral letters revealed that less than half of couples have basic tests of confirmation of ovulation and semen analysis performed in primary care. Conversely, up to a fifth of women with regular menses undergo unnecessary and expensive endocrine investigations. Between the two audit periods, significant, but modest, improvements occurred in the proportion of couples where the male partner was examined and had semen analysis performed and where the women's rubella status was checked. Care by gynaecologists: Between the two audit periods, significant changes in line with nine of the agreed audit criteria were demonstrated. two significant changes contrary to the agreed criteria also occurred. Patient satisfaction and experience: The patient survey indicated that 87% of women were satisfied with their care. However, over a third (39%) had never been asked to bring their partner to the clinic; 86% felt they had not been given enough help with emotional aspects of infertility; only a third had been given any written information and 78% expressed a wish for more written information. Conclusions: Clinical audit remains a cornerstone of national strategies to promote more uniform standards of high quality, evidence-based care. The GAPS Infertility Audit demonstrated the feasibility of conducting a national audit exercise encompassing patient management in both primary and secondary care settings. Modest changes in the process of care and in patients' experience were demonstrable. The modest extent of change confirms the view that audit and feedback may not be the most effective means of promoting improvements in practice. Further research is needed to determine obstacles to change and the most effective ways of overcoming them.

Peritoneal dialysis (PD) utilisation is falling in Western Countries. Concerns regarding reduced survival on PD, impact of inadequate dialysis on patient outcomes and the serious complication of encapsulating peritoneal sclerosis (EPS) may be contributing to the decline of PD. The exact incidence of EPS has been difficult to establish because of differences in design of published studies. In Scotland there was concern that the incidence of EPS was increasing, which prompted discussions about the future role and risks of PD. The aim of the MD was to establish an accurate incidence of EPS in Scotland and to examine complications and outcomes of PD patients to try to answer the question of who and for how long PD should be used in our population. Since 1999 all adult renal units in Scotland have completed a PD Audit form 6 monthly for every PD patient which gives details of PD population, source of new patients, reasons for stopping PD, causes of technique failure, details of all peritonitis episodes, adequacy test results and basic laboratory results. This prospectively collected data was linked to further demographic and laboratory data from the Scottish Renal Registry database for analysis. The analysis focussed on all incident patients commencing PD between 1st January 2000 and 31st December 2007 (n=1324), with follow-up to 30th June 2011. Our data analysis confirmed the ongoing fall in PD population in Scotland, and greater usage of APD. Peritonitis rates have remained steady at 1 episode every 19.9 months when averaged over the study period; similar to UK and Australasian results but worse than North American centres. Several risk factors for peritonitis were identified in our population including unit, CAPD compared to APD, diabetes mellitus (DM) in females, older age, hypoalbuminaemia, and lower residual renal function (RRF) at the start of PD. We established that the overall risk of EPS is low, but if PD is continued beyond 4 years the risk is substantial at 1 in 13 patients, with an exponentially increasing incidence with longer PD exposure. Survival is poor with 46.8% mortality at 1 year after diagnosis. No clear risk factors were apparent other than PD exposure. Analysis of patient survival identified several factors associated with poorer survival including increasing age, hypoalbuminaemia and RRF at the start of PD, presence of DM and multisystem primary renal diagnoses as well as having experienced peritonitis. The main causes of technique failure in our cohort include peritonitis (42.9%) and inadequate dialysis (22.1%). Predictors of technique failure include DM, lower RRF at the start of PD and being treated in more recent PD eras. Overall analysis of the PD cohort has shown that PD is a short-term treatment in Scotland with only a quarter of patients continuing PD beyond 3 years, with the remainder stopping for a transplant, technique failure or death. It is not possible to predict how long an individual patient will continue PD, but certain patients have poorer outcomes including the elderly (>70 years), those with DM and those hypoalbuminaemic at the start of PD. Therefore the actual number of patients who will continue PD long enough to be at significant risk of EPS is very small, and we believe the potential risk of EPS should not prevent patients from being offered PD in the first instance. Although some patients fare better on PD than others, we cannot state that any specific patient group should not be offered PD on the basis of our analyses particularly as we cannot show that they would have improved outcomes on haemodialysis. For the minority of patients with ongoing technique success at 4 years we suggest discussing ongoing PD, ensuring patients are informed about the EPS risk and a risk:benefit assessment of ongoing treatment should be decided on a case by case basis. It is likely that clinician attitude are driving the decline of PD, in the absence of evidence to show inferior outcomes on PD compared to HD. There would be an argument for actively increasing PD utilisation in Scotland, particularly among the elderly by expanding the assisted PD programs. Similarly, unless efforts are made to ensure adequate PD training and experience for nephrology trainees it is likely that PD will continue to decline.

