Embryonic stem (ES) cells represent a potentially limitless supply of tissue for &'946;-cell replacement therapy because of their properties of unlimited self renewal and pluripotency. Initial differentiation protocols for obtaining &'946;-cell from ES cells were based on a protocol for obtaining neural cells from ES cells. A differentiation protocol which mimics development in the developing embryo would be more appropriate. The aim of this study was to design a protocol for driving the differentiation of mouse ES cells towards a &'946;-cell fate using growth factors and transcription factors thought to be involved in this process in vivo. A three-step protocol was designed for the differentiation of CGR8 ES cells towards a pancreatic &'946;-cell fate. The first 2 steps involved exposure to activin A (previously proven to induce endoderm) followed by exposure to 2 factors thought to be involved in pancreas specification from endoderm in vivo: retinoic acid (RA) and fibroblast growth factor 2 (FGF2). Two Tamoxifen inducible DNA constructs, created to express the &'946;-cell specific transcription factor, Pax4, were designed for use in the third step; where the cells were exposed to 4-OH Tamoxifen thus causing Pax4 to become active. The results of the study demonstrated that activin A promotes differentiation into endoderm, with BMP4 promoting differentiation into mesoderm. One of the inducible constructs (pP6PAX4FLAG/ERT2), was highly responsive to 4-OH Tamoxifen, showing an effect after 2 hours exposure whilst the other seemed unresponsive. A protocol designed to drive differentiation of the CGR8- pP6PAX4FLAG/ERT2 clone toward a &'946;-cell phenotype did not succeed but this was probably due to problems with the early stages of differentiation and further work has the potential to result in a successful protocol.

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Macrovascular contraction and relaxation effects of metformin were measured using a Mulvany Halpern myograph. Mouse aortic ring sections were treated for 1 and 4 hours in vitro with metformin at 10-5M, and for 2, 4 and 8 weeks in vivo with metformin at 250mg/kg/day. The rings were contacted with increasing concentrations of noradrenaline (10-9M, 10-8M, 10-7M, 10-6M) in the absence and presence of metformin. Maximally contracted tissue was then relaxed using increasing acetylcholine concentrations (10-9M, 10-8M, 10-7M, 10-6M). Meformin increased the sensitivity of the aorta to noradrenaline-induced contraction. The maximal effect in vitro was seen after 4 hours giving a 221% increase in contraction after 4 hours at noradrenaline 10-6M. Acetylcholine-stimulated relaxation via endothelium also increased with metformin after 4 hours by 36.85%. The maximal effect of metformin treatment in vivo was seen on aortic contraction after 8 weeks: the effect of melformin treatment on relaxation was less marked at this time. Metformin also increased passive tension generated by the aortic vessel wall after 4 hours, which was reversed by administration of papaverine, which acts directly on vascular smooth muscle. Metformin was shown not to alter nitric oxide production by the mouse aortic wall after 1 and 4 hours in vitro. Metformin lowered basal calcium concentrations, as measured by FURA/2AM, generating a slow sustained increase in calcium release induced by noradrenaline during contraction. This research programme has shown that metformin can increase both the contraction and relaxation capabilities of aortic sections treated both in vitro and in vivo with therapeutic concentrations of metformin at 10-5M. Metformin has been shown to act directly in the vascular wall to alter vascular contractility via effects on both vascular smooth muscle and endothelium, and to influence calcium movements independently of nitric oxide.

