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The incidence of cardiomyopathy and cardiac fibrosis is markedly increased in patients with diabetes mellitus. As cardiac fibrosis is mediated by myofibroblasts, we investigated the effect of diabetes-associated collagen glycation on the conversion of cardiac fibroblasts to myofibroblasts. Collagen glycation was modeled by the glucose metabolite, methylglyoxal (MGO). Cells cultured on MGO-treated collagen exhibited increased activity of the α-smooth muscle actin (SMA) promoter, elevated levels of collagen I, α-SMA mRNA, and enhanced protein expression of α-SMA, ED-A fibronectin and cadherins. Increased expression of α-SMA was dependent on β1 integrins and on TGF-β. In collagen gel assays, MGO-collagen promoted faster contraction and cell migration was increased by MGO-collagen. In shear-force detachment assays, cells on MGO-collagen were less adherent, and β1 integrin activation and focal adhesion formation were inhibited. We conclude that collagen glycation augments the formation and migration of myofibroblasts, critical processes in the development of cardiac fibrosis in diabetes.

The incidence of cardiomyopathy and cardiac fibrosis is markedly increased in patients with diabetes mellitus. As cardiac fibrosis is mediated by myofibroblasts, we investigated the effect of diabetes-associated collagen glycation on the conversion of cardiac fibroblasts to myofibroblasts. Collagen glycation was modeled by the glucose metabolite, methylglyoxal (MGO). Cells cultured on MGO-treated collagen exhibited increased activity of the α-smooth muscle actin (SMA) promoter, elevated levels of collagen I, α-SMA mRNA, and enhanced protein expression of α-SMA, ED-A fibronectin and cadherins. Increased expression of α-SMA was dependent on β1 integrins and on TGF-β. In collagen gel assays, MGO-collagen promoted faster contraction and cell migration was increased by MGO-collagen. In shear-force detachment assays, cells on MGO-collagen were less adherent, and β1 integrin activation and focal adhesion formation were inhibited. We conclude that collagen glycation augments the formation and migration of myofibroblasts, critical processes in the development of cardiac fibrosis in diabetes.

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Objectives: Body image concerns and problematic eating attitudes and behaviours are recognised as an important concern for young people and clinicians. Identification of groups that might be at risk of developing such problems would enable prevention and early implementation strategies to be implemented. The aim of this study was to explore body image concerns and eating attitudes and behaviours in a community sample of young people aged between 12 and 17 years. Design & Method: A cross sectional between groups design was employed. Participants completed self-report assessments measuring attitudes to body shape and weight, eating attitudes, behavioural features of eating disorders and eating disorder psychopathology. Siblings of young people with type 1 diabetes (n=12) were compared to a matched control group (n=12). Comparisons were also made between males and females and between age groups in a community sample (n=75). Results: Compared with females without a diabetic sibling, females with a diabetic sibling reported more concerns about body weight and shape, greater disturbance in eating attitudes, and significantly higher levels of eating disorder psychopathology. Within the community sample, concerns about body shape and weight and disturbed eating attitudes were significantly higher in females than in males. The results highlighted a high prevalence of excessive exercise behaviour in both males and females. Excessive exercise was the only variable to differ significantly between age groups, with the highest prevalence reported in those aged 14 and 15 years. Conclusions: The findings provide tentative support for the hypothesis that siblings of young people with type 1 diabetes are at increased risk of disturbed eating attitudes and behaviours, but require replication in a larger sample. Methodological implications of the study and suggestions for further research are discussed.

Investigation into the molecular basis of diabetes-induced erectile dysfunction was carried out on corpus cavernosum tissue from control and streptozotocin (STZ)-induced diabetic rats. Increased expression of endothelial nitric oxide synthase (eNOS) and increased nitrosative stress was found in the diabetic penile tissue. Diabetic neuropathy is associated with a failure in axonal regeneration. <i>In vitro</i> axon growth, guidance and regeneration model systems were used to study both the intrinsic nature of diabetic DRG neurones to grow and respond to guidance cues, but also to study the supportive properties of the diabetic nerve environment for axon regeneration. Cyclic Adenosine-3’5-Monophosphate (cAMP) signalling is compromised in diabetic DRG neurones. Increasing cAMP in diabetic neurones reduced the deficit in neurite growth and reversed the aberrant response of diabetic neuronal growth cones to the cAMP-dependent trophic factor, nerve growth factor (NGF). Diabetic neurones display reduced expression of Exchange protein activated by cAMP (Epac), a downstream effector of cAMP. Specifically activating Epac also rescued the abnormal turning responses of diabetic DRG neurones. Impaired axonal regeneration in diabetes has been, in part, attributed to delayed Wallerian degeneration. Using the cryoculture bioassay, it was shown that during regeneration the peripheral nerve environment, of both control and diabetic rats are equally supportive for axon growth. In summary, this is the first study to imply that specific signalling mechanisms, involving the cAMP-Epac pathway, may be compromised in neurones from diabetic rats, which contribute towards reduced neurite growth and abnormal responses to axon guidance cues.

