title: Inhibition of inducible nitric oxide synthase gene expression and nitric oxide synthesis in vascular smooth muscle cells by granulocyte-colony stimulating factor in vitro.
abstract: Clinical and experimental evidence suggests that granulocyte-colony stimulating factor (G-CSF) acts as an anti-inflammatory modulator with beneficial effects in severe inflammatory diseases, e.g., sepsis and septic shock. Excessive production of nitric oxide (NO) is regarded as a potent mediator of the vascular changes leading to systemic hypotension that occurs during sepsis. Therefore, the aim of the present study was to investigate the influence of G-CSF on inducible nitric oxide synthase (iNOS) gene expression and NO synthesis in vascular smooth muscle cells (VSMC). Qualitative and quantitative analyses of iNOS cDNA revealed that G-CSF significantly reduced interferon-gamma/lipopolysaccharide (IFN-gamma/LPS) dependent iNOS gene expression (P < 0.05) following 6, 18, 24, and 48 h incubation periods. In addition, the co-application of G-CSF resulted in a decreased IFN-gamma/LPS mediated iNOS protein generation as detected by immunoblotting methods after 24 and 48 h. Measurement of the stable NO metabolites showed a significant reduction of nitrite/nitrate concentrations following co-incubation of VSMC with G-CSF + IFN-gamma/LPS (242.57 +/- 10.73 nmol NO2-/NO3-/mg cell protein, n = 8) as compared to IFN-gamma/LPS treatment (306.20 +/- 19.26 nmol NO2-/NO3-/mg cell protein, n = 8, P < 0.05) following a 24-h incubation protocol. This inhibitory effect of G-CSF was still present after a 48 h incubation period (G-CSF + IFN-gamma/LPS: 319.56 +/- 6.26 nmol NO2-/NO3-/mg cell protein; IFN-gamma/LPS: 489.20 +/- 27.15 nmol NO2-/NO3-/mg cell protein (P < 0.05), n = 8, respectively). The present findings suggest that inhibition of iNOS gene expression and NO generation in VSMC might be one of the protective anti-inflammatory effects of G-CSF during sepsis.
authors.affiliation: Department of Physiology, University of Bonn, Germany.
authors.forename: C
authors.initials: C
authors.lastname: Deetjen
authors.forename: S
authors.initials: S
authors.lastname: Frede
authors.forename: M
authors.initials: M
authors.lastname: Smolny
authors.forename: M
authors.initials: M
authors.lastname: Seibel
authors.forename: W
authors.initials: W
authors.lastname: Schobersberger
authors.forename: G
authors.initials: G
authors.lastname: Hoffmann
mesh_terms: D000818:Animals; D001483:Base Sequence; D002474:Cell-Free System; D017931:DNA Primers / Q000235:genetics; D005260:Female; D015870:Gene Expression / Q000187:drug effects; D016179:Granulocyte Colony-Stimulating Factor / Q000494:pharmacology*; D066298:In Vitro Techniques; D007371:Interferon-gamma / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology; D009131:Muscle, Smooth, Vascular / Q000187:drug effects* / Q000378:metabolism*; D009569:Nitric Oxide / Q000096:biosynthesis*; D019001:Nitric Oxide Synthase / Q000235:genetics*; D052247:Nitric Oxide Synthase Type II; D051381:Rats; D011921:Rats, Inbred WKY; D011994:Recombinant Proteins
chemical_list: D017931:DNA Primers; D008070:Lipopolysaccharides; D011994:Recombinant Proteins; D016179:Granulocyte Colony-Stimulating Factor; D009569:Nitric Oxide; D007371:Interferon-gamma; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496318:Nos2 protein, rat