Reactive Oxygen Species Differentially Modulate the Metabolic and Transcriptomic Response of Endothelial Cells

Abstract

Reactive oxygen species (ROS) are important mediators of both physiological and pathophysiological signal transduction in the cardiovascular system. The effects of ROS on cellular processes depend on the concentration, localization, and duration of exposure. Cellular stress response mechanisms have evolved to mitigate the negative effects of acute oxidative stress. In this study, we investigate the short-term and long-term metabolic and transcriptomic response of human umbilical vein endothelial cells (HUVEC) to different types and concentrations of ROS. To generate intracellular H2O2, we utilized a lentiviral chemogenetic approach for overexpression of human D-amino acid oxidase (DAO). DAO converts D-amino acids into their corresponding imino acids and H2O2. HUVEC stably overexpressing DAO (DAO-HUVEC) were exposed to D-alanine (3 mM), exogenous H2O2 (10 µM or 300 µM), or menadione (5 µM) for various timepoints and subjected to global untargeted metabolomics (LC-MS/MS) and RNAseq by MACE (Massive analysis of cDNA ends). A total of 300 µM H2O2 led to pronounced changes on both the metabolic and transcriptomic level. In particular, metabolites linked to redox homeostasis, energy-generating pathways, and nucleotide metabolism were significantly altered. Furthermore, 300 µM H2O2 affected genes related to the p53 pathway and cell cycle. In comparison, the effects of menadione and DAO-derived H2O2 mainly occurred at gene expression level. Collectively, all types of ROS led to subtle changes in the expression of ribosomal genes. Our results show that different types and concentration of ROS lead to a different metabolic and transcriptomic response in endothelial cells.

Insights into H2O2-induced signaling in Jurkat cells from analysis of gene expression

“…Treatment of PIG1 cells with 100 μM H 2 O 2 resulted in only 661 DEG after 6 h (26), but no analysis was presented on the functional classes of these genes. In HUVEC treated with 10 μM H 2 O 2 , no DEG were identified up to 900 min post-treatment, although 300 μM H 2 O 2 resulted in maximum DEG (3540) at 4.5 h (29). In contrast we identified 6803 DEG in Jurkat cells treated with a 10 μM bolus dose of H 2 O 2 .…”

“…There is an abundance of literature implicating H 2 O 2 in cellular signalling, predominantly focussing on investigating specific signalling targets. In addition, microarrays and RNA-Seq have been used to provide a global overview of the transcriptomics response of budding and fission yeasts, and cultured human cells, to H 2 O 2 (16–29). Collectively these studies support a role for H 2 O 2 in controlling the expression of a range of genes, including p53 target genes, apoptosis genes and signal transduction genes.…”

The influence of redox modulation on hypoxic endothelial cell metabolic and proteomic profiles through a small thiol‐based compound tuning glutathione and thioredoxin systems

“…Endothelial cells (ECs) are ubiquitously present across the vascular beds in the body. They represent the interface through which blood and neighboring tissue communicate via cytokine signaling, metabolites, and the exchange of gas and nutrients 1 . Their properties are tightly linked to their location due to physical and biochemical changes in the microenvironment 2,3 .…”

CircANKRD12 Is Induced in Endothelial Cell Response to Oxidative Stress

“…As redox imbalance elicits EC dysfunction, it also triggers specific redox-sensitive gene expression changes, both transcriptional and posttranscriptional [ 3 , 4 , 5 , 6 ]. Important hubs of cell response to ROS and oxidative stress are Forkhead Box O1 (FOXO1) and NFE2-like bZIP transcription factor 2 (NRF2), which activate the antioxidant defences, Nuclear Factor Kappa B Subunit 1 (NFkB), which induces the expression of pro-inflammatory cytokines, and p53, which coordinates a complex antiproliferative and pro-apoptotic transcriptional response [ 3 , 7 ].…”