Regulation of vascular endothelial growth factor expression by extra domain B segment of fibronectin in endothelial cells.

Diabetic retinopathy entails proliferation of vascular endothelial cells (ECs) and unregulated angiogenesis. We have previously shown that ECs increase the expression of an embryonic variant of fibronectin (FN), called extra domain-B FN (ED-B FN) in response to high glucose. We also showed that ED-B FN regulates EC tube morphogenesis, possibly through vascular endothelial growth factor (VEGF). In the present study, we have attempted to decipher the mechanisms by which ED-B FN may modulate EC phenotype. We hypothesized that ED-B FN regulates VEGF expression in ECs through interaction with selected integrin receptors. To test this hypothesis, we first cultured ECs in high levels of glucose to investigate for any alteration. We then used integrin-specific matrix mimetic peptides, neutralizing antibodies, and RNAi to identify the integrin(s) involved in VEGF expression. Finally, we used an animal model of diabetes to study whether these in vitro mechanisms also take place in the retina. Our results show that exposure of ECs to high levels of glucose increased VEGF expression. ED-B FN mediated this increase since knockdown of ED-B FN completely prevented glucose-induced VEGF expression. We then identified β1 integrin as the essential receptor involved in high glucose-induced VEGF expression. We also showed that diabetes increased β1 integrin and VEGF expression in the retina, which normalized upon ED-B knockdown. These findings showed that high levels of glucose in diabetes increased VEGF expression in ECs through ED-B FN and β1 integrin interaction. These results provide novel mechanistic basis of increased VEGF expression in diabetes.