Cellular stress conditions are reflected in the protein and RNA content of endothelial cell‐derived exosomes

TLDR

The vascular endothelium adapts to stress signals such as hypoxia and inflammation, and exosomes—carrying cell‑derived RNA and proteins—serve as key mediators of intercellular communication whose cargo reflects the donor cell’s condition and may protect target cells from stress. The study aimed to determine whether endothelial exosome RNA and protein profiles mirror cellular stress induced by hypoxia, inflammation, or hyperglycemia. Endothelial cells were subjected to hypoxia, TNF‑α activation, high glucose, and mannose, and exosome RNA and protein contents were profiled using microarray and quantitative proteomics, revealing 1,354 proteins and 1,992 mRNAs. The analysis revealed stress‑dependent changes in exosome RNA and protein, with hypoxia and TNF‑α activation markedly altering cargo, whereas high glucose had only minor effects on protein composition and no impact on RNA.

Abstract

The healthy vascular endothelium, which forms the barrier between blood and the surrounding tissues, is known to efficiently respond to stress signals like hypoxia and inflammation by adaptation of cellular physiology and the secretion of (soluble) growth factors and cytokines. Exosomes are potent mediators of intercellular communication. Their content consists of RNA and proteins from the cell of origin, and thus depends on the condition of these cells at the time of exosome biogenesis. It has been suggested that exosomes protect their target cells from cellular stress through the transfer of RNA and proteins. We hypothesized that endothelium-derived exosomes are involved in the endothelial response to cellular stress, and that exosome RNA and protein content reflect the effects of cellular stress induced by hypoxia, inflammation or hyperglycemia.We exposed cultured endothelial cells to different types of cellular stress (hypoxia, TNF-α-induced activation, high glucose and mannose concentrations) and compared mRNA and protein content of exosomes produced by these cells by microarray analysis and a quantitative proteomics approach.We identified 1,354 proteins and 1,992 mRNAs in endothelial cell-derived exosomes. Several proteins and mRNAs showed altered abundances after exposure of their producing cells to cellular stress, which were confirmed by immunoblot or qPCR analysis.Our data show that hypoxia and endothelial activation are reflected in RNA and protein exosome composition, and that exposure to high sugar concentrations alters exosome protein composition only to a minor extend, and does not affect exosome RNA composition.