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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 M or 300 M), or menadione (5 M) 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 M 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 M 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.
