title: Intracellular localization of interleukin-6 in eosinophils from atopic asthmatics and effects of interferon gamma.
abstract: Eosinophils, prominent cells in asthmatic inflammation, have been shown to synthesize, store, and release an array of up to 18 cytokines and growth factors, including interleukin-6 (IL-6). In this report, we show that IL-6 immunofluorescence localizes to the matrix of the crystalloid granule in peripheral blood eosinophils from atopic asthmatics using confocal laser scanning microscopy (CLSM). Granule localization of IL-6 was confirmed using dot-blot analysis and enzyme-linked immunosorbent assay (ELISA) on subcellular fractions of highly purified eosinophils produced from density centrifugation across a 0% to 45% Nycodenz gradient. IL-6 was found to coelute with eosinophil crystalloid granule marker proteins, including eosinophil peroxidase (EPO), major basic protein (MBP), arylsulfatase B, and beta-hexosaminidase. Immunoreactivity to IL-6 colocalized with granule-associated IL-2 and IL-5 in subfractionated eosinophils. We also made the novel and compelling observation that interferon gamma (IFNgamma), a Th1-type cytokine, stimulated an early elevation in eosinophil IL-6 immunoreactivity. A 2.5-fold enhancement of IL-6 immunoreactivity in eosinophil granules was observed within 10 minutes of IFNgamma treatment (500 U/mL), as determined by subcellular fractionation and CLSM. These findings suggest that IFNgamma has short-term effects on human eosinophil function and imply that a physiologic role exists for Th1-type cytokine modulation of Th2-type responses in these cells.
authors.affiliation: Pulmonary Research Group, Department of Medicine, University of Alberta, Edmonton, Alberta, Canada.
authors.forename: P
authors.initials: P
authors.lastname: Lacy
authors.forename: F
authors.initials: F
authors.lastname: Levi-Schaffer
authors.forename: S
authors.initials: S
authors.lastname: Mahmudi-Azer
authors.forename: B
authors.initials: B
authors.lastname: Bablitz
authors.forename: S C
authors.initials: SC
authors.lastname: Hagen
authors.forename: J
authors.initials: J
authors.lastname: Velazquez
authors.forename: A B
authors.initials: AB
authors.lastname: Kay
authors.forename: R
authors.initials: R
authors.lastname: Moqbel
mesh_terms: D001249:Asthma / Q000097:blood*; D003593:Cytoplasm / Q000378:metabolism; D003594:Cytoplasmic Granules / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D004804:Eosinophils / Q000378:metabolism* / Q000648:ultrastructure; D006801:Humans; D007371:Interferon-gamma / Q000494:pharmacology*; D015850:Interleukin-6 / Q000032:analysis*; D018613:Microscopy, Confocal
chemical_list: D015850:Interleukin-6; D007371:Interferon-gamma