title: Inflammatory responses to Mycoplasma hyopneumoniae in murine alveolar macrophage cell lines.
abstract: OBJECTIVE
  To investigate the mechanism by which Mycoplasma hyopneumoniae induces inflammatory responses in murine alveolar macrophage (MH-S) cells.
  
  
  METHODS
  A pathogenic strain of M. hyopneumoniae cultured in modified Friis medium was used to investigate the inflammatory response in MH-S cell lines. The effect of stimulation by M. hyopneumoniae on the production of nitric oxide (NO) and cytokines in MH-S cells and inhibition of their production, using specific inhibitors of signalling pathways, was investigated using the Griess reaction and ELISA respectively. A Western blot assay was used to confirm activation of the nuclear factor kappa B (NF-κB) and mitogen-activated protein kinase (MAPK) pathways. Nuclear translocation of NF-κB was further confirmed using transient transfection and luciferase gene reporter assay.
  
  
  RESULTS
  The results revealed dose-dependent production of NO in MH-S cells stimulated by M. hyopneumoniae. Increased concentrations of the cytokines tumour necrosis factor (TNF)-α and interleukin (IL)-1β and IL-6 were also observed (p<0.05). Using immunoblot analysis, involvement of three MAPK pathways, extracellular signal-regulated kinase I/II (ERK1/2), p38 and Jun N-terminal kinases/stress-activated protein kinases (JNK/SAPK) was confirmed. Specific inhibitors of signal pathways also demonstrated their effect on the NO and cytokine responses of MH-S cells. Degradation and phosphorylation of inhibitory kappa B (IκB)-alpha was observed, while the luciferase gene reporter assays revealed activation of NF-κB after stimulation by M. hyopneumoniae. Inhibition of NF-κB by pyrrolidine dithiocarbamate decreased M. hyopneumoniae-induced production of NO and IL-1β (p<0.05), whereas no inhibitory effect was observed on concentrations of TNF-α, and IL-6.
  
  
  CONCLUSIONS
  These findings indicate that M. hyopneumoniae induces NO and pro-inflammatory cytokines, and NF-κB and the three MAPK pathways are involved in the process.
authors.affiliation: Laboratory of Veterinary PK & PD, College of Veterinary Medicine, Kyungpook National University, Daegu, South Korea.
authors.forename: D
authors.initials: D
authors.lastname: Damte
authors.forename: S-J
authors.initials: SJ
authors.lastname: Lee
authors.forename: M-H
authors.initials: MH
authors.lastname: Hwang
authors.forename: E
authors.initials: E
authors.lastname: Gebru
authors.forename: M-J
authors.initials: MJ
authors.lastname: Choi
authors.forename: J-S
authors.initials: JS
authors.lastname: Lee
authors.forename: H
authors.initials: H
authors.lastname: Cheng
authors.forename: S-C
authors.initials: SC
authors.lastname: Park
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D002460:Cell Line; D016207:Cytokines / Q000032:analysis / Q000096:biosynthesis*; D004797:Enzyme-Linked Immunosorbent Assay; D016676:Macrophages, Alveolar / Q000276:immunology / Q000378:metabolism / Q000382:microbiology*; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000276:immunology; D045705:Mycoplasma hyopneumoniae / Q000276:immunology*; D016328:NF-kappa B / Q000276:immunology; D009569:Nitric Oxide / Q000032:analysis / Q000096:biosynthesis*
chemical_list: D016207:Cytokines; D016328:NF-kappa B; D009569:Nitric Oxide; D020928:Mitogen-Activated Protein Kinases