Inhibition of inducible nitric oxide synthase gene expression by glucocorticoid-induced protein(s) in lipopolysaccharide-stimulated J774 cells.
Glucocorticoids inhibit inducible-type NO synthase activity in a variety of cell types. We report here that proteins recovered from the medium of dexamethasone-treated J774 macrophages (1, 10, 100 microg/ml) inhibited lipopolysaccharide-stimulated nitrite generation by 10.0 +/- 3.0%, 32.3 +/- 5.3% and 55.0 +/- 6.0%, respectively, and inducible NO synthase mRNA expression in these cells. Immunoblotting analysis of crude and partially purified glucocorticoid-induced proteins with an anti-lipocortin-1 polyclonal antiserum revealed the presence of lipocortin-1-like immunoreactive species with a molecular mass of 35-37 kDa. Furthermore, inhibition of lipopolysaccharide-induced nitrite production by glucocorticoid-induced proteins in J774 cells was reversed by addition of anti-lipocortin-1 neutralizing polyclonal antibody (1:60 dilution; 4 h before lipopolysaccharide). Comparison of glucocorticoid-induced proteins inhibition of both nitrite production and inducible NO synthase mRNA expression suggests that these effects result mainly from inhibition of lipopolysaccharide-mediated inducible NO synthase gene expression. These results indicate that negative regulation of inducible NO synthase by glucocorticoids is, at least in part, mediated by glucocorticoid-induced proteins that involve also members of the lipocortin-like superfamily.
D'Acquisto
F
F
Department of Experimental Pharmacology, University of Naples Federico II, Italy.
Cicatiello
L
L
Iuvone
T
T
Ialenti
A
A
Ianaro
A
A
Esumi
H
H
Weisz
A
A
Carnuccio
R
R
D000818:Animals; D017305:Annexin A1 / Q000276:immunology; D002478:Cells, Cultured; D017077:Culture Media, Conditioned; D003907:Dexamethasone / Q000494:pharmacology*; D004790:Enzyme Induction; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology*; D005938:Glucocorticoids / Q000494:pharmacology*; D015151:Immunoblotting; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008264:Macrophages / Q000187:drug effects*; D051379:Mice; D019001:Nitric Oxide Synthase / Q000235:genetics*; D052247:Nitric Oxide Synthase Type II; D009573:Nitrites / Q000378:metabolism; D013268:Stimulation, Chemical
D017305:Annexin A1; D017077:Culture Media, Conditioned; D005938:Glucocorticoids; D008070:Lipopolysaccharides; D009573:Nitrites; D003907:Dexamethasone; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse