Abstract 13280: Localized Hydrogen Peroxide Production and Differential Activation of Redox-sensitive Transcription Factors Involved in Atherogenesis
Reactive oxygen species modulate key physiological responses, yet ROS are also implicated in vascular disease states. At low concentrations, the stable ROS hydrogen peroxide (H2O2) activates eNOS and enhances vascular barrier function. Yet at higher concentrations, vascular ROS lead to oxidative stress, increase permeability and promote atherosclerosis. The molecular events underlying the transition from “physiological” to “pathological” ROS in the endothelium are incompletely understood. Laminar shear stress activates “atheroprotective” redox-modulated transcription factors whereas turbulent/oscillatory flow activates “atherogenic” transcription factors that are sensitive to cellular redox state. In these studies, we constructed differentially-targeted HyPer biosensors for H2O2 to identify the intracellular organelles and pathways responsible for dynamic mechanochemical regulation of endothelial H2O2 metabolism. We discovered that laminar (physiological) shear stress increases H2O2 in the endothelial cell nucleus much more than in the cytosol. By contrast, oscillatory (pathological) shear increases H2O2 in the cytosol significantly more than in the nucleus. We next generated H2O2 in specific subcellular locales in endothelial cells using recombinant constructs expressing a D-amino acid oxidase (DAAO) that robustly generates H2O2 upon addition of D-alanine to the cells. By transfecting a series of differentially targeted DAAO constructs into endothelial cells, we discovered that generation of H2O2 in distinct subcellular compartments differentially modulates transcriptional activation. Generation of H2O2 by DAAO expressed in the endothelial cell nucleus led to enhanced transcription of Nrf2-modulated genes, whereas generation of H2O2 by DAAO targeted to the cytosol instead activated NF-κB-regulated genes. This differential transcriptional response regulated by H2O2 generated in distinct subcellular locales provides new insights into the roles of ROS and eNOS in mechanochemical coupling. Taken together, these findings have important implications for our understanding of flow-responsive genes in the normal blood vessel as well as in disease states associated with disordered blood flow and oxidative stress.
Author Disclosures: B. Steinhorn: None. V. Belousov: None. T. Michel: Honoraria; Modest; Pfizer. Consultant/Advisory Board; Modest; Ionis Pharmaceuticals.
- © 2016 by American Heart Association, Inc.