Extra domain-B containing fibronectin (EDB(+) FN), a recently proposed marker of angiogenesis, has been shown to be expressed in a number of human cancers and in ocular neovascularization in patients with proliferative diabetic retinopathy. To gain molecular understanding of the functional significance of EDB(+) FN, we have investigated possible regulatory mechanisms of induction and its role in endothelial cell proliferation and angiogenesis. Human vascular endothelial cells were cultured in high levels of glucose, and fibrogenic growth factors, transforming growth factor-beta1 (TGF-beta1) and endothelin-1 (ET-1). Our results show that high glucose levels, TGF-beta1, and ET-1 upregulated EDB(+) FN expression. Treatment of cells exposed to high glucose with TGF-beta1 neutralizing antibody and ET receptor antagonist prevented high glucose-induced EDB(+) FN expression. In order to elucidate the functional significance of EDB(+) FN upregulation, cells were subjected to in vitro proliferation and angiogenesis assays following EDB peptide treatment and specific EDB(+) FN gene silencing. Our results show that exposure of cells to EDB peptide increased vascular endothelial growth factor (VEGF) expression, endothelial proliferation, and tube formation. Furthermore, specific EDB(+) FN gene silencing prevented both basal and high glucose-induced VEGF expression and reduced the proliferative capacity of endothelial cells. In conclusion, these results indicate that EDB(+) FN is involved in endothelial cell proliferation and vascular morphogenesis, findings which may provide novel avenues for the development of anti-angiogenic therapies.

These results indicate an important contribution of both ET-1 and NHE-1 in the pathogenesis of diabetic cardiomyopathy. These data suggest that NHE-1 may act as a downstream mediator in the production of ET-induced functional and structural changes in the myocardium in diabetes.

These results indicate that diabetes-induced vascular hypertrophy and remodeling is associated with reexpression of embryonic forms of FN and myosin heavy chain. Such changes are ET-dependent and may be mediated via TGF-beta1 and angiotensin.

Aims/hypothesis: Hyperglycaemia-induced oxidative stress is implicated in the pathogenesis of chronic diabetic complications. Glucose-mediated oxidation of LDL may result in increased oxidative stress and vascular endothelial cell dysfunction via interaction with a cell surface scavenger receptor, CD36. In this study, we investigated the role of CD36 in cultured microvascular endothelial cells (MVECs) and in the heart by using an animal model of chronic diabetes. Methods: Cultured MVECs were subjected to varying glucose concentrations and assayed for alteration in CD36 gene expression and protein levels. To assess for oxidised LDL (ox-LDL) uptake, MVECs exposed to low and high glucose were treated with ox-LDL (80 μg/ml), a ligand for CD36. Haem oxygenase-1 (HO-1) and endothelin-1 (ET-1) induction, as well as oxidative stress were determined. The role of glucose-induced CD36 alteration in ox-LDL uptake was also assayed following post-transcriptional CD36 gene silencing. For in vivo studies, CD36 mRNA and oxidative DNA and protein damage were measured in heart tissues of 1-month-old diabetic Sprague-Dawley rats. Results: We found that glucose increased CD36 mRNA and protein levels in MVECs. High levels of glucose also augmented ox-LDL uptake, in association with increasing HO-1 and ET-1 mRNA levels. CD36 gene silencing prevented glucose-induced CD36 alteration, reduced ox-LDL uptake, and prevented HO-1 and ET-1 up-regulation. Similar to in vitro studies, diabetic heart tissues exhibited increased CD36 mRNA levels and increased oxidative DNA and protein damage. Conclusions/interpretation: Our results provide evidence that up-regulation of CD36 may have a role in increasing oxidative stress in MVECs and the heart in chronic diabetes.

No abstract