Introduction: Options for imaging in acute stroke are expanding with the potential to select therapy based on imaging targets, as well as providing additional diagnostic and prognostic information. Multimodal CT has been used to image the ischemic penumbra, infarct core, and to detect leptomeningeal collateral flow although the optimum way to image these variables is not clear. Methods: In addition to a systematic literature review of imaging for leptomeningeal collaterals, Data from observational studies of acute stroke which employed multimodal CT imaging on admission and follow up was used to evaluate feasibility of acute stroke imaging with CT and MRI, Perfusion thresholds for core and ischemic penumbra, methods to quantify leptomeningeal collateral flow and sensitivity of non contrast CT for detecting infarct core pixels. Results: Advanced imaging in acute stroke and at follow up was more feasible with CT compared to MRI with the possible suggestion that imaging with MRI alone could introduce a bias regarding age and clinical severity for patients entered into clinical studies Heterogeneity in grading and detecting collateral flow was found in the literature providing an opportunity to devise a novel assessment method. Well developed collaterals were associated with imaging and clinical markers for good outcome as well as some potential biomarkers including atrial fibrillation and blood fibrinogen level. Relative cerebral blood flow and delay time were found to be the best predictors on infarct core and ischemic penumbra after derivation of optimum perfusion thresholds and subsequent validation in independent patient groups. Pixel based comparison of infarct core on CT perfusion and non contrast CT highlighted the lack of sensitivity of CT for detecting infarct core based on Hounsfield unit value alone. Conclusion: Multimodal CT for acute stroke assessment offers the potential for measuring infarct core, ischemic penumbra and leptomeningeal collateral flow status rapidly according to novel grading scales and thresholds and provides information on tissue viability which cannot be detected on non-contrast CT. Further evaluation on the impact additional imaging should have in clinical practice is needed.

Rheumatoid Arthritis (RA) is a chronic, progressive, multisystem inflammatory disorder for which there is, at present, no cure. It affects up to 1% of the population resulting in chronic pain, disability and, through loss of function, may lead to loss of employment. It is associated with major co-morbidities that account for premature mortality. There is now extensive published research that suggests early treatment with disease modifying drugs can retard joint damage and improve outcome. In a proportion, drug- free remission is possible. However, there remain both individuals with persistently active disease despite standard drug treatments and those with longstanding disease not exposed to effective early treatment that remain relatively unresponsive to therapy. There is a growing literature that epigenetic modifications may underpin, or at least accelerate the development of many autoimmune disorders. These include alterations in DNA methylation patterns, histone tail modifications, post-translational mRNA regulation by microRNA and combinations therein. Having established the human genome project and underlying human DNA sequence, the recognition of dynamic epigenetic regulation of the genome has added further complexity. Few data however are currently available in ‘real-world’ cohorts of patients. In order to explore the hypothesis that specific epigenetic changes may underpin differences in response to therapy, I first examined the characteristics of a cohort of fifty RA patients with longstanding and active disease (DAS28 >3.2) despite receipt of standard therapies (disease modifying drugs (DMARD) and biologic therapies. This included a detailed examination of clinical characteristics, immune profile, inflammatory markers and burden of co-morbid complications such as vascular disease and depression. Outcomes such as disability, quality of life assessments and fatigue were evaluated by means of previously validated questionnaires. These groups were assessed at baseline, three months and six months. I then measured one of the many epigenetic marks, namely microRNA, of this cohort. We analyzed the accessible profile of peripheral RA CD14+ cell microRNAs in treatment resistant RA patients, in healthy controls, DMARD inadequate responders and DMARD good responders in order to determine the presence of a microRNA profile indicative of biologic resistance. An analysis of the serum cytokine profile of the biologic resistant and DMARD resistant groups was also performed. Finally, to extend the analysis beyond conventional clinical and novel molecular biomarkers the influence of additional patient factors such as coping and illness perception were evaluated by questionnaire to determine subjective disease severity in discrete patient groups. Active inflammatory disease was present as judged by the DAS28 score and there was some improvements seen over the six-month assessment period reflecting treatment changes in all groups. Substantial disability and impaired quality of life was found particularly in the therapeutic resistant group but also in those with inadequate response to DMARD, and remained relatively unresponsive to treatment escalation. Clinical variables, deprivation, quality of life and fatigue were strongly correlated with mood suggesting close interactions and resultant increase in disease activity as measured by the DAS28. Multiple cardiovascular risk factors were determined