The problem: Around 300 million people worldwide have asthma and prevalence is increasing. Support for optimal self-management can be effective in improving a range of outcomes and is cost effective, but is underutilised as a treatment strategy. Supporting optimum self-management using digital technology shows promise, but how best to do this is not clear. Aim: The purpose of this project was to explore the potential role of a digital intervention in promoting optimum self-management in adults with asthma. Methods: Following the MRC Guidance on the Development and Evaluation of Complex Interventions which advocates using theory, evidence, user testing and appropriate modelling and piloting, this project had 3 phases. Phase 1: Examination of the literature to inform phases 2 and 3, using systematic review methods and focussed literature searching. Phase 2: Developing the Living Well with Asthma website. A prototype (paper-based) version of the website was developed iteratively with input from a multidisciplinary expert panel, empirical evidence from the literature (from phase 1), and potential end users via focus groups (adults with asthma and practice nurses). Implementation and behaviour change theories informed this process. The paper-based designs were converted to the website through an iterative user centred process (think aloud studies with adults with asthma). Participants considered contents, layout, and navigation. Development was agile using feedback from the think aloud sessions immediately to inform design and subsequent think aloud sessions. Phase 3: A pilot randomised controlled trial over 12 weeks to evaluate the feasibility of a Phase 3 trial of Living Well with Asthma to support self-management. Primary outcomes were 1) recruitment & retention; 2) website use; 3) Asthma Control Questionnaire (ACQ) score change from baseline; 4) Mini Asthma Quality of Life (AQLQ) score change from baseline. Secondary outcomes were patient activation, adherence, lung function, fractional exhaled nitric oxide (FeNO), generic quality of life measure (EQ-5D), medication use, prescribing and health services contacts. Results: Phase1: Demonstrated that while digital interventions show promise, with some evidence of effectiveness in certain outcomes, participants were poorly characterised, telling us little about the reach of these interventions. The interventions themselves were poorly described making drawing definitive conclusions about what worked and what did not impossible. Phase 2: The literature indicated that important aspects to cover in any self-management intervention (digital or not) included: asthma action plans, regular health professional review, trigger avoidance, psychological functioning, self-monitoring, inhaler technique, and goal setting. The website asked users to aim to be symptom free. Key behaviours targeted to achieve this include: optimising medication use (including inhaler technique); attending primary care asthma reviews; using asthma action plans; increasing physical activity levels; and stopping smoking. The website had 11 sections, plus email reminders, which promoted these behaviours. Feedback during think aloud studies was mainly positive with most changes focussing on clarification of language, order of pages and usability issues mainly relating to navigation difficulties. Phase 3: To achieve our recruitment target 5383 potential participants were invited, leading to 51 participants randomised (25 to intervention group). Age range 16-78 years; 75% female; 28% from most deprived quintile. Nineteen (76%) of the intervention group used the website for an average of 23 minutes. Non-significant improvements in favour of the intervention group observed in the ACQ score (-0.36; 95% confidence interval: -0.96, 0.23; p=0.225), and mini-AQLQ scores (0.38; -0.13, 0.89; p=0.136). A significant improvement was observed in the activity limitation domain of the mini-AQLQ (0.60; 0.05 to 1.15; p = 0.034). Secondary outcomes showed increased patient activation and reduced reliance on reliever medication. There was no significant difference in the remaining secondary outcomes. There were no adverse events. Conclusion: Living Well with Asthma has been shown to be acceptable to potential end users, and has potential for effectiveness. This intervention merits further development, and subsequent evaluation in a Phase III full scale RCT.