Diabetes mellitus designates a range of metabolic disorders characterised by hyperglycaemia due to deficient or absent insulin secretion, insulin action, or both. In particular, Type 1 diabetes is characterised by a total lack of endogenous insulin secretion which has to be replaced by exogenous insulin to control the plasma glucose concentration. An extracorporeal wearable artificial pancreas (AP) has been a research aim for over three decades. The research is motivated by the need to improve glucose control. Results of a major study, the Diabetes Control and Complications Trial (DCCT), have demonstrated that improvements in glucose control prevent or delay long term complications, which are the main causes of morbidity and mortality in subjects with Type 1 diabetes. Prior to a clinical evaluation, performance of new medical devices can be tested in silico. Such an approach has been adopted extensively by the pharmaceutical industry in the development of new drugs. In silica testing benefits from relatively low financial, human, and time costs by comparison with the resources required for a full clinical evaluation. The aims of the present thesis are to identify components of the AP, integrate them into a simulation environment, and design an in silico evaluation strategy for the development of closed-loop algorithms with the ultimate goal to assess safety and efficacy prior to clinical evaluation. In the present work, submodels of metabolic processes were linked to represent the characteristics of the glucoregulation in Type 1 diabetes. The submodels were associated with sets of parameters to account for variability in population and individual responses to meals and insulin therapy. The model of glucoregulation in Type 1 diabetes was extended by models of subcutaneous (sc) glucose sensing and sc insulin delivery to represent all aspects of the AP. A systematic approach was developed and employed to evaluate, in silica, the potential and limitations of an AP glucose controller. This was exemplified by evaluating a nonlinear model predictive controller. The robustness of the AP was explored by hypothesising various perturbations induced by different system components. A further objective included the establishment of a qualitative grading scheme of glucose control from the clinical viewpoint. This was followed by a comparison between results from simulations and a clinical trial of 24 hours, which gave the proof of concept of in silica testing. It was found that despite discrepancies due to initial conditions and meal differences, the simulations indicated well the outcome of the clinical trial. In conclusion, the thesis demonstrates the significant potential of in silica testing to make predictions about system behaviour aiding the assessment of safety and efficacy of control algorithms during the development of an AP.

The increasing comorbid burden of tuberculosis (TB) and diabetes mellitus (OM) worldwide requires the management of all stakeholders including pharmacists. This raises the question whether current single disease management system fulfils patients' health needs and whether pharmacists could effectively play a role in enhancing the joint management of these two commonly associated diseases. Pharmacists have begun to provide pharmaceutical care through pharmacist-led medication therapy adherence clinics and clinical pharmacy services for several diseases and conditions (e.g. OM, asthma) in some public hospitals in Malaysia but are yet to be involved in the management of TB. The management of TB has been largely delivered through directly observed treatment (OOT) as high level of adherence to treatment is vital. However, little is known on how TB patients with OM are being managed and how these patients cope with their medication. The aim of this study was to develop a pharmaceutical care service for patients with TB and OM. The first three phases (preclinical, phase 1 and phase 2) of the UK Medical Research Council framework for the development of complex interventions to improve health was applied to develop a pharmaceutical care service for patients with both TB and DM in a tertiary hospital in Malaysia. First, literature relating to TB and OM was reviewed (preclinical). Second, the pharmaceutical care needs of TB and DM patients were explored using semi-structured interviews with twenty patients, three physicians, three nurses, and a focus-group with four pharmacists (phase 1). Third, action research was conducted to assess the feasibility of providing a pharmaceutical care service for patients with TB and OM (phase 2). This study received ethical approval from the Medical Research and Ethics Committee (MREC), Ministry of Health, Malaysia. Patients and health care professionals reported several medication-related issues in the phase 1 study. Patients were most inclined to discuss their concerns about their medication. Patients also tended to display different attitudes towards medication-taking, depending on their beliefs, the severity of illnesses, perceived efficacy of the treatment, and the severity of medication-related problems. The findings also revealed that many of these concerns had not been discussed with their physicians. This was also caused by the patients' and