DHEA is an adrenal derived hormone with a unique secretory pattern with highest serum concentrations observed in middle age. Individuals with adrenal insufficiency exhibit gross DHEA deficiency though its replacement in this context is not commonplace. DHEA is known to behave as a pro-hormone with the ability to be converted into either androgenic or oestrogenic terminal hormones whether this is its sole physiological role still remains unclear. Various animal and in vitro studies have suggested that treatment with DHEA can precipitate improvements in body fat, adipocytokine profiles, insulin resistance and estimates of vascular disease. Human studies have demonstrated inconsistent results and have tended to focus on the physiological deficiency of DHEA associated with normal aging and not the pathological DHEA deficiency seen in adrenal insufficiency. The aims of this thesis were: (1) To determine the effect of DHEA on preadipocyte (cell line and primary) proliferation and differentiation and to examine the mechanisms behind any observed effects. (2) To evaluate the effect of replacing DHEA on vascular function and body composition in subjects with primary and secondary adrenal insufficiency. (1) DHEA inhibited proliferation in all preadipocytes examined secondary, at least in part, to cell cycle blockade. DHEA inhibited adipogenesis in omental but not subcutaneous derived preadiocytes. (2) Arterial stiffness and endothelial function was not affected the total population but stratification by study group showed that DHEA replacement reduced central blood pressure in patients with secondary adrenal insufficiency. Body composition was not affected in either subject group.

In this thesis we investigate the interaction between drugs and toxins with membranes using red blood cells (RBCs) as morphoelastic probes. Using fluctuation spectroscopy, we were able to probe the RBC mechanical response to a simulated diabetic environment and to investigate the effect of metformin, one of the most widely used medicines to treat diabetes. Healthy RBCs were incubated in high levels of glucose or glucose and metformin and their mechanical properties were tested upon the exposure to oxidation with hydrogen peroxide (H2O2). My results show that the response to oxidation and glycation is different for different donors, with some donors more susceptible to oxidation than others. I have also found that glycated cells are more susceptible to oxidation with H2O2 than control and metformin treated RBCs. Metformin treated RBCs show a response to oxidation similar to control cells which suggests that metformin may have some antihyperglycaemic and antioxidant effects which could preserve the RBCs membrane elasticity within the normal limits, counteracting the adverse effects of oxidative stress. The interaction between the RBC membrane and two of the Clostridium perfingens toxins, α-toxin and NetB, is next studied in this thesis. Using fluctuation and absorbance spectroscopy, changes in the RBCs morphology caused by the toxins can be monitored allowing us to describe the course of the toxin membrane interaction. I conclude that the two toxins studied in this thesis have two different mechanisms of action. Both toxins produce a decrease in the cell radius but through two different mechanisms. NetB causes a decrease in the cell radius by forming large pores in the red cell membrane allowing for quick lysis and the exchange of material across the membrane. Whereas α-toxin causes a decrease in the cell radius by hydrolysing specific lipids in the cell membrane without necessarily causing the formation of membrane pores. These differences in the interactions between the two toxins and the red cell membrane have distinct fingerprints in the evolution of the cell shape and membrane thermal fluctuation dynamics. Fluctuation and absorbance spectroscopy were also used to investigate the effect of nitroglycerine (GTN) on the RBC morphology and mechanical properties. This study was prompted by a recent report in the literature that related decreases in the viscosity of whole blood to changes in the membrane surface charge. My results show that changes in the electrophoretic mobility of GTN-treated RBCs strongly depend on the incubation time. Cells incubated in GTN for 5 minutes decreased their mobility by about 20% whereas cells incubated for 20 minutes increased their mobility by about the same amount. Further investigations on the RBC morphology showed that GTN causes changes in the RBC shape. The matching times scales between those experiments and the electrophoretic experiments made me conclude that RBCs shape may play a role in the electrophoretic mobility of the RBCs treated with GTN. The main results obtained in this thesis demonstrate the viability of the idea of using RBCs as morphoelastic probes, which can provide detailed information about the interaction of solutes of interest and the plasma membrane. At the end of this thesis I propose use of RBCs in such a capacity to monitor the progression of disease by comparing the cell elastic state to clinical markers of disease.

Heparan sulfate proteoglycans (HSPGs) exist in pancreatic beta cells, and HS seems to modulate important interactions in the islet microenvironment. However, the intra-islet structures of HS in health or altered glucose homeostasis are currently unknown. This work shows that distinct spatial distribution of HS domains is present in islets in the adult, that intra-islet HS domains are mostly conserved between rodents and humans, and that HS is abundant in glucagon producing islet alpha cells. In beta cells HS is characterised by 2-O, 6-O and N- sulfated moieties, whereas HS in alpha cells is N-acetylated, N-, and 2-O sulfated and low in 6-O groups. Differential expression of three HS modifying genes in alpha and beta cells was observed and may account for the different HS patterns. Immunoelectron microscopy localized the alpha cell HS epitopes in alpha cell intracellular granules. Furthermore, FGF1 and FGF2 were present in alpha cells, whereas functional FGFRs exist in beta cells, but not in the alpha cell line aTC1-6, or in primary alpha cells in islets. Intra-islet FGF21 was shown in beta cells with low levels of expression, bound mainly to the co-receptor βKlotho in unstimulated conditions and FGF21 gene expression was down-regulated by hyperglycaemia in vitro. In contrast, islet and beta cell FGF1 was constitutively expressed in vitro, in leptin resistant Zucker rats and in insulinopenic, hyperglycaemic rats. FGF1 induced signalling was dependent on 2-O, and 6-O HS sulfation in beta cells, and HS desulfation reduced beta cell proliferation and potentiated oxidant induced apoptosis. In leptin resistant animals and in islets from streptozotocin treated rats there was a reduction in alpha cell HS expression. These data demonstrate a distinct HS expression patterns in alpha and beta islet cells and propose a novel role for alpha cells as a source of paracrine FGF ligands to neighbouring beta cells with specific cell-associated HS domains mediating the activation and diffusion of paracrine ligands. The intracellular alpha cell HS that localises in vesicles, suggests a role for HS in the regulation of vesicle trafficking and/or hormone secretion.