Mesenchymal stem cells (MSC) have been used clinically to treat a range of conditions, and have been shown to enhance skin wound healing in vivo. There is evidence suggesting that the beneficial effects of MSC transplantation may be mediated more through their paracrine activity than through their differentiation into mature cell types. This thesis has sought to examine the effects of MSC secreted factors on dermal fibroblasts, keratinocytes, and endothelial cells, using an in vitro wound healing model. The results presented demonstrate that mesenchymal stem cell-conditioned culture medium (MSC-CM) enhances wound closure by dermal fibroblasts and keratinocytes through increased skin cell migration. MSC-CM was similarly shown to enhance wound closure by endothelial cells. The contents of MSC-CM were analysed by immunological techniques and mass spectrometry to reveal targets for further investigation. These included cytokines, notably interleukin-6 (IL-6), interleukin-8 (IL-8) and transforming growth factor-beta one (TGF-β1); and extracellular matrix (ECM) components, notably collagen type I, fibronectin and decorin. Examining these factors individually or in combination with each other or MSC-CM demonstrated differential and interdependent effects. Fibronectin was generally stimulatory to wound closure, whilst decorin was inhibitory. The cytokines studied had variable effects, e.g. TGF-β1 was stimulatory to fibroblast migration in combination with collagen type I, elicited no effect in combination with fibronectin, and was inhibitory to keratinocyte migration in both conditions. This study has thereby progressed our understanding of how MSC secreted factors influence skin wound healing and suggests that further investigation of MSC activity is needed in order to inform the eventual clinical use of MSC to treat severe or chronic skin wounds. With further work, MSC-related therapy, using MSC transplantation or the delivery of MSC secreted products, might provide a preferable alternative to current treatment strategies for these wounds, either in terms of patient benefit or clinical cost reduction.

Clinicians are ‘medical knowledge experts’ who can use their training, their experience, and evidence based medicine to diagnose and determine healthcare options available to patients, but patients are ‘personal preference experts’. Shared decision-making (SDM) depends on the combination of the different clinician and patient expertise, but to date efforts to increase SDM have largely not addressed how to enable patients to contribute their expertise to the process. The work in this thesis describes the development and pilot-testing of a theory-based intervention, which aims to prepare patients to participate in SDM. Development and pilot-testing was guided by the MRC’s framework for developing complex interventions. A systematic review revealed patient-reported barriers and facilitators to participating in SDM (key barriers included knowledge and perceived power imbalance) and a literature review found that existing interventions do not sufficiently address patient-reported barriers; thus there was scope to develop on more comprehensive theory-based intervention. The Behaviour Change Wheel guide was used to develop a theory-based intervention: an 8-page booklet entitled ‘Your Health, Your Choice’. Pre-testing with lay users, clinicians and organisational representatives revealed positive responses to the booklet’s key messages and design. Preliminary pilot-testing in a breast care setting showed evidence of high reach, dose and usage, and potential for the intervention to change patients attitudes towards the patient role in consultations (i.e. should patients be involved in SDM). However, the booklet had less impact on patients’ perceptions of whether clinicians want patients to become more involved (i.e. would patients be able to become involved.) Overall, preliminary findings suggest that the intervention could be a useful tool for preparing patients for SDM and for changing patients’ attitudes towards patient involvement. However, preparation for SDM must be followed by enablement by willing and skilled clinicians, and delivered within a supportive organisation.

