The thesis comprises a literature review of the existing research on interventions to improve self-management of diabetes. An empirical paper describing the conception, designing and evaluation of a new psychological intervention for people with diabetes and self-management problems. The intervention is a mindfulness-based approach formulated for this population. The intervention was piloted with a group and individuals. The evaluation uses both qualitative and quantitative methodology to assess the acceptability, safety and feasibility of the new intervention. The third section of the thesis is a critical appraisal of the process of conducting the empirical research.

Behavioural interventions targeting physical activity alone produce clinically significant improvements in long-term glycaemic control in adults with type 2 diabetes. Effective translation of physical activity behavioural interventions into routine primary care is hindered by the lack of evidence-based training resources to equip healthcare professionals with the knowledge, skills and confidence to deliver behavioural interventions to their patients. This PhD thesis describes the development and open pilot testing of an evidence-informed multifaceted physical activity behaviour change intervention ‘Movement as Medicine for Type 2 Diabetes’ targeting: (i) consultation behaviour in primary healthcare professionals (online training programme); and (ii) physical activity behaviour in adults with Type 2 diabetes (patient toolkit delivered by primary healthcare professionals). Informed by multi-methodological development work (workshops to identify information/support needs of patients and healthcare professionals; systematic review to identify effective physical activity behaviour change components; and usability testing), Movement as Medicine for Type 2 diabetes was piloted in two primary care practices over a two month period. A qualitative process evaluation and treatment fidelity assessment were used to optimise the intervention by identifying barriers and enabling factors to implementation and informed revisions to content and study procedures. Six primary healthcare professionals completed the online training programme and delivered the toolkit to participating patients (N=30) during diabetes review appointments. Transferability of behaviour change techniques to other areas of practice was identified as a salient facilitator for healthcare professionals, although several implementation challenges were identified (e.g. previous negative experiences with supporting patients to increase physical activity behaviour). Intervention components were delivered by healthcare professionals to a satisfactory level of fidelity. Patients reported physical activity monitoring resources and review sessions as particularly beneficial components of the patient toolkit. Movement as Medicine for Type 2 diabetes was found to be acceptable and feasible in the primary care setting. Open pilot methodology facilitated optimisation of the intervention ahead of a planned pilot randomised controlled trial.

Background: Prevalence of type 2 diabetes (T2D) is rapidly increasing worldwide, linked to the obesity epidemic. There is substantial research evidence for T2D prevention by lifestyle interventions in high-risk individuals. The span of this research provides a unique case study with which to critically examine general guidance for development and evaluation of interventions to improve health. My research question is how might can effective, equitable and sustainable service provision for T2D prevention in high-risk individuals be achieved? Methods: Five papers reporting my empirical T2D prevention research form the core of my thesis. This research extends from the European Diabetes Prevention Study (EDIPS) RCT to the ‘New life, New you’ (NLNY) feasibility study. NLNY is a community based lifestyle intervention to reduce T2D incidence that is delivered by fitness trainers in North East England. To inform my research question I have reviewed intervention guidance history. I have then used T2D prevention as a case study, supported by my empirical research experience, to analyse this guidance Findings: Development of the NLNY intervention built on the EDIPS RCT evidence and experience. Pilot evaluation of NLNY suggests a feasible and acceptable intervention that is likely to be effective in preventing T2D. Prevention of T2D provided a useful exemplar for analysis of intervention guidance and highlighted strengths and limitations of existing guidance models. This analysis led to a proposed new guidance framework. Conclusions: The NLNY intervention provides a potential service provision model for T2D prevention in high-risk individuals. Well planned effectiveness and cost-effectiveness evaluation of the NLNY intervention is now needed. The analysis of intervention guidance and the proposed new framework will contribute to developing a robust study design. If effectiveness of the NLNY intervention is demonstrated there is potential for this community based intervention model to be further developed and adapted.

The incidence of Type 2 Diabetes Mellitus (T2DM) is increasing rapidly, therefore there is a need to detect this disease earlier and more efficiently, and also to identify novel risk factors that may aid both its detection and prevention. Aims: 1) To discuss the benefits and disadvantages of using HbA1c for diagnosis of T2DM and impaired glucose regulation (IGR). 2) To explore the impact on prevalence of using HbA1c to detect T2DM and IGR in global and local populations. 3) To determine if diagnostic cut-points are equivalent in different ethnic groups 4) To determine the use of the triglyceride-to-HDL ratio and its association with insulin resistance and whether statins and liver enzymes predict T2DM. 5) To investigate if Vitamin D deficiency has a role in the prevention of T2DM by designing a 6 month randomised controlled trial on vitamin D replacement. Key findings: 1) Using HbA1c for diagnosis has some logistical advantages over glucose testing, but may not detect the same people as having T2DM or IGR. 2) In Leicestershire, using HbA1c will increase numbers of people with T2DM and IGR. On global level, there will be regional variation on the effect on prevalence. 3) HbA1c, fasting and two hour plasma glucose are independently higher in South Asians (SA). Complications of T2DM may begin earlier in SA. 4) The triglyceride-to-HDL ratio associates with insulin resistance in Europeans and SA men but not women. Statin therapy reduces the risk of incident T2DM. Liver enzymes predict T2DM in Europeans but not SA. 5) Vitamin D deficiency may form a target for reducing insulin resistance in SA – the final results of the VITALITY study in 2015 will contribute to evidence base in this area. Conclusions: In this thesis I have explored new ways of detecting T2DM and IGR by using HbA1c and what impact this may have; also novel risk factors for T2DM have been investigated that may help improve methods of both earlier detection and prevention of T2DM.

