Protein Modification by O-Linked GlcNAc Reduces Angiogenesis by Inhibiting Akt Activity in Endothelial Cells

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Abstract
Objective—Glucose flux through the hexosamine biosynthesis pathway (HBP) has been implicated in the development of diabetic vascular complications. O-linked N-acetylglucosamine (O-GlcNAc) modification on protein is the major mechanism mediating the actions of the HBP. Impaired angiogenesis is well-recognized in diabetes; however, the mechanisms are not completely defined. Here, we investigated the role of protein O-GlcNAc modification in angiogenesis.Methods and Results—In a mouse aortic ring assay, elevated O-GlcNAc levels induced by high-fat diet, streptozotocin-induced diabetes, or in vitro glucosamine treatment were associated with impaired angiogenesis. In cultured human umbilical vein endothelial cells and EA.hy926 endothelial cells, glucosamine increased protein O-GlcNAc modification and inhibited cell migration and capillary-like structure formation. Conversely, removal of O-GlcNAc by adenoviral-mediated overexpression of O-GlcNAcase improved these steps of angiogenesis. Also, high concentrations of glucose reduced capillary-like structure formation of human umbilical vein endothelial cells. Akt was recognized by an O-GlcNAc specific lectin, and glucosamine increased the amounts of Akt protein in these lectin precipitates. Increased glycosylation paralleled reduced Akt activity in endothelial cells.Conclusion—These results suggest that elevated protein O-GlcNAc modification through the HBP impairs angiogenesis in endothelial cells, possibly by inhibiting Akt signaling.