Complex changes in the mechanical, biochemical, structural and electrical properties of the heart, which may be responsible for the development of an early diastolic dysfunction and increased incidence of cardiac arrhythmias in diabetic patients, are characteristic features of diabetic cardiomyopathy. In spite of the accumulating knowledge obtained from different models of diabetes, the mechanism of diabetic cardiac dysfunction still remains elusive [1
The current study demonstrates that STZ-induced diabetes in mice is associated with a marked depression of both systolic and diastolic function of the left ventricle. These results are consistent with earlier reports showing depressed cardiac function in different mouse [3
] models of type 1 diabetes and compromised cardiac conductivity and/or performance reported in diabetic patients. The results presented here document that in STZ-induced mouse model of type 1 insulin-dependent diabetes, the impaired cardiac function is associated with increased myocardial XO activity, oxidative/nitrosative stress and fibrosis, which can be attenuated by pharmacological inhibition of the XO by ALP.
Multitude of experimental and clinical studies have suggested that increased sympathetic activity, activated cardiac reninangiotensin system, myocardial ischaemia/functional hypoxia and elevated circulating levels of glucose result in oxidative/nitrosative stress and inflammation, eventually culminating in cellular dysfunction and death in cardiomyocytes and endothelial cells (both apoptotic and necrotic) promoting increased remodelling and fibrosis, processes which play a critical role in the development of subsequent diabetic cardiomyopathy [3
]. Furthermore, increased oxidative/nitrosative stress may also lead to inactivation of key cardiac ion channels by oxidation, nitration and/or nitrosylation, thereby contributing to impaired repolarization reported in both diabetic animals as well as in human beings with type 1 diabetes.
Hyperglycaemia triggers oxidative stress via
numerous mechanisms involving activation of the polyol pathway, glucose auto-oxidation, alterations of cellular redox state, increased formation of diacylglycerol and the subsequent activation of protein kinase C, and accelerated nonenzymatic formation of advanced glycation end products. Superoxide anion appears to play a particularly important role in the pathogenesis of diabetic cardiovascular dysfunction, and this reactive oxidant was reported to activate many of the above-mentioned pathways [6
]. The cellular sources of superoxide in anion in diabetes are multiple and may include NAD(P)H and XO, the mitochondrial respiratory chain among many others [19
]. Recent studies suggest that XO may play an important role in the generation of free radicals in diabetes. There is an elevation in the plasma and liver XO levels in type 1 diabetic patients [32
] and increased endothelial superoxide formation in aorta from alloxan-induced diabetic rabbits can be blocked by the XO inhibitor ALP [32
]. Diabetes also causes an increase of XO activity in the liver of rats, and XO is released from the liver of these animals [32
]. Increased plasma XO activity in diabetic mice correlates with the degree of superoxide generation 2 weeks after the onset of diabetes, and can be normalized by pre-treatment with XO inhibitors ALP or oxypurinol [33
]. In type 1 diabetic patients, ALP treatment attenuated the degree of oxidative stress (haemoglobin glycation, glutathione oxidation and lipid peroxidation) [32
], whereas in type 2 diabetic patients with mild hypertension, prolonged treatment with ALP resulted in significant improvements in peripheral endothelium-dependent vasorelaxant function [34
]. Consistently with these results, we found increased XO activity in the liver, myocardium and serum of diabetic mice, which could be normalized by ALP treatment. ALP treatment also attenuated the myocardial ROS generation in our mouse model of diabetes. In contrast, ALP had no effects on diabetes-induced increased expression of various isoforms of NAD(P)H oxidases.
Through the activation of NF-kB hyperglycaemia-induces increased ROS generation favours increased expression of iNOS, which can increase the generation of NO in diabetic hearts [28
]. Superoxide anion interacts with nitric oxide, forming the oxidant peroxynitrite (ONOO−
), which attacks various biomolecules, leading to—among other things—the production of a modified amino acid, nitrotyrosine [28
]. Small amount of peroxynitrite may be generated in normal cells and may play various physiological regulatory functions (e.g.
in signalling processes; for reviews see [36
]). However, several lines of evidence support the pathogenetic role of excessive endogenous peroxynitrite formation in diabetic cardiovascular [6
] and other complications [41
] both in experimental animals and in human beings. For example, the degree of cell death and/or dysfunction correlates with levels of NT in endothelial cells, cardiomyocytes and fibroblasts from myocardial biopsies of diabetic patients [9
], hearts of experimental diabetic rats or mice [3
] and hearts perfused with high glucose concentrations [5
]. The NT immunoreactivity is increased in the microvasculature of type 2 diabetic patients and correlates with fasting blood glucose, HbA1c, intracellular adhesion molecule, vascular cellular adhesion molecule and endothelial dysfunction [42
]. Peroxynitrite has been reported to attack various biomolecules, leading to compromised cardiovascular function and cell death (both apoptotic and necrotic) in hearts and other tissues via
multiple complex mechanisms [6
]. One of these pathways involves DNA strand breakage and consequent activation of the nuclear enzyme PARP, which emerges as a major effector pathway in the development of various interrelated diabetic complications [8
], including cardiomyopathy [7
]. Consistently with the importance of these pathways in the development of diabetic cardiomyopathy, we found increased iNOS (but not eNOS) expression, NT generation, apoptosis and PARP activity in diabetic heart accompanied by increased fibrosis. Remarkably, ALP treatment attenuated not only the oxidative/nitrosative stress-PARP pathway, apoptosis but also fibrosis in diabetic hearts.
Based on the results of the current study, we conclude that the XO-derived superoxide production contributes to the development of diabetic cardiomyopathy, and XO inhibition with ALP improves diabetes-induced cardiac dysfunction by decreasing oxidative/nitrosative stress and the activation of its downstream effector pathways (e.g.
PARP) and fibrosis. This coupled with the beneficial effects of ALP in various animal models of heart failure, also observed in small-scale human clinical trials (reviewed in [20
]), is particularly encouraging from the therapeutic point of view forecasting multiple possible benefits of ALP treatment in diabetic patients.