High glucose (HG) induces oxidative injury to cultured human umbilical vein endothelial cells (HUVECs). Recent studies have discovered 4-octyl itaconate (OI) as a novel and cell permeable Nrf2 (nuclear-factor-E2-related factor 2) activator. Its potential activity in HG-treated HUVECs was tested here. In HUVEC5 01 disrupted Keapl-Nrf2 association, causing Nrf2 protein accumulation and nuclear translocation, as well as transcription and expression of Nrf2-ARE-dependent genes, including HO1, NQO1 and GCLM. Significantly, pretreatment with 01 potently inhibited HG (40 mM glucose)-induced death and apoptosis of HUVECs. Moreover, 01 potently inhibited HG-induced reactive oxygen species (ROS) production, lipid peroxidation, superoxide accumulation and mitochondrial depolarization in HUVECs. Activation of Nrf2 is required for 01-induced cytoprotection in HUVEC5. Nrf2 shRNA or knockout (by CRISPR/Cas9 method) reversed 01-mediated HUVEC protection against HG. Further studies showed that Keapl silencing or Cys151S mutation mimicked and nullified 01-induced activity in HUVECs. Taken together, we conclude that 01 activates Keapl-Nrf2 signaling to protect HUVEC5 from HG. (C) 2018 Elsevier Inc. All rights reserved.