Nepalolide A inhibits the expression of inducible nitric oxide synthase by modulating the degradation of IkappaB-alpha and IkappaB-beta in C6 glioma cells and rat primary astrocytes.

Abstract

1 The effects of nepalolide A on the expression of inducible nitric oxide synthase (iNOS) caused by incubation with lipopolysaccharide/interferon-gamma (LPS/IFN-gamma) or tumour necrosis factor-alpha/interleukin-1beta/IFN-gamma (TNF-alpha/IL-1beta/IFN-gamma, mixed cytokines) in C6 glioma cells and primary astrocytes of rat were investigated. The mechanisms by which nepalolide A confers its effect on iNOS expression were also elucidated. 2 Treatment with LPS/IFN-gamma and mixed cytokines for 24 h elicited the induction of iNOS activity as determined by nitrite accumulation in the culture medium and assay of enzyme activity. Nepalolide A at 10 microM abrogated the LPS/IFN-gamma- and mixed cytokines-mediated induction of iNOS by more than 90% in C6 glioma cells, and by 80% for mixed cytokines-induced induction of iNOS in primary astrocytes. The effect of nepalolide A (2-10 microM) was concentration-dependent. 3 The inhibition of iNOS induction by nepalolide A was attributed to decreases in the content of iNOS protein and the level of iNOS mRNA, as measured by immunoblotting and reverse transcriptase-polymerase chain reaction. 4 Electrophoretic mobility shift assay was used to evaluate the effect of nepalolide A on the activation of nuclear factor-kappaB (NF-kappaB). Results showed that nepalolide A diminished the LPS/IFN-gamma-mediated association of NF-kappaB with consensus oligonucleotide in a concentration-dependent manner. The activation of NF-kappaB by mixed cytokines was modulated both in the extent of activation and in its time-course by nepalolide A. 5 The ability of nepalolide A to inhibit NF-kappaB activation was further confirmed by studies on the degradation of the inhibitor of NF-kappaB, IkappaB, as measured by immunoblotting. 6 The present study demonstrates that the attenuation of NF-kappaB activation by nepalolide A was mediated by blockade of the degradation of IkappaB, leading to suppression of the expression of iNOS.

• Authors: C N Wang, Y J Shiao, Y L Lin, C F Chen
• Journal: British journal of pharmacology (Br J Pharmacol) Vol. 128 Issue 2 Pg. 345-56 (Sep 1999) ISSN: 0007-1188 [Print] England
• PMID: 10510444 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Anthracenes
• DNA-Binding Proteins
• Enzyme Inhibitors
• I kappa B beta protein
• I-kappa B Proteins
• NF-kappa B
• Nfkbia protein, rat
• Nitrites
• Oligonucleotides
• RNA, Messenger
• Recombinant Proteins
• nepalolide A
• NF-KappaB Inhibitor alpha
• Interferon-gamma
• Nitric Oxide Synthase
• Nitric Oxide Synthase Type II
• Nos2 protein, rat

Topics

• Animals
• Anthracenes (pharmacology)
• Astrocytes (drug effects, metabolism)
• Biotransformation (drug effects)
• Blotting, Western
• Brain Neoplasms (metabolism)
• Cell Survival (drug effects)
• Cells, Cultured
• DNA-Binding Proteins (metabolism)
• Depression, Chemical
• Electrophoresis
• Enzyme Inhibitors (pharmacology)
• Glioma (metabolism)
• I-kappa B Proteins
• Interferon-gamma (antagonists & inhibitors, pharmacology)
• NF-KappaB Inhibitor alpha
• NF-kappa B (antagonists & inhibitors, biosynthesis)
• Nitric Oxide Synthase (antagonists & inhibitors, biosynthesis)
• Nitric Oxide Synthase Type II
• Nitrites (metabolism)
• Oligonucleotides (metabolism)
• Precipitin Tests
• RNA, Messenger (biosynthesis)
• Rats
• Rats, Sprague-Dawley
• Recombinant Proteins