title: GM-CSF and TNFα modulate protein expression of human neutrophils visualized by fluorescence two-dimensional difference gel electrophoresis.
abstract: Increased serum levels of TNFα and GM-CSF are found in various chronic inflammatory diseases and these cytokines affect the function of circulating and tissue neutrophils. TNFα- and GM-CSF-induced protein expression profiles could, therefore, serve as biomarker for the action of these cytokines in vivo. We stimulated human peripheral neutrophils with TNFα and GM-CSF in vitro and analyzed changes in their proteome by fluorescence two-dimensional difference gel electrophoresis (2D-DIGE). We report the differential expression of 3 and 18 protein spots following TNFα and GM-CSF stimulation, respectively. Differences in protein expression induced by TNFα were limited and did not show discriminatory power in a principal component analysis, whereas the profile induced by GM-CSF did. TNFα- and GM-CSF-induced both de novo IL-1β and sIL-1Ra protein expression as detected by Western blot analysis, which confirmed proper neutrophil activation by these cytokines in vitro. Mass spectrometry analysis of cytokine-regulated protein spots resulted in the identification of 8 proteins. Among the identified proteins, enolase 1 and annexin A1 might function as markers for peripheral neutrophil activation. In conclusion, a proteomic analysis of neutrophils by 2D-DIGE provides proof-of-principle that cytokine-induced protein profiles can serve as biomarkers for the action of individual cytokines in vivo.
authors.affiliation: Department of Respiratory Medicine, University Medical Center Utrecht, Heidelberglaan 100, 3584CX Utrecht, The Netherlands.
authors.forename: Jeroen D
authors.initials: JD
authors.lastname: Langereis
authors.forename: Lorenza
authors.initials: L
authors.lastname: Franciosi
authors.forename: Laurien H
authors.initials: LH
authors.lastname: Ulfman
authors.forename: Leo
authors.initials: L
authors.lastname: Koenderman
mesh_terms: D015153:Blotting, Western; D002853:Chromatography, Liquid; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D005453:Fluorescence; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000097:blood / Q000502:physiology*; D006801:Humans; D009504:Neutrophils / Q000378:metabolism*; D025341:Principal Component Analysis; D053719:Tandem Mass Spectrometry; D014409:Tumor Necrosis Factor-alpha / Q000097:blood / Q000502:physiology*
chemical_list: D014409:Tumor Necrosis Factor-alpha; D016178:Granulocyte-Macrophage Colony-Stimulating Factor