Hydrogen sulfide (H2S) has been proposed as a novel endogenous regulator of vascular tone and inflammation. Novel slow release H2S donors (SRHOs) (e.g. GYY4137) have been shown to regulate blood pressure in experimental/genetically induced hypertension, and to inhibit tissue damage, oedema and inflammatory signalling in sepsis and arthritis. All these conditions involve perturbed mitochondrial function, oxidative stress and eventually apoptotic cell death. In the literature, the majority of studies pertaining to the vascular effects of H2S have been limited to the use of simple sulfide salts, NaSH and Na2S, which release H2S far too quickly to be physiologically relevant. Indeed, a study comparing NaSH and GYY 4137 showed that the level of H2S released from NaSH was rapidly high (400I-lM in the first 20min) and declined dramatically until disappearance. In contrast, H2S release from GYY4137 was low and continuous in time (-40I-lM up to 7 days). Therefore, to circumvent this problem, our team has designed several novel SRHOs, such as GYY4137, AP39, AP123, AP67 and AP72. These compounds offer the advantage over sulfide salts (i) to be not toxic (ii) to release H2S in a slow and sustained manner, (iii) to exert their effects at considerably lower H2S concentrations, and (iiii) to selectively target the mitochondria (in the case of AP39 and AP123). The latter combine a mitochondria-targeting moiety (triphenylphosphonium), a C2-C18 linker and H2S releasing moieties (dithiolethione or 4-hydroxythiobenzamide respectively). Preliminary experiments on these mitochondria-targeting SRHO showed a C1Q linker to be optimal; therefore AP39 and AP123 were chosen. Human cerebral microvascular endothelial cells (hCMEC/03) were exposed to oxidative stress agents, mitochondrial toxins and apoptosis inducing drugs in the presence/absence of the SRHOs. Cell toxicity was assessed using alamarBlue and flow cytometry, whereas cytosolic/mitochondrial reactive oxygen species (ROS) generation was estimated using H20CFOA, MitoSOX Red and Oihydroethidium fluorescence (spectrophotometry, flow cytometry). Mitochondrial membrane potential (~4Jm) was assessed using TMRM. Western immunoblotting and fluorescence activity assays were used to determine caspase3/7 activation/activity. Quantification of cytochrome c release from the 3 mitochondria was also attempted. Intracellular levels of ATP were assessed by the luciferase activity assay. Some antioxidant assays were finally used to determine whether the potential antioxidant effects of the SRHDs would be due to any scavenging effect. Cell viability was significantly preserved by all SRHDs. Besides, the potency of cytoprotection was substantially increased by the targeting mitochondria SRHD (AP39/AP123) from the j..IM to the nM range. The collapse of L14Jm normally observed in toxic conditions was inhibited by SRHDs. Overall "ROS" generation was markedly decreased after treatment with SRHDs. AP67 and AP72 inhibited caspase 3/7 activity when apoptosis was induced by staurosporine/etoposide. Cytochrome c release was positively modulated with our donors, as well as the ATP synthesis. These data suggest that SRHDs can inhibit/reverse oxidative stressmediated cellular injury, and highlight the increased potency of the mitochondria- targeting H2S donors AP39 and AP123 compared to GVY4137, AP67 and AP72. The measurement of the antioxidant capacity in cell-free system of the SRHDs showed (i) no scavenging effect of AP39 and AP123, confirming their release of H2S only in the mitochondria (ii) a scavenging effect of GVY4137, AP67 and AP72 only for high concentrations (>250j..lM), confirming that the antioxidant effects of these SRHDs in our cell system was unlikely due to any scavenging effect. Strategies increasing H2S bioavailability, in particular targeting mitochondria, may represent a new therapeutic opportunity to limit mitochondrial dysfunction.

