title: Ghrelin inhibits LPS-induced release of IL-6 from mouse dopaminergic neurones.
abstract: BACKGROUND
  Ghrelin is an orexigenic stomach hormone that acts centrally to increase mid-brain dopamine neurone activity, amplify dopamine signaling and protect against neurotoxin-induced dopamine cell death in the mouse substantia nigra pars compacta (SNpc). In addition, ghrelin inhibits the lipopolysaccharide (LPS)-induced release of pro-inflammatory cytokines from peripheral macrophages, T-cells and from LPS stimulated microglia. Here we sought to determine whether ghrelin attenuates pro-inflammatory cytokine release from dopaminergic neurones.
  
  
  RESULTS
  The dopaminergic SN4741 cell-line, which derives from the mouse substantia nigra (SN) and expresses the ghrelin-receptor (growth hormone secretagogue receptor (GHS-R)) and the ghrelin-O-acyl transferase (GOAT) enzyme, was used to determine the neuro-immunomodulatory action of ghrelin. We induced innate immune activation via LPS challenge (1 μg/ml) of SN4741 neurones that had been pre-cultured in the presence or absence of ghrelin (1, 10, 100 nM) for 4 h. After 24 h supernatants were collected for detection of IL-1 beta (IL-1β ), TNF alpha (TNF-α) and IL-6 cytokines via enzyme linked immunosorbent assay (ELISA) analysis. Nuclear translocation of the transcription factor nuclear factor kappa B (NF-κB) was analyzed by Western blotting, and to determine viability of treatments a cell viability assay and caspase-3 immunohistochemistry were performed.We provide evidence that while IL-1β and TNF-α were not detectable under any conditions, SN4741 neurones constitutively released IL-6 under basal conditions and treatment with LPS significantly increased IL-6 secretion. Pre-treatment of neurones with ghrelin attenuated LPS-mediated IL-6 release at 24 h, an affect that was inhibited by the GHS-R antagonist [D-Lys3]-GHRP-6. However, while ghrelin pre-treatment attenuated the LPS-mediated increase in NF-κB, there was no alteration in its nuclear translocation. Cell viability assay and caspase-3 immunocytochemistry demonstrated that the results were independent from activation of cytotoxic and/or apoptotic mechanisms in the neuronal population, respectively.
  
  
  CONCLUSIONS
  Our results provide evidence that the gut-hormone, ghrelin, attenuates IL-6 secretion to LPS challenge in mid-brain dopaminergic neurones. These data suggest that ghrelin may protect against dopaminergic SN nerve cell damage or death via modulation of the innate immune response.
authors.affiliation: Molecular Neuroscience, Swansea University, Swansea, SA28PP, UK.
authors.forename: Amy L
authors.initials: AL
authors.lastname: Beynon
authors.forename: M Rowan
authors.initials: MR
authors.lastname: Brown
authors.forename: Rhiannon
authors.initials: R
authors.lastname: Wright
authors.forename: Mark I
authors.initials: MI
authors.lastname: Rees
authors.forename: I Martin
authors.initials: IM
authors.lastname: Sheldon
authors.forename: Jeffrey S
authors.initials: JS
authors.lastname: Davies
mesh_terms: D000818:Animals; D002460:Cell Line; D059290:Dopaminergic Neurons / Q000378:metabolism*; D054439:Ghrelin / Q000502:physiology*; D015850:Interleukin-6 / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000037:antagonists & inhibitors / Q000502:physiology*; D051379:Mice; D000093242:Interleukin-6 Inhibitors
chemical_list: D054439:Ghrelin; D015850:Interleukin-6; D008070:Lipopolysaccharides; D000093242:Interleukin-6 Inhibitors