physicians' tendencies to prioritise the management of TB, and unintentionally neglecting other comorbidities especially when patients were primarily managed at the chest clinic. Other difficulties identified in comorbid management included delayed initiation of both TB and OM treatment, chest physicians' lack of confidence in managing 'difficult' OM in TB patients and the burden of attending multiple clinics for patients. Health care professionals believed that pharmacist-led medication therapy adherence clinics (MTACs) encouraged the provision of patient-centred care, enhanced pharmacist-patient communication, created opportunities for inter-professional interactions and could be used as a model to provide pharmaceutical care services. Health care professionals urged pharmacists to play a role in the management of TB and OM by providing patient education and counselling. The phase 2 study revealed that the prevalence of OM in TB patients was 15%. Action research allowed the researcher, together with a hospital pharmacist, to identify pharmaceutical care needs in TB and OM patients, and fulfilled some of them. Pharmaceutical care issues identified included lack of medication adherence, poor management of OM, the need to manage adverse drug reactions, and the lack of frequent monitoring of certain monitoring parameters for TB, OM and other comorbidities at the chest clinic. Many patients had uncontrolled OM, however, many were more likely to be adherent to TB medication than medication of OM and/or other conditions. As a follow-up action, pharmacists advised these patients to place equal importance to TB and non-TB related management. Additionally, pharmacists also made treatment recommendations and referred patients to their chest physicians for further management of medication-related problems. Nevertheless, there were barriers that impinged the provision of pharmaceutical care service. The barriers include the lack of space with privacy to provide education and counselling to patients; the unavailability of medication records and other clinical information for comorbidities at the chest clinic; and the lack of time to develop inter-professional relationship. Despite the need to address the barriers, the provision of pharmaceutical care service to TB and OM patients was feasible as pharmacists were able to integrate TB and OM management by identifying, communicating, and resolving some medication-related problems. In summary, this study provided the groundwork by conducting phase 1 and phase 2 study prior to developing a full-fledged pharmaceutical care service for TB and DM patients. Future work can be done to improve the service through critical analysis of the challenges faced in the developmental phase with the effectiveness of the service care plan assessed through a randomised controlled trial (RCT).

Background: Attaining optimal glucose control in type 2 diabetes (T2DM) is essential to minimise complications, but difficult to achieve in practice with declining β-cell function in patients. Aim: To evaluate the efficacy and feasibility of a protocol-led, treat-to-target approach emphasising earlier oral hypoglycaemic agent (OHA) combination and insulin use to target basal and prandial glucose in primary care. Methods: T2DM patients aged 40-75 years, with glycosylated haemoglobin (HbA_1C) 6.4-10.0% on diet or oral monotherapy in 7 practices were asked to participate in a protocol-led, target-driven programme of care using early OHA and insulin therapy combinations to target basal and prandial glucose. Recruited patients were randomised to pre-specified algorithms with monthly therapy adjustment aiming for fasting plasma glucose values (FPG) <6mmol/L and 2-hour post-prandial (PPG) <8mmol/L. Isophane or humalog insulin was added where glucose targets were not attained. All patients were reviewed at one year. Glucose, weight and cardiovascular (CVS) risk factor outcomes were measured and the feasibility, acceptability and safety of the programme evaluated with validated questionnaires. Results: 345 patients were studied. 60 participated in the glucose study and 285 received standard care. 65% of recruited patients were male, with mean (SD) age 61.0 (8.2) years, BMI 29.8 (5.3) kg/m2, HbA_1C 7.5 (0.9)% and median (IQR) T2DM duration 3 (1-5) years. 41 completed the study. Glucose control was significantly improved in recruited compared to non-recruited patients with a mean overall 0.8% HbA_1C reduction and 72% patients achieving HbA_1C<7% and 64% <6.5% at 1 year. Therapy use was also significantly increased with 50% requiring 3 therapies, 30% 2 and 10% monotherapy. 24 (59%) of recruited patients received isophane and 16 (39%) humalog to achieve targets. Glucose optimisation was achieved without significant hypoglycaemia or adverse events and patient quality of life (QoL) and therapy satisfaction remained high. CVS risk was reduced in recruited patients. Conclusions: A more structured, target-driven approach to glucose optimisation can successfully achieve normoglycaemia in a selected primary care T2DM population without significant adverse events, hypoglycaemia. or QoL impairment in patients. A principal barrier to wider implementation of this programme appears to be practice and patient participation.