Expression of inducible nitric oxide synthase in bovine corneal endothelial cells and keratocytes in vitro after lipopolysaccharide and cytokines stimulation.

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State: Public
Version: Final published version
Serval ID
serval:BIB_24E2BEFD91F3
Type
Article: article from journal or magazin.
Collection
Publications
Institution
Title
Expression of inducible nitric oxide synthase in bovine corneal endothelial cells and keratocytes in vitro after lipopolysaccharide and cytokines stimulation.
Journal
Investigative Ophthalmology and Visual Science
Author(s)
Dighiero P., Behar-Cohen F., Courtois Y., Goureau O.
ISSN
0146-0404 (Print)
ISSN-L
0146-0404
Publication state
Published
Issued date
1997
Peer-reviewed
Oui
Volume
38
Number
10
Pages
2045-2052
Language
english
Notes
Publication types: Journal Article ; Research Support, Non-U.S. Gov't Publication Status: ppublish
Abstract
PURPOSE: To determine whether bovine corneal endothelial (BCE) cells and keratocytes express the inducible form of nitric oxide synthase (NOS) after exposure to cytokines and lipopolysaccharide (LPS), and to study the regulation of NOS by growth factors.
METHODS: Cultures of bovine corneal endothelial cells and keratocytes were exposed to increasing concentrations of LPS, interferon-gamma (IFN-gamma), and tumor necrosis factor-alpha (TNF-alpha). At selected intervals after exposure, nitrite levels in the supernatants were evaluated by the Griess reaction. Total RNA was extracted from the cell cultures, and messenger RNA levels for inducible NOS (NOS-2) were measured by reverse transcription-polymerase chain reaction (RT-PCR).
RESULTS: Exposure of BCE cells and keratocytes to LPS and IFN-gamma resulted in an increase of nitrite levels that was potentiate by the addition of TNF-alpha. Analysis by RT-PCR demonstrated that nitrite release was correlated to the expression of NOS-2 messenger RNA in BCE cells and keratocytes. Stereoselective inhibitors of NOS and cycloheximide inhibited LPS-IFN-gamma-induced nitrite release in both cells, whereas transforming growth factor-beta (TGF-beta) slightly potentiated it. Fibroblast growth factor-2 (FGF-2) inhibited LPS-IFN-gamma-induced nitrite release and NOS-2 messenger RNA accumulation in keratocytes but not in BCE cells.
CONCLUSIONS: The results demonstrate that in vitro activation of keratocytes and BCE cells by LPS and cytokines induces NOS-2 expression and release of large amounts of NO. The high amounts of NO could be involved in inflammatory corneal diseases in vivo.
Keywords
Animals, Cattle, Cells, Cultured, Corneal Stroma/cytology, Corneal Stroma/drug effects, Cytokines/pharmacology, DNA Primers/chemistry, Endothelium, Corneal/cytology, Endothelium, Corneal/drug effects, Enzyme Induction, Enzyme Inhibitors/pharmacology, Growth Substances/pharmacology, Lipopolysaccharides/pharmacology, Nitric Oxide Synthase/biosynthesis, Nitric Oxide Synthase/drug effects, Nitrites/metabolism, Polymerase Chain Reaction, RNA, Messenger/metabolism, Salmonella typhimurium
Pubmed
Web of science
Create date
10/04/2014 13:25
Last modification date
20/08/2019 13:03
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