Effects of subclinical hypothyroidism on bone mineral density and cardiometabolic-risk: genetic and immune influences Subclinical hypothyroidism (SH) affects 3-8% of the population and is associated with hypertension, dyslipidaemia and altered bone mineral density (BMD). Metabolic anomalies in thyroid disease have been attributed to thyroid hormone variation but mice deficient in thyrotropin (TSH) receptor (TSHR) have low BMD, despite normal thyroid hormones, suggesting that TSH/TSHR function in bone is important. SH has several aetiologies including thyroid autoimmunity characterised by thyroid peroxidase autoantibodies (TPO-Ab), inactivating TSHR mutations (TSHR-M) and FOXE1 polyalanine tract length (FOXE1-PTL) polymorphisms. I hypothesise differential bone effects in SH relating to these causes. Similarly the effects of SH on metabolic outcomes is unclear and may also depend on SH aetiology. Principle aims - 1. To determine the prevalence of heterozygous TSHR-Ms in SH. 2. To evaluate body composition and metabolic parameters according to i) TSH, T4 and T3; ii) TPO-Ab; iii) TSHR-M or polymorphism; iv) FOXE1-PTL polymorphism. 156 women and 52 men, mean age 51 with primary untreated SH were recruited. Blood samples were obtained for biochemistry (thyroid & lipid profiles, TPO-Ab, HOMA-IR) and blood pressure (BP) and anthropometric data collected. TSHR and FOXE1 were genotyped. Dual-energy X-ray absorptiometry generated Z-scores. Stepwise multivariate regression analyses were performed. Half of the cohort had TPO-Ab; 6% had TSHR-Ms (essentially TPO-Ab negative) and 60% expressed FOXE114/14-PTL. TSH and TPO-Ab associated negatively with BMD-Z at lumbar spine but not hip whereas free-T3 and male gender associated negatively at both sites. TSHR-M status did not influence BMD-Z despite lower free-T3 relative to TSH. Free-T3 associated positively with BP and HOMA-IR. FOXE1-PTL14/14 associated positively with free-T3 and negatively with BP. TSH, TSHR-M and TPO-Ab status showed no metabolic associations but unexpectedly TSH showed a positive association with T3. SH is considered homogenous but my results illustrate its heterogeneity and highlight the need for studies accounting for aetiology in order to optimise clinical management.

In the UK alone 28 000 sudden cardiac deaths a year are found to be unrelated to structural heart problems, likely having ion channel malfunction and arrhythmia as an underlying cause. With the discovery of human induced pluripotent stem cells (hiPSC), a new field has emerged, allowing the creation of personalised cellular models of disease that can be studied in vitro. In this study we aim to functionally characterize and validate an in vitro model of Catecholaminergic Polymorphic Ventricular Tachycardia type 1 (CPVT1), an adrenergically elicited arrhythmic condition caused by a mutation in the cardiac ryanodine receptor (S2246L). The cardiac ryanodine receptor (RyR2) is an intracellular ion channel that releases Ca2+ stores essential to cardiac contraction during EC-coupling. Inherited defects in the RyR2 can result in an intracellular calcium leak during diastole that may lead to arrhythmia. The most frequent arrhythmic events at a cellular level in CPVT cardiomyocytes (CM) are delayed afterdepolarizations (DADs) although early afterdepolarizations (EADs) and triggered activity (TA) may also occur. With the need of obtaining a human cell based disease model to establish in vitro valid assays for CPVT study and drug screening, a standardized quantitative approach is ideal. Nonetheless, because the quantitative definition of these arrhythmic endpoints is subjective by nature, most studies using hiPSC CPVT derived CM have assessed these cells only from a qualitative point of view. Using whole cell patch clamp in the ruptured membrane modality and Ca2+ fluorescence imaging, this study aimed to provide a quantitative characterization of arrhythmic endpoints in a CPVT hiPSC model. Pharmacological rescue was carried out using flecainide, propafenone, dantrolene (10μM for all drugs) and propranolol (1μM) in the presence of isoprenaline. Our results show that the average response of hiPSC CPVT CM to recapitulate disease is heterogeneous and therefore not a consistent in vitro model of disease.