Neonatal diabetes is a rare, genetically heterogeneous disorder. Mutations in 19 genes are known to cause the disease, including several transcription factors involved in pancreatic development. The aim of this thesis is to comprehensively assess the spectrum and role of mutations in known and novel pancreatic transcription factors in patients with pancreatic agenesis and neonatal diabetes. The first section gives an introduction to neonatal diabetes with a focus on pancreatic transcription factor mutations that cause neonatal diabetes in humans. This chapter is followed by a section describing the main methodologies used throughout the thesis. Biallelic mutations in the pancreatic transcription factor PDX1 have been shown to cause pancreatic agenesis. Chapter 1 investigates the possibility of hypomorphic PDX1 mutations causing isolated permanent neonatal diabetes (PNDM) without pancreatic hypoplasia. We identified 4 such patients, demonstrating that mutations in PDX1 can cause isolated PNDM. Chapter 2 describes the use of exome sequencing to identify novel disease genes, by looking for de-novo mutations in two patients with pancreatic agenesis. Both patients had mutations in the transcription factor gene GATA6. Sequencing of GATA6 in a cohort of 27 patients with pancreatic agenesis identified heterozygous mutations in 13 patients. All 15 patients present additional extra-pancreatic features. In Chapter 3 the role of GATA6 mutations was further investigated in patients with neonatal diabetes, which led to the identification of an additional 14 cases. Familial studies showed that GATA6 mutations can also cause adolescent/adult-onset diabetes. Taken together, these two chapters show that GATA6 mutations cause a variable diabetic phenotype, ranging from pancreatic agenesis, to adult-onset diabetes with or without extra-pancreatic features. Chapter 4 describes three patients with diabetes and chromosome rearrangements deleting GATA4, a transcription factor closely related to GATA6. Mutation analysis of all patients with neonatal diabetes of unknown genetic aetiology identified two additional cases, suggesting that GATA4 mutations are a cause of neonatal and childhood-onset diabetes. Chapter 5 illustrates the use of a targeted capture next-generation sequencing assay to simultaneously investigate all known neonatal diabetes genes in patients with neonatal diabetes of unknown genetic aetiology. This study investigates the impact of early, comprehensive genetic testing in a large international cohort of 1020 neonatal diabetes patients. Chapter 6 summarises the main findings of each chapter, describes their wider impact, and suggests areas of future work.

Treatment response in type 2 diabetes varies greatly: the same glucose lowering medication may have a marked effect in one individual but little effect in another. This is likely to have at least a partly biological basis (type 2 diabetes is a highly heterogeneous condition), raising the possibility of a personalised or 'stratified' approach to treatment where the therapy most likely to be effective for an individual is prescribed based on that person's clinical characteristics or biomarkers. One aspect of type 2 diabetes that varies greatly is beta cell insulin secretion. While many patients will be highly insulin resistant with relatively preserved insulin secretion, others are insulin sensitive with marked beta cell failure. It is plausible that these differences will be associated with variation in treatment response, particularly to treatments exerting their effect through beta cell dependant mechanisms. Insulin secretion is best assessed in clinical practice by measuring C-peptide, co-secreted in equimolar amounts to endogenous insulin. However the measurements commonly used in research are not practical for general clinical use. The aim of this thesis is to assess practical methods of measuring insulin secretion that could be incorporated into routine clinical practice and to examine the utility of these methods in diabetes treatment stratification. In Chapter 1 we review the current evidence on asseSSing insulin secretion and its potential use in diabetes treatment stratification. 3 In Chapters 2 and 3 we assess practical alternatives to the 'gold standard' mixed meal tolerance test stimulated C-peptide for assessing insulin secretion. We show that urine C-peptide creatinine ratio (UCPCR) and fasting C-peptide provide valid alternatives to stimulated blood tests for clinical practice, with high sensitivity and specificity for commonly used clinical thresholds. We demonstrate that a mixed meal test to assess insulin secretion can be undertaken with concurrent insulin given, retaining a high discriminative value for absolute insulin deficiency despite a marked reduction in hyperglycaemia. In Chapter 4 we demonstrate a strong inverse relationship between postprandial glucose rise and C-peptide, including fasting C-peptide and UCPCR. We demonstrate that those who have preserved insulin secretion have markedly reduced response to prandial (meal time) fast acting insulin. In Chapters 5 and 6 we explore problems of confounding by baseline glycaemia in studies of diabetes treatment stratification; we demonstrate that adjustment for baseline HbA 1 c in studies of stratification is both necessary and appropriate. In Chapter 7 we demonstrate in a large prospective cohort that markers of insulin deficiency, including low fasting C-peptide and positive GAD antibodies, are associated with reduced glycaemic response to GLP-1 agonist therapy in patients with a clinical diagnosis of type 2 diabetes. Participants who were Cpeptide negative and/or autoantibody positive had markedly reduced response to these therapies. An overview of the major finding of each chapter and their implications potential future research are discussed in Chapter 8.

