Human coronary endothelial cell activation by endotoxin is characterized by NF-κB activation and TNF-α synthesis

Emily L. Chan, Sandra B. Haudek, Brett P. Giroir, Joseph T. Murphy

Research output: Contribution to journalArticle

25 Scopus citations

Abstract

Tumor necrosis factor-α (TNF-α), an early inflammatory mediator typically regulated by nuclear factor kappa B (NF-κB), plays a critical role in the development of cardiovascular dysfunction in sepsis. While several myocardial cell types synthesize TNF-α, the importance of the myocardial endothelium in sepsis-related cardiac cytokine production is unclear. To determine the role of the human coronary artery endothelial cell (HCA-EC) in the cytokine response to endotoxin we measured in vitro TNF-α synthesis, TNF-α mRNA, and the associated NF-κB response to LPS. To determine the magnitude of the HCA-EC response we assessed the TNF-α and NF-κB response to LPS in a human monocytic cell line (THP-1) as well. We observed an increase in supernatant TNF-α from LPS-stimulated HCA-EC (12 h) that was ablated by the proteosome inhibitor, ALLN (N-acetyl-Leu-Leu-norleucinal). Similarly, ALLN-sensitive TNF-α was produced by monocytes following LPS, although at concentrations 100-fold higher than HCA-EC. TNF-α mRNA from HCA-EC was detected at 60 min in LPS-stimulated cells, but not in unstimulated cells or cells pretreated with ALLN. NF-κB p50/p65 subunits were detectable in endothelial nuclear protein 60 min following LPS. In contrast, NF-κB subunits from monocytes were detected at 15 min. Also, while ALLN only attenuated endothelial NF-κB translocation, monocyte NF-κB translocation was completely inhibited. These data suggest endotoxin-stimulated human coronary endothelial cells express TNF-α, which is regulated in part by NF-κB activation, in a manner and degree distinct from human monocytes.

Original languageEnglish (US)
Pages (from-to)349-354
Number of pages6
JournalShock
Volume16
Issue number5
DOIs
StatePublished - Nov 2001

Keywords

  • Electrophoretic mobility shift assay
  • Endothelium
  • Human coronary endothelial cells
  • Human monocyte
  • Lipopolysaccharide
  • N-acetyl-Leu-Leu-norleucinal
  • RT-PCR
  • Sepsis

ASJC Scopus subject areas

  • Emergency Medicine
  • Critical Care and Intensive Care Medicine

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