Adiposity has been shown to secrete bioactive cytokines and growth factor known as adipocytokines, they can contribute to obesity, diabetes and complications of diabetes. Visfatin is a novel adipocytokine, and it was shown to exert insulin-mimetic effects in stimulating glucose transport and induced triglyceride accumulation in preadipocytes and triglyceride synthesis from gluvose. Visfatin plasma levels are increased in morbid obesity and type 2 diabetes mellitus. These finding indicate that visfatin may play a role in the association between visceral obesity and increased metabolic risk, visfatin gene suggested that genetic variation in the visfatin gene may, indeed, have a minor effect on visceral and subcutaneous visfatin messenger RNA expression profiles and parameters of glucose and insulin metabolism. In this study, we explored the relationships between the plasma level of visfatin and genetic single nucleotide polymorphisms (SNPs) of visfatin gene in type 2 diabetes mellitus (T2DM) with and without macrovascular disease. Plasma visfatin was found to be elevated significantly in T2DM with macrovascular disease patients. Moreover, waist to hip ratio was independently associated with plasma visfatin level. There were statistically significant differences in visfatin -948 G/T genetic variants distribution between T2DM with macrovascular disease and the T2DM control group. The visfatin -948 G/T heterozygotes showed higher mean high-density lipoprotein cholesterol than the carriers of the G allele. The results of the current study indicated that plasma visfatin levels were associated with macrovascular complications in type 2 diabetes. However, the definite roles of visfatin in the pathogenesis of insulin resistance, glucose and lipid metabolism are unclear. The observation of changes in the plasma concentrations of visfatin seen in T2DM and T2DM with macrovascular diseases may exert beneficial effects in understanding roles of visfatin in physiologic activity and metabolic disorder. Further studies are needed to elucidate the mechanisms behind visfatin overexpression in humans.

The prevalence of diabetes is increasing rapidly around the world. People with diabetes are 2–4 times more likely to die from cerebro and cardio-vascular causes than people with no history of diabetes, even after controlling for other risk factors. Atherosclerosis, the underlying cause of most cardiovascular disease (CVD), is accelerated in people with diabetes, but several clinical trials have questioned the efficacy of glucose lowering therapies. A better understanding of the molecular pathways by which diabetes accelerates atherosclerosis will expand the scope of current targets and strategies for more effective therapies. In this thesis we investigate a novel mechanism and establish and characterize new hyperglycemic mouse models for the study of diabetic atherosclerosis. Firstly, we investigate the effects of hyperglycemia on the vasa vasorum, the microvascular network that surrounds and supplies large vessels, and correlate those effects to the development of atherosclerosis. In normoglycemic ApoE-/- mice, the vasa vasorum expands as atherosclerotic lesions grow. However, in hyperglycemic ApoE-/- mice there is no significant neovascularization of the vasa vasorum despite the enhanced atherosclerotic development. We hypothesize that the ability of hyperglycemia to disrupt vasa vasorum neovascularization may promote the development and progression of atherosclerosis in diabetes. Secondly, we establish, characterize and manipulate a new model of hyperglycemia-induced atherosclerosis: the ApoE-/-:Ins2+/Akita mouse. We describe sex-specific differences of the ApoE-/-:Ins2+/Akita mouse model. Male ApoE-/-:Ins2+/Akita mice develop chronic hyperglycemia and accelerated atherosclerosis. Castration slows atherosclerosis in ApoE-/-:Ins2+/Akita mice but enhances it in normoglycemic controls. Female ApoE-/-:Ins2+/Akita mice are only transiently hyperglycemic but still present with accelerated atherosclerosis. Ovariectomized ApoE-/-:Ins2+/Akita mice are chronically hyperglycemic and show indications of advanced atherosclerosis. Lastly, we investigate the effects of a western-type diet on the hyperglycemic ApoE-/-:Ins2+/Akita mice. We demonstrate the pernicious phenotype of the mice leading to a significantly shortened lifespan correlated with massive atherosclerosis that extends to the aortic sinus, ascending and descending aorta, brachiocephalic artery and coronary arteries. In conclusion we provide insights for a new mechanism by which hyperglycemia may accelerate atherosclerosis and possible role of the vasa vasorum in the progression of atherosclerosis in hyperglycemic mice. We also establish new mouse models that illuminate the action of sex hormones on pancreatic beta-cell function and the vasculature. These models will provide a test bed to further study sex hormone effects, as well as the diabetic pathways that promote atherosclerosis. / Thesis / Doctor of Philosophy (PhD)

