Elevated intracellular cyclic AMP (cAMP) inhibits release of tumor necrosis factor-α (TNF-α) from porcine vascular tissue and cultured smooth muscle cells

W. H. Newman, L. M. Zhang, S. K. Leeper-Woodford, D. H. Lee, M. R. Castresana

Research output: Contribution to journalArticle

Abstract

TNF-α is a key cytokine in septic shock. We previously reported that human vascular tissue produced TNF-α, when stimulated by bacterial lipopolysaccharide (LPS) (Crit. Care Med. 24: 294, 1996). Here, we studied the role of intracellular cAMP in the regulation of production of TNF-α. Smooth muscle cells (SMC) were cultured from pig coronary artery. Cells were incubated with 20 μg/ml LPS alone or plus forskolin (FSK). At zero time, 1, 3 and 6 hr, medium was removed from the cells and analyzed for TNF-α by L929 cell cytotoxicity assay. Intracellular cAMP was analyzed by radioimmunoassay. Similar experiments were performed on sections of coronary vessel. LPS alone stimulated the release of TNF-α from SMC at 1, 3 and 6 hr. For instance at 1 hr, TNF-α in the medium from LPS stimulated cells was 9.4 ± 2.0 units/mg/cell protein vs. 1.1 ± 0.4 units/mg/cell protein in medium from control cells (p < 0.05). FSK inhibited LPS stimulated TNF-α release in a concentration-dependent manner. In the presence of LPS plus 1 or 10 μM FSK, TNF-α release at 1 hr was 3.3 ± 1.1 and 0.8 ± 0.2 units/mg cell protein, respectively (p < 0.05 vs. LPS alone). A similar suppression of TNF-α by FSK was seen at 3 and 6 hr. Intracellular cAMP increased from 10.1 ± 0.7 pmol/mg cell protein to 31.8 ± 3.3 within 5 min. in the presence of 1 μM FSK and remained elevated at 18.2 ± 2.9 pmol/mg protein at 6 hr. FSK also inhibited LPS stimulated TNF-α release from coronary segments. These results indicate that porcine vascular smooth muscle cells release TNF-α when stimulated with LPS. This release is inhibited by elevated intracellular cAMP. We propose that TNF-α from a vascular source could play a role in the hypotension of septic shock.

Original languageEnglish (US)
Pages (from-to)A261
JournalFASEB Journal
Volume11
Issue number3
StatePublished - Dec 1 1997
Externally publishedYes

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry
  • Molecular Biology
  • Genetics

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