The background to this research is the primary lymphoedema clinic held at St George's Hospital, London. This is a multidisciplinary clinic run by the combined expertise of dermatology and genetic teams, and is the only one of its kind in the UK. The aim of the service is to diagnose, investigate and manage patients with primary lymphoedema. To date, 7 genes are known to cause a phenotype where lymphoedema predominates. The main aim of my MO(Res) project was to use and refine Connell et aI's diagnostic classification pathway (Connell, Brice et al. 2010) for patients with primary lymphoedema in order to identify phenotypically similar groups of patients who could then be the subject of molecular investigation. This, in turn, would hopefully lead to the discovery of new genotypes responsible for subtypes of primary lymphoedema . Lymphangiogenesis, lymphoedema, its clinical features, diagnosis and investigation modalities, and the well -established phenotypes of primary lymphoedema are discussed in the introduction. The laboratory techniques that have been used (including Sanger sequencing and next generation sequencing) are described. Subsequent chapters describe the individual projects that have been carried out in an effort to phenotype and genotype cohorts of patients with primary lymphoedema, including the discovery of a new causal gene for primary lymphoedema. One chapter is dedicated to the development of magnetic resonance lymphangiography (MRL) as an investigative tool in patients with lymphoedema to improve accurate phenotyping of the subgroups, as current imaging techniques are limited. Finally, Connell et aI's diagnostic pathway has been updated on the basis of new findings.

There have been significant changes in the management of patients with diabetes in the United Kingdom (UK) over the last 20 years, with the introduction of analogue insulins and the drive toward tighter glycaemic control, with its concomitant risk of increased hypoglycaemia. This thesis describes changes in insulin prescribing in the UK between 1999 and 2009, examines the associations between a patient's characteristics and the type of insulin they are prescribed, and between a patient's insulin exposure and their risk of serious hypoglycaemic events. Data for the study was obtained from the General Practice Research Database (GPRD), a UK prirpary-care database providing anonymised patient data for approximately 690 of the UK population. A cohort of insulin-using patients was identified, from which subgroups of patients newly-starting insulin therapy and patients switching insulins were extracted for analysis. There was a rapid uptake of analogue insulins and by 2009 they accounted for more than 8090 of insulin prescriptions. While some of a patient's characteristics, including age and ethnicity, influenced the choice between analogue and human insulin in new insulin users, the largest effect size was the calendar year in which therapy started: clinician (or patient) choice seemed to be th~ driving factor. The risk of serious hypoglycaemic events under exposure to human or analogue insulin was examined in new users and insulin switchers. After adjusting for patient characteristics, some of which, including ethnicity, were associated with a change in the risk of an event, the study confirmed that new users of long-acting analogue insulins had a lower risk of serious hypoglycaemic events compared with users of human insulin. However, following a switch from human insulin no difference in the risk of serious hypoglycaemic events was found in patients switching to either analogue insulin or to a different combination of human insulins.

Diabetes mellitus is a chronic disorder characterised by a blood glucose level persistently above normal due to loss of insulin production from the pancreas or peripheral tissues becoming resistant to insulin. Currently, treatment for diabetes accounts for 12% of the world's total health expenditure. There are a range of therapies available for the treatment of T2DM; however treatments can be expensive and associated with adverse effects. The studies conducted in this thesis investigated the ability of seaweed extracts to exert anti-diabetic activities through established pharmaceutical mechanisms. Some extracts from species of Irish seaweed were :1 able to reduce glucose diffusion, inhibit alpha glucosidase activity, inhibit DPP-4 activity, and also promote GLP-l secretion and biosynthesis in an enteroendocrine cell model. Furthermore, acute oral administration of Porphyra linearis reduced glucose excursions in normal mice. A commercial seaweed extract (Digestica®) was able to potently inhibit alpha glucosidase activity in a non-competitive manner and in concentrationdependent fashion. Digestica extracts were able to inhibit DPP-4 activity, again in a concentration-dependent manner. Digestica promoted GLP-l and GIP secretion in vitro and increased GLP-l biosynthesis in an enteroendocrine cell model. Digestica was able to prevent the induction of T2DM in a high fat fed mouse model by preventing the development of glucose intolerance and insulin resistance. Moreover, Digestica acted as a therapeutic for T2DM improving glucose tolerance and insulin sensitivity in the same animal model. Selected marine compounds commonly found in seaweeds were also investigated for their anti-diabetic effects in vitro with promising results. These studies demonstrate potent anti-diabetic effects displayed by seaweed extracts and common marine compounds both in vitro and in vivo without exhibiting cytotoxic effects showing their potential use as inexpensive treatments for T2DM, perhaps as food supplements or natural therapeutics.