title: Macrophages inhibit Salmonella typhimurium replication through MEK/ERK kinase and phagocyte NADPH oxidase activities.
abstract: Host responses during the later stages of Salmonella-macrophage interactions are critical to controlling infection but have not been well characterized. After 24 h of infection, nearly half of interferon-gamma-primed murine RAW 264.7 macrophage-like cells infected by Salmonella enterica serovar Typhimurium contained filamentous bacteria. Bacterial filamentation indicates a defect in completing replication and has been previously observed in bacteria responding to a variety of stresses. To understand whether macrophage gene expression was responsible for this effect on Salmonella Typhimurium replication, we used gene arrays to profile interferon-gamma-primed RAW 264.7 cell gene expression following infection. We observed an increase in MEK1 kinase mRNA at 8 h, an increase in MEK protein at 24 h, and measured phosphorylation of MEK's downstream target kinase, ERK1/2, throughout the 24-h infection period. Treatment of cells with MEK kinase inhibitors significantly reduced numbers of filamentous bacteria observed within macrophages after 24 h and increased the number of intracellular colony-forming units. Phagocyte NADPH oxidase inhibitors and antioxidants also significantly reduced bacterial filamentation. Either MEK kinase or phagocyte oxidase inhibitors could be added 4-8 h after infection and still significantly decrease bacterial filamentation. Oxidase activity appears to mediate bacterial filamentation in parallel to MEK kinase signaling, while inducible nitric-oxide synthase inhibitors had no significant effect on bacterial morphology. In summary, Salmonella Typhimurium infection of interferon-gamma-primed macrophages triggers a MEK kinase cascade at later infection times, and both MEK kinase and phagocyte NADPH oxidase activity impair bacterial replication. These two signaling pathways mediate a host bacteriostatic pathway and may play an important role in innate host defense against intracellular pathogens.
authors.affiliation: Department of Microbiology and Immunology, University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada.
authors.forename: Carrie M
authors.initials: CM
authors.lastname: Rosenberger
authors.forename: B Brett
authors.initials: BB
authors.lastname: Finlay
mesh_terms: D000818:Animals; D001483:Base Sequence; D002455:Cell Division / Q000276:immunology*; D002460:Cell Line; D017931:DNA Primers; D005455:Fluorescent Antibody Technique; D007371:Interferon-gamma / Q000494:pharmacology; D048728:MAP Kinase Kinase Kinase 1; D008264:Macrophages / Q000187:drug effects / Q000276:immunology*; D051379:Mice; D019255:NADPH Oxidases / Q000378:metabolism*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D012486:Salmonella typhimurium / Q000166:cytology*; D015398:Signal Transduction
chemical_list: D017931:DNA Primers; D012333:RNA, Messenger; D007371:Interferon-gamma; D019255:NADPH Oxidases; D017346:Protein Serine-Threonine Kinases; D048728:MAP Kinase Kinase Kinase 1; C482578:Map3k1 protein, mouse