title: Expression of pro-inflammatory cytokines by flow-sorted alveolar macrophages in severe pneumonia.
abstract: The aim of the present study was to further characterize the role of alveolar macrophages (AM) in acute human lung inflammation by evaluating their capacity to produce pro-inflammatory cytokines such as tumour necrosis factor (TNF)-alpha, interleukin (IL)-6 and IL-8. Patients with severe community-acquired pneumonia (CAP; n=12) and healthy volunteers (n=10) underwent bronchoalveolar lavage (BAL). AM were separated to high purity (>96%) using fluorescence-activated cell sorting. We determined the TNF-alpha, IL-6 and IL-8 cytokine gene expression in AM ex vivo using semiquantitative reverse transcriptase polymerase chain reaction (RT-PCR). Moreover, we measured in vitro unstimulated, lipopolysaccharide (LPS)- and LPS/interferon-gamma inducible TNF-alpha, IL-6 and IL-8 cytokine release and evaluated samples of BAL fluids for the same pro-inflammatory cytokines using an enzyme-linked immunosorbent assay (ELISA). We found increased TNF-alpha, IL-6 and IL-8 messenger ribonucleic acid (mRNA) levels in AM from CAP patients that were significantly elevated only for IL-8. When challenged with endotoxin in vitro, AM obtained from CAP patients showed a strongly reduced potential to release TNF-alpha and IL-6 compared to healthy controls, whereas IL-8 secretion did not differ significantly between groups. Moreover, stimulation of AM from CAP patients with LPS plus IFN-gamma augmented TNF-alpha and IL-6 cytokine release to near normal levels. Interestingly, no TNF-alpha protein was measured in BAL samples from CAP patients, whereas IL-6 and IL-8 protein levels were found to be significantly increased. Together, highly purified alveolar macrophages from community-acquired pneumonia patients show relatively low ex vivo tumour necrosis factor-alpha and interleukin-6 but not interleukin-8 messenger ribonucleic acid levels that are associated with a decreased pro-inflammatory cytokine release in vitro which, however, can be restored by concurrent interferon-gamma stimulation.
authors.affiliation: Dept of Internal Medicine, Justus-Liebig-University, Giessen, Germany.
authors.forename: U
authors.initials: U
authors.lastname: Maus
authors.forename: S
authors.initials: S
authors.lastname: Rosseau
authors.forename: U
authors.initials: U
authors.lastname: Knies
authors.forename: W
authors.initials: W
authors.lastname: Seeger
authors.forename: J
authors.initials: J
authors.lastname: Lohmeyer
mesh_terms: D000328:Adult; D001992:Bronchoalveolar Lavage Fluid / Q000166:cytology; D016022:Case-Control Studies; D002469:Cell Separation; D017714:Community-Acquired Infections / Q000276:immunology / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D007371:Interferon-gamma / Q000494:pharmacology; D015850:Interleukin-6 / Q000096:biosynthesis* / Q000235:genetics; D016209:Interleukin-8 / Q000096:biosynthesis* / Q000235:genetics; D008070:Lipopolysaccharides / Q000494:pharmacology; D016676:Macrophages, Alveolar / Q000276:immunology / Q000378:metabolism*; D008297:Male; D008875:Middle Aged; D018410:Pneumonia, Bacterial / Q000276:immunology / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D015850:Interleukin-6; D016209:Interleukin-8; D008070:Lipopolysaccharides; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D007371:Interferon-gamma