and, having applied cardiovascular risk scoring systems, unmet treatment of modifiable risk was demonstrated. Exploratory analysis of candidate microRNA -34a, -27b and -125a showed no correlation with clinical or biochemical variables other than swollen joint counts but differential expression between study groups. Exploratory microarray profiling between the four study groups demonstrated a number of differentially regulated microRNA. Of these, a unique microRNA profile of the biologic resistant group was found. MicroRNA-423 and -1275 showed higher expression in the biologic resistant group and fell in parallel with the DAS28 reduction between study visits raising their potential utility as biomarkers. MicroRNA-3178 showed higher relative expression in the biologic resistant group. Cytokine profiles demonstrated significant differences vs healthy controls but biologic resistant, DMARD resistant and DMARD good responder groups were less distinct and individual cytokines failed to discriminate in these study groups. Cytokine profiling did not correlate with observed clinical variables, inflammatory markers nor central processes such as mood or fatigue. Finally, those coping strategies favoured were adaptive and problem based. These were unaffected by the high prevalence of mood disturbance. Conversely, illness perception was influenced by mood and both affected subjective disease assessments. The strong influence of mood and fatigue raise the hypothesis that blunted treatment response may be partially driven by these variables. Ultimately we seek to explain, identify and target those patients with aggressive disease as early intervention may prevent established disease and it's accompanying co-morbid conditions. Undoubtedly, a personalised assessment of disease variables and co-morbid conditions is necessary where treatment response is being evaluated. In such a way, significant cardiovascular morbidity and mortality may be prevented. The question of true biologic resistance remains open. Undoubtedly residual inflammation exists in longstanding RA but significant ‘disease activity’ may be explained at least in part by those subjective clinical variables influenced by both external and internal factors. The identification of a ‘biologic resistant’ microRNA profile could act both as a biomarker of treatment response in longstanding disease, superior to the DAS28 scoring system and, through target identification, better understanding of the regulation of the molecular pathways of inflammation operating in such patients. In this way novel pathways of treatment resistance may be exposed and novel treatment targets revealed. However, mood and thus illness perception also contribute to resistance to therapy and should be sought, characterized, and directly addressed to add to the global improvements in outcome that we seek in a holistic model of care in the rheumatic diseases.

The stability study of drugs of forensic interest in human post-mortem blood is an important forensic study, because in some cases, a requirement for the laboratory to undertake a full drug screening is after a few months of storage due to a need for new evidence. Therefore, it is necessary and important to know if drugs are stable over a period of time under different conditions to enable a solid interpretation to be made from any results. Some studies have been published on the stability of drugs at different temperatures but none had covered the whole set of drugs that has been studied in this thesis. The periods of study that have been covered by other studies varied from a few days to a maximum of 70 weeks, but again not all drugs have been covered. The drugs studies in this thesis are two sets of drugs, benzodiazepines and tricyclic antidepressants, their stability being determined over twelve months and at three different temperatures 25,5 and -20°C. In this thesis, blood was 'spiked' with eight drugs, Diazepam, Temazepam, Triazolam, Desmethyldiazepam, Amitriptyline, Nortriptyline, Imipramine and Chlorpromazine. The samples were stored with blanks at different temperatures for different storage times. Each month a number of samples were removed from storage and analysed to test the effect of storage time and temperature on drug concentration. Different solid phase and liquid-liquid extraction methods were tested for the determination of benzodiazepines. Liquid-liquid extraction methods for - 2 - the determination of Diazepam, Temazepam, desmethyldiazepam and Triazolam proved after study to be tedious and time-consuming. A method based on solid phase extraction was used to determine the four benzodiazepine drugs. The extraction method gave good recoveries and was highly efficient. The method of analysis used for the determination of stability of benzodiazepine drugs was the high performance liquid chromatography (HPLC) method. Tricyclic antidepressant drugs are the other drugs studied for their stability in blood. Different solid phase extraction methods were used for the determination of drugs in post-mortem blood but gave poor recoveries. The best method of extraction used was a liquid-liquid extraction method which yielded high recoveries and proved to be quick. The method of analysis used for the determination of tricyclic antidepressants for the purpose of stability of the study was gas chromatography (GC). At a recognised toxic level for each drug under study a reasonable amount of the drug was found to be detectable after one year at storage regardless of the storage temperature or media. The decrease rate of each drug concentration with time at the three storage conditions (25, 5 and -200e) was obtained.