Acute post-prandial hyperglycaemia may trigger acute increases in systemic inflammation and oxidative stress, potential risk factors for cardiovascular disease. We aimed to investigate the effect of both an oat-rich diet and standard dietary advice on glycaemic control, post-prandial glycaemia, inflammation and oxidative stress in volunteers with diet-managed type 2 diabetes. Method In a randomised cross-over design 30 volunteers with type 2 diabetes followed for two 8 week periods either an oat-rich diet or a control diet based on standard dietary advice. Volunteers attended at baseline, week 8 and 16. Measurements included basic clinical parameters, baseline blood tests followed by consumption of a standard test-meal with glucose, insulin and markers of inflammation and oxidative stress quantified during the post-prandial period. During each dietary period volunteers underwent a three day period of free-living continuous glucose monitoring (CGMS) during which time they kept a food diary. At the study end, the post-prandial response to an iso-energetic meal rich in oats was assessed. Results There were no diet-related differences in parameters of glycaemic control or post-prandial glycaemia based on CGMS data. Dietary intervention also had no effect on either the glycaemic or insulinaemic responses to the test-meal and there were no differences in the acute responses to the standard or oat-based test meals. Chronic intervention with the oat-based diet increased fasting adiponectin concentrations (P=0.06) and post-prandial ORAC concentrations (P<0.05) but had no effect on fasting CRP, ORAC, OxLDL or TBARS concentrations or the post-prandial response of CRP, adiponectin or TBARS to the standard test-meal. Following the oat-based test meal, adiponectin concentrations declined less (P<0.05) while CRP concentrations increased less (P<0.05) compared with the standard test-meal. Conclusion The oat-rich diet exerted anti-inflammatory and anti-oxidant effects in both the basal and post-prandial states independent of any effect on glycaemia. These findings may have implications for nutritional management of cardiovascular risk.

Type-2 diabetes (T2D), a multi-factorial disease characterised by chronic hyperglycaemia, is caused by a complex interaction between genetic and environmental factors. Genetic and phenotypic characterisations of diabetic patients suggest that a combination of &beta;-cell failure, culminating in defective insulin secretion, as well as impairment of glucagon secretion in &alpha;-cells is central to the aetiology of the disease. Mouse models represent a valuable tool in such investigations. With the advent of large-scale genetic tools, a myriad of novel susceptibility loci associated with T2D have been identified. For many of these genes, it is unclear how genetic variation is linked to increased disease susceptibility. Our first study elucidates the implication of a transcription factor, SOX4, which is believed to underlie a T2D susceptibility locus (CDKAL1) in human. Using an N-ethyl-N-nitrosourea (ENU) mouse model, we explored &beta;-cell function in mice carrying a point mutation in Sox4 (Sox4mt mice). This mouse strain displayed a significant reduction in glucose-stimulated insulin secretion (GSIS) that was associated with a 2-fold increase in wild-type Sox4 expression. The exocytotic events in mutant &beta;-cells, as measured by single-vesicle (on-cell) capacitance measurements, suggested the presence of a persistent fusion pore. Subsequent failure of fusion pore expansion beyond the initial 1–2 nm results in an incomplete insulin release due to steric hindrance (insulin diameter: 3–4 nm). The proportion of full fusion events diminished in favour of kiss-and-run events in mutant &beta;-cells. Stxbp6, which encodes amisyn, was shown to be the target gene of Sox4. Increased expression of amisyn, a protein previously shown to be involved in the stabilisation of the fusion pore in chromaffin cells, was observed in islets isolated from Sox4mt mice. The possible involvement of amisyn is further suggested by the finding that overexpression of amisyn mimicked the effect of the Sox4 mutation and resulted in reduced insulin secretion. Knockdown of amisyn expression restored the secretory defect in Sox4mt-overexpressing cells. Importantly, the effect of the Sox4 mutation was recapitulated by the overexpression of Sox4. Similar effects were obtained in the human &beta;-cell line EndoC-&beta;H2. We also observed a negative correlation between SOX4 expression and GSIS in a large collection of human islet preparations. There was also a positive correlation between SOX4 expression and STXBP6 (amisyn) expression and a tendency towards increased SOX4 expression in islets from organ donors with T2D. The second part of the thesis focuses on the role of the Krebs cycle enzyme fumarate hydratase (FH) in insulin release. Ablation of the Fh1 gene (which is initially implicated as a tumour suppressor in hereditary leiomyomatosis and renal cell cancer) in pancreatic &beta;-cells led to a complete loss of GSIS, as determined by ex vivo pancreatic perfusion studies, although this was not associated with any detectable abnormalities in [Ca²⁺]_i homeostasis, ATP production or glucose oxidation. The phenotype was rescued by the introduction of the human orthologue FH into the cytosol alone or in both the cytoplasm and mitochondria of Fh1 knockout (Fh1KO) mice, confirming the role of Fh1 in insulin secretion. Moreover, the addition of exogenous glutamate, previously implicated as a 2^nd messenger between glucose metabolism and insulin secretion, corrected the insulin secretory defect in Fh1^-/- &beta;-cells. We hypothesise that the loss of GSIS in Fh1KO mice results from enhanced anaplerosis, which is necessary to replenish Krebs cycle reactants. Consequently, this is followed by the depletion of the intracellular amino acid pool (including glutamate). Thus, our study demonstrates that the pancreatic Fh1KO mouse is a novel model of severe hyperglycaemia that harbours dysregulated metabolic features at the interface between both cancer and diabetes. The final study investigates the effect of ageing, a risk factor for T2D, on glucose-stimulated insulin and glucagon release. However, GSIS increased rather than decreased with ageing in both human and mouse islets (6 and 20 mmol/l glucose). Notably, ageing was not associated with reduced insulin content. Normal calcium homeostasis was observed in old (24-month-old) mice, demonstrating that the glucose sensing machinery was intact. In human islets, the inhibitory effect of glucose on glucagon secretion deteriorated with age. In the oldest group (>60 years of age), the inhibitory effect was completely abolished with 20 mmol/l glucose, while 6 mmol/l glucose only achieved less than 20% inhibition (which was not statistically significant). Our study reports the exciting possibility that hypersecretion of glucagon represents a link between senescence and increased diabetes risk.