Abnormal glucose metabolism, including new-onset diabetes after transplantation (NODAT), is a common complication following kidney transplantation. Better understanding of the causes, associations, prediction and outcomes of NODAT in the modern era of kidney transplantation is essential. Our central hypothesis was that early NODAT is distinct from type 2 diabetes mellitus (T2DM), and is due to factors unique to the transplant setting, of which the predominant factor is the use of specific immunosuppressive agents (calcineurin inhibitors-CNIs), and that traditional risk factors for T2DM are the more significant factors late after transplantation. In a series of observational studies, we found that recipient age, body mass index and baseline plasma glucose levels were associated with the development of NODAT, both early and late after transplantation. Exposure to tacrolimus and being transplanted in an older era were associated with early NODAT development, but had no effect on late NODAT. There was increasing insulin resistance but no compensatory increase in insulin secretion in patients developing NODAT, suggesting an effect of CNIs. In an observational study using paired oral glucose tolerance tests, there was worsening of glucose tolerance late after transplantation. Metabolic syndrome was a risk factor for this deterioration. Finally, in an epidemiological study, we show that immunosuppression regimens in Cardiff have evolved, with the introduction of induction therapy and tacrolimus as the CNI of choice. Blood tacrolimus levels, corticosteroid exposure and acute rejection rates were lower in a recent era of transplantation, as was the incidence of NODAT. NODAT developing within the first year, higher systolic blood pressure and higher serum creatinine level were all associated with increased mortality. In conclusion, traditional T2DM risk factors are important in causing both early and late NODAT, with a strong influence from immunosuppressive agents early after transplantation. NODAT and other cardiovascular risk factors were associated with mortality. Therefore, less diabetogenic immunosuppressive regimes and interventions to reduce hyperglycaemia may not improve mortality unless other cardiovascular factors are also managed simultaneously.

Background: Medication adherence has shown to be problematic for many renal transplant recipients. While factors promoting or inhibiting medication adherence have been extensively researched, little is known about the processes leading to this behaviour as perceived by kidney transplant recipients. Also, no research on the perspectives of German kidney transplant recipients has yet been carried out. Research Question: The question underpinning this research was: “How do German renal transplant recipients perceive the processes leading to medication adherence or non-adherence?” Methods: Following informed consent, telephone interviews with 17 German renal transplant recipients were conducted, transcribed verbatim, and analysed according to the tenets of constructive Grounded Theory, until theoretical saturation was reached. The research has been approved by the research ethics committees of the School of Healthcare Sciences and the German Society of Nursing Science. Results: This research established the theory of medication-taking as a symbol of living with a chronic condition. This theory is underpinned by two categories: in the category reflecting on one’s own position, the participants discussed their role regarding the intake of medication, which was perceived very ambivalently and as just one component of self-management following transplantation. In the category experiencing facilitators and challenges, participants reported factors supporting or impeding medication-taking. Crucially, these are perceived very individually: what one finds helpful or challenging may be perceived in a fundamentally different way by someone else. Conclusions: This research has similar findings to other research in this field, such as the fact that renal transplantation is not a cure for a chronic condition. However, in contrast to other research, it has found a strong connection between medication-taking and participants’ self-reflection of being chronically ill. In this regard, it has emphasised the need for individualised care, preferably in the form of a team approach that includes patients and families as well as the different healthcare professions.

An influx of leucocytes, particularly macrophages, into the glomerulus and cortical interstitium, is characteristic of most forms of progressive renal disease. Previous work in our lab has demonstrated that leucocyte adherence to primary cultures of human renal fibroblasts stimulated ICAM-1 induction (1). Induction was initiated as a result of the ICAM-1/ β2 integrin interaction, and was mimicked by crosslinking ICAM-1 with specific antibodies. The aim of my research was to examine the effect of leucocyte/fibroblasts interaction on ICAM-1 and HAS2 expression, and to investigate the mechanisms involved. I have shown induction of HAS 2 mRNA, the principal synthase involved in generation of the extracellular matrix polysaccharide hyaluronan (HA), in parallel with ICAM-1 in the fibroblasts. There were both cell-contact and soluble factor-mediated components to this activation. In addition there was an exponential rise in this induction when monocytes were activated. This was mediated by ICAM-1 on the fibroblast surface. Two major pro-inflammatory cytokines, TNFα and IL 1β, were shown to be potential soluble factor involved. In conclusion, I was able to show two possible mechanisms through which mononuclear cells activate fibroblasts and induce ICAM 1 and HAS 2, thus perpetuating an inflammatory reaction. The first was through interaction with ICAM-1 on the fibroblast surface the second was through the release of pro-inflammatory cytokines.

