MicroRNA-9 rescues hyperglycemia-induced endothelial cell dysfunction and promotes arteriogenesis through downregulating Notch1 signaling.

Abstract

Hyperglycemia-induced endothelial dysfunction plays a major role in the pathogenesis of diabetic vascular complications. MicroRNAs are potential therapeutic agents to improve hyperglycemia-induced endothelial dysfunction. This study examined the relationship of miR-9 with Notch1 signaling in hyperglycemia-induced endothelial dysfunction. Human umbilical vein endothelial cells (HUVECs) were exposed to 30mM glucose concentration. Cell viability including proliferation, adhesion, migration and tube formation was significantly impaired. Quantitative real time polymerase chain reaction (qRT-PCR) or Western blot demonstrated that miR-9 expression remarkably decreased and expression of Notch1 and its effectors (Hes1, Hey1, Hey2) were upregulated. Transfection with miR-9 improved cell function, inhibited mRNA and protein expression of Notch1 and its effectors. Although basal expression of the arterial endothelium biomarker Ephrin B2 was almost undetectable in HUVECs, double-label immunofluorescence revealed that transfection with miR-9 upregulated Ephrin B2 expression. By contrast, such protective effects of miR-9 overexpression were eliminated due to use of miR-9 inhibitor. Dual luciferase assay further confirmed a significant inverse correlation between miR-9 and Notch1. In addition, Notch1 overactiviation was mimicked in HUVECs by transfecting with Notch1 intracellular domain (NICD1). MiR-9 significantly inhibited NICD1 mRNA expression and alleviated hyperglycemia-induced injury of the NICD1-overexpressing cells. Taken together, our data support upregulating miR-9 expression as a potential therapeutic strategy to antagonize hyperglycemia-induced injury by inhibiting Notch1 signaling.