Active NF-E2-related factor (Nrf2) contributes to keep endothelial NO synthase (eNOS) in the coupled state: role of reactive oxygen species (ROS), eNOS, and heme oxygenase (HO-1) levels

J Biol Chem. 2009 Nov 13;284(46):31579-86. doi: 10.1074/jbc.M109.009175. Epub 2009 Sep 21.


The aim of our study was to examine in detail the impact of NF-E2-related factor (Nrf2) activation on endothelial cell function with focus on redox homeostasis and the endothelial nitric oxide synthase (eNOS) system. We administered 2-cyano-3,12-dioxooleana-1,9-dien-28-oic imidazolide (CDDO-IM), a known activator of Nrf2, to primary human umbilical vein endothelial cells. Activation of Nrf2 by CDDO-IM increased the amount of bioavailable nitric oxide (NO), a major contributor to vascular homeostasis, in naive and stressed cells. Concomitantly, intracellular reactive oxygen species were dose-and time-dependently reduced. In apparent contrast to elevated NO levels, eNOS protein expression was transiently decreased in an Nrf2-dependent manner. Employing pharmacological inhibitors as well as a small interfering RNA approach, we identified de novo protein synthesis of heme oxygenase 1 (HO-1) and its enzymatic activity as cause for the observed reduction of eNOS. We hypothesize that under redox stress, when the availability of tetrahydrobiopterin, a pivotal stoichiometric cofactor for eNOS, is limited, activation of Nrf2 leads (a) to transient reduction of eNOS protein levels and (b) to an antioxidant defense in human umbilical vein endothelial cells. Both activities ensure that a stoichiometric ratio of eNOS and tetrahydrobiopterin is sustained and that the risk of eNOS uncoupling is reduced. Our study is the first to provide a causal link between Nrf2 activation and eNOS expression and NO levels, respectively.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antioxidants / metabolism
  • Biopterin / analogs & derivatives
  • Biopterin / metabolism
  • Cells, Cultured
  • Endothelium, Vascular / cytology
  • Endothelium, Vascular / drug effects
  • Endothelium, Vascular / metabolism
  • Heme Oxygenase-1 / antagonists & inhibitors
  • Heme Oxygenase-1 / metabolism*
  • Humans
  • Imidazoles / pharmacology
  • Immunoblotting
  • NF-E2-Related Factor 2 / antagonists & inhibitors
  • NF-E2-Related Factor 2 / physiology*
  • Nitric Oxide / metabolism
  • Nitric Oxide Synthase Type III / antagonists & inhibitors
  • Nitric Oxide Synthase Type III / metabolism*
  • Oleanolic Acid / analogs & derivatives
  • Oleanolic Acid / pharmacology
  • Oxidation-Reduction
  • Reactive Oxygen Species / metabolism*
  • Umbilical Veins / cytology
  • Umbilical Veins / drug effects
  • Umbilical Veins / metabolism


  • 1-(2-cyano-3,12-dioxooleana-1,9-dien-28-oyl) imidazole
  • Antioxidants
  • Imidazoles
  • NF-E2-Related Factor 2
  • NFE2L2 protein, human
  • Reactive Oxygen Species
  • Biopterin
  • Nitric Oxide
  • Oleanolic Acid
  • NOS3 protein, human
  • Nitric Oxide Synthase Type III
  • Heme Oxygenase-1
  • sapropterin