Objectives. To investigate diabetes prevalence, incidence, mortality trends, the effects of hyperglycaemia and blood pressure, diabetes and hypertension treatment, and the effect of screening detection on total and cardiovascular disease (CVD), myocardial infarction (MI) and stroke incidence. Study population and methods. Between 1972 and 2001 all patients with diabetes, some detected clinically and some by case-finding procedures (screening), were entered in a diabetes register at Laxå Primary Health Care Center in Sweden. The register included information on medical treatment and laboratory data as well as information on mortality and morbidity from National Registers. The register was supplemented with five non-diabetic subjects, matched to each diabetes patients by age, sex, and year of detection. Results. During the study period 776 new diabetes cases was found, 36 type 1 diabetes mellitus and 740 type 2 diabetes mellitus. Age standardised incidence and prevalence rates for type 1 and type 2 diabetes did not increase over time. Diabetic patients had 17% higher mortality rate than non-diabetic persons, 22% in women and 13% in men. The corresponding over-mortality in CVD was 33%, 41% in women and 27% in men. CVD mortality decreased across time in non-diabetic subjects and in diabetic men but not in diabetic women. Results regarding coronary heart disease (CHD) were similar. CVD incidence increased with fasting blood glucose (FBG), body mass index (BMI), mean arterial blood pressure (MABP), and decreased with metformin treatment and sulfonylurea. Myocardial infarction incidence increased with FBG, BMI and MABP, and decreased with metformin treatment. Stroke incidence increased with MABP. There was no difference in prognoses between those detected by screening or clinically. Conclusions. Diabetes prevalence and incidence did not change over time. The over-mortality according to diabetes was moderate. CVD and MI during follow up were negatively affected by hypertension and hyperglycaemia, and positively by pharmacological diabetic treatment. For stroke no pharmacological protective effect was seen. Screening did not improve prognosis.

Diabetic vasculopathy (DV) is the most important consequence of chronic hyperglycemia in patients with diabetes mellitus (DM). This thesis explores the interaction of blood, bone and kidney in the pathogenesis of DV by i) reviewing the current understanding of pathogenesis of macrovascular and microvascular diseases in DM to identify gaps in literature and generate hypotheses relating to various facets of DV ii) undertaking a series of prospective studies to examine these hypotheses iii) analysing the findings and integrating any new information obtained from the clinical studies into the current knowledge base and iv) generating hypotheses upon which future work might be based. The literature search was carried out with the aim of understanding current concepts of pathogenesis of DV and its potential modulators. The original reviews resulting from this process are presented in chapters 2 to 4. A series of pilot studies reported in chapters 7 to 11, were then carried out to interrogate hypotheses originating from this process. The first study was carried out in healthy individuals to define the biological variation of potential modulators of DV, namely erythropoietin (EPO), parathyroid hormone, 25 hydroxyvitamin D and 1, 25-dihydroxyvitamin D to facilitate the design and interpretation of subsequent studies. It revealed a wide biological variation of these modulators in the healthy population thus,emphasizing the need to have a control group in the subsequent study population. To examine whether tubulointerstitial dysfunction occurs before the onset of microalbuminuria, a measurement of the above mentioned parameters was carried out along with markers of tubulointerstitial injury in patients with type 1 and type 2 DM without microalbuminuria and in non-diabetic controls. It was found that tubulointerstitial dysfunction with low levels of EPO and 1, 25-dihydroxyvitamin D and higher excretion of tubular injury markers, occurs before the onset of microalbuminuria. Subsequently, diabetic and nondiabetic chronic kidney disease (CKD) patients with EPO deficiency anaemia were examined to study the effects of EPO therapy on the excretion of tubular injury markers. However, in these patient groups, we were unable to demonstrate an effect of EPO therapy on the markers of tubular injury in spite of a beneficial haematological response. To examine whether vascular calcification (VC) and bone mineral density (BMD) were linked in patients with diabetes mellitus and to explore their relationship to modulators of DV, an assessment of VC and BMD was undertaken in patients with type 2 DM with different degrees of proteinuria and normoalbuminuria. VC was assessed by CT scan and BMD by a DEXA scan. Modulators of DV were measured including serum Osteoprotegerin (OPG) and receptor activator of nuclear factor kappa-b-ligand (RANKL). The findings were i) a high prevalence of VC and osteopenia in normoalbuminuric type 2 DM patients with normal serum creatinine ii) a weak inverse relationship between VC and osteopenia iii) proteinuric patients had worse VC but not osteopenia iv) weak relationships between OPG levels and both VC and osteopenia, masked by age in multivariate analysis. The final study examined the relationship between modulators of DV, including OPG and RANKL, and the degree of CKD. It was found that abnormalities of OPG and RANKL occur before the onset of microalbuminuria and progress with deterioration of renal function. Compared to nondiabetics, DM patients have higher OPG levels in the predialysis phase and lower levels in haemodialysis phase, a phenomenon that might indicate endothelial exhaustion in dialysis patients with DM. The derangements associated with DV seem to occur earlier than previously thought. Further work is required to untangle these complexities and to define the contribution of factors such as the adverse blood milieu, the vasculature, abnormal bone and mineral metabolism, and early tubulointerstitial damage. The findings from the studies reported here may help in the formulation of new hypotheses, which might contribute to future work in this area.