title: Hydrogen peroxide signals E. coli phagocytosis by human polymorphonuclear cells; up-stream and down-stream pathway.
abstract: Hydrogen peroxide (Η2Ο2) is produced during a variety of cellular procedures. In this paper, the regulatory role of Η2Ο2, in Escherichia coli phagocytosis by the human polymorphonuclears, was investigated. White blood cells were incubated with dihydrorhodamine (DHR) in order to study H2O2 synthesis and E. coli-FITC to study phagocytosis. Flow cytometry revealed increased synthesis of H2O2 in polymorphonuclears which incorporated E. coli-FITC. The blocking of H2O2 synthesis by specific inhibitors, N-ethylmaleimide (ΝΕΜ) for NADPH oxidase and diethyldithiocarbamate (DDC) for superoxide dismutase (SOD), decreased E. coli phagocytosis, as well. Immunoblot analysis of white blood cell protein extracts revealed that the blocking of NADPH oxidase and SOD decreased ERK-1/2 phosphorylation, while it had no effect on JNK and p38. Confocal microscopy showed that phosphorylation of MAPKs and phagocytosis solely occur in the polymorphonuclear and not in mononuclear cells. The use of specific MAPKs inhibitors showed that all of them are necessary for phagocytosis, but only phospho-p38 affects H2O2 synthesis. The blocking of JNK phosphorylation, in the presence of E. coli, evoked a further decrease of cytoplasmic p47 thus increasing its translocation onto the plasma membrane for the assembly of NADPH oxidase. It appears that newly synthesised H2O2 invigorates the phosphorylation and action of ERK-1/2 in E. coli phagocytosis, while phospho-JNK and phospho-p38 appear to regulate H2O2 production.
authors.affiliation: Laboratory of Biology, Department of Biology, University of Patras, Greece.
authors.forename: Michalis
authors.initials: M
authors.lastname: Petropoulos
authors.affiliation: Laboratory of Biology, Department of Biology, University of Patras, Greece.
authors.forename: Georgia
authors.initials: G
authors.lastname: Karamolegkou
authors.affiliation: Laboratory of Biology, Department of Biology, University of Patras, Greece.
authors.forename: Eleftheria
authors.initials: E
authors.lastname: Rosmaraki
authors.affiliation: Laboratory of Biology, Department of Biology, University of Patras, Greece. Electronic address: stsakas@upatras.gr.
authors.forename: Sotiris
authors.initials: S
authors.lastname: Tsakas
mesh_terms: D004050:Ditiocarb / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004926:Escherichia coli / Q000737:chemistry / Q000276:immunology*; D005033:Ethylmaleimide / Q000494:pharmacology; D016650:Fluorescein-5-isothiocyanate / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry; D005786:Gene Expression Regulation / Q000187:drug effects* / Q000276:immunology; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism* / Q000494:pharmacology; D048670:MAP Kinase Kinase 4 / Q000235:genetics / Q000276:immunology; D019950:Mitogen-Activated Protein Kinase 1 / Q000235:genetics / Q000276:immunology; D048052:Mitogen-Activated Protein Kinase 3 / Q000235:genetics / Q000276:immunology; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D009504:Neutrophils / Q000187:drug effects / Q000276:immunology / Q000378:metabolism*; D010587:Phagocytosis / Q000187:drug effects / Q000235:genetics; D010766:Phosphorylation / Q000187:drug effects; D061251:Primary Cell Culture; D015398:Signal Transduction / Q000276:immunology*; D013194:Staining and Labeling; D013482:Superoxide Dismutase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D048051:p38 Mitogen-Activated Protein Kinases / Q000235:genetics / Q000276:immunology
chemical_list: D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D004050:Ditiocarb; D006861:Hydrogen Peroxide; D013482:Superoxide Dismutase; D019255:NADPH Oxidases; C535150:MAPK1 protein, human; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D048051:p38 Mitogen-Activated Protein Kinases; D048670:MAP Kinase Kinase 4; D016650:Fluorescein-5-isothiocyanate; D005033:Ethylmaleimide
keywords: Hydrogen peroxide; Phagocytosis; Polymorphonuclears; Signalling