title: Activation of superoxide formation and lysozyme release in human neutrophils by the synthetic lipopeptide Pam3Cys-Ser-(Lys)4. Involvement of guanine-nucleotide-binding proteins and synergism with chemotactic peptides.
abstract: Upon exposure to the bacterial chemotactic peptide fMet-Leu-Phe, human neutrophils release lysozyme and generate superoxide anions (O2.-). The synthetic lipoamino acid N-palmitoyl-S-[2,3-bis(palmitoyloxy)-(2RS)-propyl]-(R)-cysteine (Pam3Cys), which is derived from the N-terminus of bacterial lipoprotein, when attached to Ser-(Lys)4 [giving Pam3Cys-Ser-(Lys)4], activated O2.- formation and lysozyme release in human neutrophils with an effectiveness amounting to about 15% of that of fMet-Leu-Phe. Palmitic acid, muramyl dipeptide, lipopolysaccharide and the lipopeptides Pam3Cys-Ala-Gly, Pam3Cys-Ser-Gly, Pam3Cys-Ser, Pam3Cys-OMe and Pam3Cys-OH did not activate O2.- formation. Pertussis toxin, which ADP-ribosylates guanine-nucleotide-binding proteins (G-proteins) and functionally uncouples formyl peptide receptors from G-proteins, prevented activation of O2.- formation by fMet-Leu-Phe and inhibited Pam3Cys-Ser-(Lys)4-induced O2.- formation by 85%. Lipopeptide-induced exocytosis was pertussis-toxin-insensitive. O2.- formation induced by Pam3Cys-Ser-(Lys)4 and fMet-Leu-Phe was enhanced by cytochalasin B, by a phorbol ester and by a diacylglycerol kinase inhibitor. Addition of activators of adenylate cyclase and removal of extracellular Ca2+ inhibited O2.- formation by fMet-Leu-Phe and Pam3Cys-Ser-(Lys)4 to different extents. Pam3Cys-Ser-(Lys)4 synergistically enhanced fMet-Leu-Phe-induced O2.- formation and primed neutrophils to respond to the chemotactic peptide at non-stimulatory concentrations. Our data suggest the following. (1) Pam3Cys-Ser-(Lys)4 activates neutrophils through G-proteins, involving pertussis-toxin-sensitive and -insensitive processes. (2) The signal transduction pathways activated by fMet-Leu-Phe and Pam3Cys-Ser-(Lys)4 are similar but not identical. (3) In inflammatory processes, bacterial lipoproteins and chemotactic peptides may interact synergistically to activate O2.- formation, leading to enhanced bactericidal activity.
authors.affiliation: Institut für Pharmakologie, Freie Universität Berlin, Federal Republic of Germany.
authors.forename: R
authors.initials: R
authors.lastname: Seifert
authors.forename: G
authors.initials: G
authors.lastname: Schultz
authors.forename: M
authors.initials: M
authors.lastname: Richter-Freund
authors.forename: J
authors.initials: J
authors.lastname: Metzger
authors.forename: K H
authors.initials: KH
authors.lastname: Wiesmüller
authors.forename: G
authors.initials: G
authors.lastname: Jung
authors.forename: W G
authors.initials: WG
authors.lastname: Bessler
authors.forename: S
authors.initials: S
authors.lastname: Hauschildt
mesh_terms: D004305:Dose-Response Relationship, Drug; D004357:Drug Synergism; D005089:Exocytosis / Q000187:drug effects; D019204:GTP-Binding Proteins / Q000502:physiology*; D006801:Humans; D066298:In Vitro Techniques; D008074:Lipoproteins / Q000494:pharmacology*; D009113:Muramidase / Q000378:metabolism*; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology*; D009504:Neutrophils / Q000187:drug effects* / Q000378:metabolism; D009842:Oligopeptides / Q000494:pharmacology*; D010972:Platelet Activating Factor / Q000494:pharmacology; D013329:Structure-Activity Relationship; D013481:Superoxides / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
chemical_list: D008074:Lipoproteins; D009842:Oligopeptides; D010972:Platelet Activating Factor; D014409:Tumor Necrosis Factor-alpha; D013481:Superoxides; C061108:N-palmitoyl-S-(2,3-bis(palmitoyloxy)propyl)cysteinyl-seryl-lysyl-lysyl-lysyl-lysine; D009240:N-Formylmethionine Leucyl-Phenylalanine; D009113:Muramidase; D019204:GTP-Binding Proteins