High glucose (HG) induces vascular endothelial cell injury. However, the underlying mechanisms are poorly understood. Circular RNA HIPK3 (circHIPK3) is a highly conserved non-coding RNA. Here we show that circHIPK3 is downregulated in HG-treated human umbilical vein endothelial cells (HUVECs) and in primary aortic endothelial cells (HAECs) from diabetic patients. In both HUVECs and HAECs, lentivirus-mediated circHIPK3 overexpression inhibited HG-induced cell death and apoptosis. Contrarily, circHIPK3 silencing by targeted siRNA exacerbated HG-induced endothelial cell death and apoptosis. Further, circHIPK3 downregulation by HG caused microRNA-124 (miR-124) accumulation in HUVECs and HAECs. On the contrary, miR-124 inhibition by the adeno-associated virus (AAV)-packed miR-124 inhibitor protected endothelial cells from HG. Together, circHIPK3 downregulation mediates HG-induced endothelial cell injury. Targeting circHIPK3-miR-124 pathway could potentially be a novel approach for the treatment of diabetic-associated vascular injury. (C) 2018 Elsevier Inc. All rights reserved.