Endothelial nitric-oxide synthase enhances lipopolysaccharide-stimulated tumor necrosis factor-alpha expression via cAMP-mediated p38 MAPK pathway in cardiomyocytes

J Biol Chem. 2003 Mar 7;278(10):8099-105. doi: 10.1074/jbc.M207288200. Epub 2002 Dec 27.


The purpose of this study was to investigate the role of endothelial nitric-oxide synthase (eNOS), cAMP, and p38 MAPK in tumor necrosis factor-alpha (TNF-alpha) expression induced by lipopolysaccharide (LPS). LPS dose- and time-dependently induced phosphorylation of p38 MAPK and TNF-alpha expression in neonatal mouse cardiomyocytes. TNF-alpha expression was preceded by p38 MAPK phosphorylation, and selective inhibition of p38 MAPK abrogated LPS-induced TNF-alpha expression. Deficiency in eNOS decreased basal and LPS-stimulated TNF-alpha expression in cardiomyocytes. NOS inhibitor l-NAME attenuated LPS-induced p38 MAPK phosphorylation and TNF-alpha production in wild-type cardiomyocytes, whereas NO donor 2,2'-(hydroxynitrosohydrazono)bis-ethanamine (DETA-NO) (2 microm) or overexpression of eNOS by adenoviral gene transfer restored the response of eNOS(-/-) cardiomyocytes to LPS. These effects of NO were mediated through cAMP-dependent pathway based on the following facts. First, deficiency in eNOS decreased basal levels of intracellular cAMP, and DETA-NO elevated intracellular cAMP levels in eNOS(-/-) cardiomyocytes. Second, a cAMP analogue 8-Br-cAMP mimicked the effect of NO in eNOS(-/-) cardiomyocytes. Third, either inhibition of cAMP or cAMP-dependent protein kinase attenuated LPS-stimulated p38 MAPK phosphorylation and TNF-alpha production in wild-type cardiomyocytes. In conclusion, eNOS enhances LPS-stimulated TNF-alpha expression in cardiomyocytes. Activation of p38 MAPK is essential in LPS-stimulated TNF-alpha expression. Moreover, the effects of NO on LPS-stimulated TNF-alpha expression are mediated through cAMP/cAMP-dependent protein kinase-dependent p38 MAPK pathway in neonatal cardiomyocytes.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Animals, Newborn
  • Base Sequence
  • Cyclic AMP / metabolism*
  • DNA Primers
  • Lipopolysaccharides / pharmacology*
  • Mice
  • Mice, Inbred C57BL
  • Mitogen-Activated Protein Kinases / metabolism*
  • Myocardium / cytology
  • Myocardium / enzymology
  • Myocardium / metabolism*
  • Nitric Oxide / metabolism
  • Nitric Oxide Synthase / metabolism*
  • Nitric Oxide Synthase Type II
  • Nitric Oxide Synthase Type III
  • Phosphorylation
  • Tumor Necrosis Factor-alpha / genetics*
  • Tumor Necrosis Factor-alpha / metabolism
  • p38 Mitogen-Activated Protein Kinases


  • DNA Primers
  • Lipopolysaccharides
  • Tumor Necrosis Factor-alpha
  • Nitric Oxide
  • Cyclic AMP
  • Nitric Oxide Synthase
  • Nitric Oxide Synthase Type II
  • Nitric Oxide Synthase Type III
  • Nos3 protein, mouse
  • Mitogen-Activated Protein Kinases
  • p38 Mitogen-Activated Protein Kinases