No description available.

Sera from patients with thyroid disorders or diabetes were tested for the presence of soluble immune complexes using a number of established or modified assay systems. In addition, a new and sensitive assay for the detection of 'complexes' was developed. The first assay system applied was based on the inhibition of the antibody dependent cell-mediated cytotoxic (ADCC) activity of rat spleen cells by soluble complexes which compete with antibody-coated target cells for Fe receptors on the effector cells.

No description available.

No description available.

Background: The increased prevalence of Type 2 Diabetes (T2D) in the UK has seen the adoption of empowerment models of T2D management. Research exists which contrasts patients’ and healthcare professionals’ perspectives of T2D management. However, no studies explicitly contrast healthcare professionals’ and patients’ perspectives within the empowerment approach. The overall aim of this study was to explore healthcare professionals’ and patients’ perspectives of managing T2D in a context where empowerment is the prevailing health paradigm. The three research questions informing the aim sought to explore: 1) What are patients’ and healthcare professionals’ perceptions, understanding and experiences of successful and unsuccessful (un/successful) T2D management? 2) What barriers and enablers do patients and healthcare professionals perceive, understand and experience in relation to managing T2D on a day-to-day basis? and 3) What similarities and differences emerge in patients’ and professionals’ perceptions, understanding and experiences of managing T2D on a day-to-day basis? Methods: This is cross-sectional, qualitative research using maximum-variation sampling with healthcare professional and patient participants in an empowerment-based T2D patient participation group. Semi-structured interviews (N = 25 patients / N=10 healthcare professionals), focus groups (3 x N = 12 patients) and open-ended questionnaires (N = 6 patients) were used. Data were analysed thematically using framework analysis. Findings: Patient management of T2D developed from factors in their personal and social contexts. T2D affected patients in differing ways across the course of the illness (i.e. diagnosis, adaption and eventual self-management) and patients had different resources available in their social contexts with which to manage these effects. Diagnosis was shaped largely by the different types of uncertainty patient participants experienced on their diagnostic route, and the progession of the illness prior to detection and diagnosis which shaped the barriers and enablers they experienced. Healthcare professionals, on the other hand saw diagnosis as a springboard to self-management and tended to interpret patients’ experiences of uncertainty as ‘resistance’. Therefore, barriers to responding to diagnosis were seen as largely patient-related. Patient participants reported adapting to living with T2D as an ongoing process of adjusting their personal (and wider) relationships, as well as social activities, to ensure their T2D-related needs were met. Conversely, although healthcare professionals saw adaption as important, it was seen as a brief adjustment period after diagnosis and before full self-management. This highlighted another area where healthcare professionals and patients gauged successful management differently, and saw different barriers and enablers. Patients also experienced varied barriers and enablers and evaluated successful management using diverse criteria, largely shaped by factors in their social context. Healthcare professionals expected patients to ‘own their illness’ which was seen to reduce pressure on finite health resources, and that clinical advice would create behaviour change. Healthcare professionals’ perspectives on successful self-management revolved around clinical evidence, the healthcare system and socio-contextual constraints, and portrayed barriers and enablers to managing T2D largely as patient or healthcare system related. Discussion: Similarities and differences in healthcare professionals’ and patients’ perspectives of T2D management were seen. These occurred as a result of the fit between patients’ styles of self-management and healthcare professionals’ expectations surrounding behaviour change and health resources management. These differences were also evident during the diagnosis and adaption stages of the illness. Conclusion: Management of T2D was seen and experienced differently by healthcare professionals and patients where empowerment of the patient to self-manage was the prevailing paradigm. The lack of fit develops largely due to the different evaluative contexts and criteria which healthcare professionals and patients use to manage T2D, and the different expectations that healthcare professionals and patients have of one another.

No description available.

The aims of this thesis were to establish the prevalence of erectile impotence in male diabetics, investigate the aetiology and follow the natural history in order to gain a fuller understanding of the condition. From a cohort of 541 diabetic men aged 20-59 years it was found that 190 (35%) were impotent. Impotence was associated with age, retinopathy and symptomatic peripheral and autonomic neuropathy. Subgroups of patients were investigated in more detail using cardiovascular autonomic function tests and by the non-invasive evaluation of bladder function. Impotent diabetic men had more abnormalities in bladder function. The severity of bladder dysfunction correlated with the degree of abnormality in cardiovascular autonomic function tests. Four hundred and sixty-six of the original cohort were followed prospectively over a five year period. Only 11 of those originally impotent had regained potency while 78 out of 275 who were originally potent became impotent. Apart from age, the most important factors, present originally which increased the likelihood of developing impotence were poor diabetic control, heavy alcohol intake and the presence of retinopathy or intermittent claudication. Sixty-three patients died, but impotence per se was not associated with increased mortality. The thesis concludes by discussing the implications of these data for the prevention and management of erectile impotence in diabetic men.