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ItemContractile dysfunction of the heart in early diabetesWaddingham, Mark Thomas ( 2016)The prevalence of heart failure, especially heart failure with preserved ejection (HFpEF), is increasing annually in part due to an ageing population and the dramatically increased incidence of obesity, insulin resistance, prediabetes and diabetes worldwide. It is well established that patients with type-1 diabetes (T1DM) and type-2 diabetes (T2DM) are at a significantly increased risk of developing HFpEF. HFpEF is a progressive condition and its earliest manifestations are subtly impaired myocardial function that is termed diabetic cardiomyopathy (DCM). At present, there is only a limited understanding of the underlying pathophysiological mechanisms that drive the development of DCM and eventual HFpEF in early T1DM and T2DM. Therefore, the aim of this thesis was to further explore mechanisms that could drive the development of DCM and HFpEF using rat models of early T1DM and T2DM. Using synchrotron radiation as a source for small angle x-ray scattering (SAXS) in the in situ beating rat heart, we are able to measure actin-myosin cross-bridge (CB) dynamics in the entire cardiac cycle, in real time. In the first part of this thesis, we were able to demonstrate that chronic inhibition of the RhoA/Rho-kinase (ROCK) pathway with fasudil improved regionally impaired diastolic myosin head extension and depressed systolic mass transfer in the myocardium of rats with early T1DM. Further, we were also able to demonstrate that global left ventricular (LV) systolic performance was significantly improved in diabetic rats treated with fasudil. These results suggest that the activation of the ROCK pathway is involved in the development of early DCM in the context of T1DM. The Goto-Kakizaki (GK) rat is a non-obese model of spontaneous T2DM, which makes it a useful model to examine the effects of early T2DM on the myocardium without the added complication of obesity. Utilising the synchrotron radiation SAXS technique in the in situ beating heart preparation, we are able to demonstrate that young GK rats (10-12 weeks old) with early T2DM (prediabetes) exhibit impaired basal diastolic myosin head extension and reduced systolic myosin mass transfer in the deeper myocardial layer, the subendocardium. Interestingly, basal global cardiac function and β-adrenergic mediated positive inotropy was preserved in young GK rats. We speculate that a combination of cardiomyocyte hypertrophy and enhanced epicardial fibre function are the most likely mechanisms for the preserved global LV function in young GK rats. Diabetes is rarely seen in isolation in patients and commonly coexists with hypertension. The interaction of diabetes and hypertension is known to exacerbate myocardial dysfunction and accelerate the development of HFpEF, but the precise mechanisms remain elusive. The GK rat is a model of salt-sensitive hypertension induced by exposure to a high-salt diet (6% NaCl) for eight weeks. Thus, we examined if the interaction of prediabetes and salt-sensitive hypertension exacerbated myocardial dysfunction and accelerated the development of HFpEF in young GK rats. In GK rats exposed to a HS diet, we observed subtle declines in basal global diastolic and systolic LV function. Limited contractile reserve is a key feature of clinical HFpEF. Consistent with this, we were able to show that regional contractile reserve was limited at the fibre-level in the subepicardial and subendocardial fibre layers of the myocardium in GK rats maintained on a HS diet. These results suggest that limited contractile reserve at the fibre-level may be an early manifestation of HFpEF. In summary, this thesis has demonstrated that the ROCK pathway is involved in the evolution of DCM in T1DM, possibly by modulating actin-myosin interactions in the cardiac cycle. Our results also indicate that impaired CB dynamics is a feature of early T2DM DCM, although global cardiac function is preserved. Importantly, we have demonstrated that limited contractile reserve at the fibre-level may be an early manifestation of HFpEF in the presence of early T2DM and hypertension. Although further work is required to identify the specific molecular mechanisms that drive the impaired actin-myosin CB dynamics in early T1DM and T2DM, this thesis provides novel information of the pathophysiological features of contractile dysfunction of the heart in early diabetes.
ItemThe role of the coronary vasculature and myocardium in the pathogenesis of diabetic cardiomyopathyJenkins, Mathew James ( 2012)The prevalence of diabetes is increasing worldwide. This poses a significant threat to human health, as diabetes is associated with an increased risk of mortality due to cardiovascular disease. In particular, diabetic patients develop diabetic cardiomyopathy (DCM), characterised by impaired cardiac muscle contraction and relaxation, leading to left ventricle (LV) muscle stiffness and congestive heart failure. Previous studies suggest that changes in the coronary vasculature and cardiac subcellular function may account for the progression to DCM, however as yet this has not been assessed in vivo. Synchrotron radiation (SR) now makes possible novel imaging and diffraction techniques, to investigate the role these mechanisms play in the early development of DCM, where clinical intervention is most efficacious. To assess coronary function in vivo we validated the use of SR imaging to detect and quantify regional differences in resistance microvessel calibre. In type 1 diabetic rodents we found that although endothelium-dependent and –independent vasodilatory responses in individual coronary vessels are preserved, following inhibition of NO and PGI2 production, there is evidence of localised focal and segmental constrictions. This demonstrates, for the first time, localised coronary microvascular endothelial dysfunction in early-stage type 1 diabets (T1D). Contributing to this diabetic coronary impairment is the RhoA/Rho-kinase (ROCK) pathway, which had previously been shown to play a role in endothelial dysfunction and coronary vasospasm. Our data further support a role for ROCK in early diabetic coronary dysfunction, as following nitric oxide synthase/cyclooxygenase blockade, ROCK inhibition greatly reduced regional segmental constrictions and completely alleviated persistent focal stenoses in diabetic animals. Together, these findings provide strong evidence that early vascular dysfunction may contribute to the development of DCM. In addition, although characterised by global cardiac impairment, the role subcellular changes in the sarcomere play in DCM progression is not known. SR, as a source for small-angle X-ray diffraction, allows the assessment of cardiomyocyte cross-bridge dynamics (CB) and myosin interfilament lattice spacing in situ and in real time. Using SR, our data shows that in early T1D, CB dynamics are abnormal in the beating hearts and this is directly related to impaired LV function. The change in CB dynamics is caused by myosin head displacement from actin filaments, but notably is not related to estimated sarcomere length or myofilament order. SR X-ray diffraction thus provides a robust method to assess cardiac CB dynamics in situ and for the first time we provide evidence that impairment in the regulation of myosin head extension in T1D hearts contributes to DCM. Currently 85-90% of diabetics have T2D and it is therefore critical that these coronary microvascular and cardiac subcellular impairments in T1D are explored in T2D. As such, rodent models which account for the environmental factors important in the human development of DCM are required. We conducted a comprehensive characterisation of cardiac function and structure in diet-induced rodent models of obesity, insulin resistance and T2D, and uncovered mild systolic dysfunction in fructose fed and mild diastolic dysfunction in high fat fed rodents. Furthermore, we demonstrated mild contractile dysfunction in high fat fed low dose streptozotocin rodents. The characterisation of only mild cardiac dysfunction, in spite of the lengthy time course used, suggests further refinement is required to achieve more robust DCM models. In summary, through the validation of novel SR imaging and diffraction techniques our data has confirmed a role for coronary microvascular dysfunction, via the ROCK pathway and cardiac subcellular impairment, via reduced myosin head extension, in the development of DCM. In addition, further studies investigating rodent models of T2D and DCM are required. These findings provide a strong basis for the future development of novel therapies aimed at preventing and/or reversing the decline in cardiac function associated with diabetes.