A 6-month intervention study with a quasi-experimental design and a 3-month follow-up was conducted to evaluate the effectiveness of an Internet-based, step-wise, tailored-feedback intervention promoting four key components of the Mediterranean diet (vegetables, fruits, nuts and seeds, legumes and ratio of monounsaturated to saturated fat). Fifty-three (intervention group) and nineteen (control group) healthy females, aged 25-55 years, were recruited from the Universities of Glasgow and Glasgow Caledonian, Scotland, respectively. Participants in the intervention group received tailored dietary and psychosocial feedback and Internet nutrition education over a 6-month period, while participants in the control group were provided with minimally-tailored dietary feedback and general healthy-eating brochures. Internet education was provided via an innovative Mediterranean Eating website. Between group comparisons carried out on an "intention-to-treat" basis, providing the strongest evidence of the effect of the intervention, showed that participants in the intervention group had made more favourable changes to their fruit, nut and seed intake over the 6-month intervention, as well as increased their vegetable intake over the 9-month trial. Over both the 6-month intervention and 9-month trial, participants in the intervention group had more favourable levels of HDL-cholesterol and ration of total:HDL-cholesterol, a higher proportion progressed through the stages of behavioural change regarding legumes and olive intake and self-efficacy skills were generally increased, compared with the control group. Participants in the control group however, showed more favourable urinary electrolyte levels throughout the study. Within group comparisons showed that at 6 months, participants in the intervention group had significantly increased their intake of vegetables, fruits, legumes, as well as the MUFA:SFA ratio in their diet, had increased their mean total MDS and had significantly increased plasma HDL-cholesterol levels and a reduced ratio of total:HDL-cholesterol, as well as higher nutrition knowledge scores compared with baseline. In addition, a higher percentage of participants in this group were in the action and maintenance stage of behavioural change for vegetables, legumes and olive oil consumption, as well as generally showing more favourable attitudes and self-efficacy skills towards consumption of most of the food components promoted by the study at 6 months. These changes were generally maintained at 9 months, when additional decreases in blood pressure and an increase in total cholesterol, compared with baseline, were reported. Participants in the control group increased their intake of legumes, as well as their mean total MDS, and had significantly reduced urinary sodium levels at 6 months, compared with baseline. In addition, a higher efficacy skills generally decreased, compared with baseline. These changes were not maintained at 9 months, but at this time point participants in this group had a higher nutrition knowledge score, compared with baseline.

Healthcare associated infection confers a significant burden of morbidity and mortality to renal patients and to renal dialysis patients in particular. Sepsis is second only to cardiovascular disease as the leading documented cause of death in patients requiring renal replacement therapy. Gram positive bacteraemia is common in the renal replacement therapy population and is highly associated with indwelling haemodialysis catheter use. Optimal prevention and management of bacteraemia in this setting has not been fully determined and requires a multidisciplinary and multifaceted approach. Each of the studies in this thesis investigates an aspect of healthcare associated infection in nephrology within the theme of exploring clinical problems arising from the development of antibiotic resistance or antibiotic associated infections in renal patients. Initially we examined risk factors and outcomes of acute kidney injury requiring renal replacement therapy in a tertiary renal unit and critical care population prior to and subsequent to a change in antimicrobial guidelines in response to an outbreak of Clostridium difficile associated disease. We performed this study to address concerns that the increase in the empiric use of gentamicin may have led to an increased incidence of acute kidney injury and a greater requirement for emergency renal replacement therapy. Secondly we explored the clinical implications of gram positive infection in a renal unit population by performing a retrospective review of Staphylococcus aureus and coagulase negative staphylococcal bacteraemia over a 2 year period with particular attention to admission rates, vascular access intervention, antibiotic resistance, metastatic infection and mortality. Thirdly we have analysed S. aureus toxin genes and assessed the epidemiology of S. aureus colonisation and infection to improve our understanding of the virulence of S. aureus in different patient populations including a large haemodialysis unit in Glasgow. Finally we undertook a prospective double blind randomised controlled trial of probiotic milk drink and placebo in renal unit inpatients commencing antibiotic therapy to assess if a probiotic was effective in the prevention of antibiotic associated diarrhoea and Clostridium difficile associated diarrhoea. We performed this study as patients with chronic kidney disease are at increased risk of infection and have a significant antibiotic burden, which can lead to antimicrobial resistance, antibiotic associated diarrhoea and pseudomembranous colitis due to Clostridium difficile infection. The study of healthcare associated infection is an evolving field and involves complex interactions between colonisation and infection. There is increasing emphasis on prevention of infection and minimising complications and side effects associated with standard antimicrobials. The rising incidence of multiresistant bacterial infections is likely to result in increasing focus on preventive bundles of care and alternatives to antimicrobial therapy such as the use of probiotics. The findings of this thesis contribute to the goal of prevention of antibiotic resistance and multiresistant infections in renal patients although further research is required.