The cardiac ryanodine receptor (RyR2) contains structural elements within the channel pore that function as gates to regulate the release of intracellular calcium, initiating cardiac muscle contraction. The precise regulation of these gates is critical in maintaining normal cardiac function, and channel dysfunction, resulting in altered calcium handling, underlies the mechanisms of arrhythmia and sudden cardiac death. The enormous size of RyR2 has impeded the gathering of detailed structural information, hence the structural determinants for channel gating remain unknown. Structural modelling studies have revealed similarities between the RyR2 pore and the K+ channel, KcsA, providing a framework in which to test channel gating mechanisms. A region termed the selectivity filter is a gating component in K+ channels, involved in inactivation and flicker closings, and its conformation is maintained by a transient hydrogen-bonding network. This project examined the role of the RyR2 selectivity filter in channel gating by generating mutants at analogous positions to KcsA that either disrupted (Y4813A, D4829A and Y4839A) or maintained (Y4813W, D4829E and Y4839W) a proposed hydrogen-bonding network, and assessed their intracellular Ca2+ release, ryanodine modification and biophysical properties. Y4813A and D4829A had drastic effects on channel function, whereas retaining physicochemical properties of conservative mutations, Y4813W and Y4839W, maintained the functional characteristics of WT RyR2. Flicker closings were affected by Y4839A mutation however, in general, single-channel gating for Y4813W, Y4839A and Y4839W was comparable to WT RyR2. Interestingly, monitoring single-channels for prolonged periods revealed novel insights into channel behaviour, characterised by inherent fluctuations in channel activity under steady-state conditions. This thesis reveals that the selectivity filter region is an important component for RyR2 channel function. However, it remains unclear whether the selectivity filter regulates channel gating, as the proposed hydrogen-bonding network would not be possible due to altered residue distances revealed from recent high-resolution RyR structural models.

Background: Differences in regional lumbar angles in sitting have been observed between subclassified groups of NSCLBP patients. However, differences during standing posture, range of movement and functional tasks, as well as differences in thoracic kinematics, have not been explored to date, despite classification-based cognitive functional therapy (CB-CFT) approaches being proposed to be effective for these subgroups. Methods: Spinal kinematics and trunk muscle activity of 27 Flexion Pattern (FP), 23 Active Extension Pattern (AEP) and 28 healthy controls were recorded (using 3D motion analysis (Vicon®) and surface electromyography) during: usual sitting, usual standing, flexion, extension, sit-to-stand -to-sit, reach up, stepping up and down, lifting and replacing a box and bending (and return) to pick up a pen tasks. Midpoint regional sagittal spinal angles and normalised amplitude sEMG for trunk muscles bilaterally were compared between groups. Statistical analysis was conducted using one-way ANOVAs (kinematics) and Kruskal-Wallis (muscle activity) tests. Results: Significant differences were observed between the AEP and FP groups in the upper lumber and lower thoracic spine during most postures and tasks. Some significant differences were also observed between the FP and control groups in these regions. Additionally, significant differences in the total lumbar spine between AEP and FP groups were occasionally evident. No differences in any other spinal region (or between AEP and control groups) were observed. Some significant differences (p < 0.05) in unilateral muscle activity were also observed between the NSCLBP and healthy control groups. Conclusion: The study findings further validate the classification approach (O’Sullivan, 2005). It highlighted that kinematic differences were observed to consistently occur in the thoraco-lumbar region during both static postures and functional tasks. Sub-division of regional spinal angles is key to identifying sub-group differences. These findings can inform novel CB-CFT interventions and highlights the need for targeted thoraco-lumbar spinal movement re-education strategies in NSCLBP subgroups.