title: Endogenous morphine levels are increased in sepsis: a partial implication of neutrophils.
abstract: BACKGROUND
  Mammalian cells synthesize morphine and the respective biosynthetic pathway has been elucidated. Human neutrophils release this alkaloid into the media after exposure to morphine precursors. However, the exact role of endogenous morphine in inflammatory processes remains unclear. We postulate that morphine is released during infection and can be determined in the serum of patients with severe infection such as sepsis.
  
  
  METHODS
  The presence and subcellular immunolocalization of endogenous morphine was investigated by ELISA, mass spectrometry analysis and laser confocal microscopy. Neutrophils were activated with Interleukin-8 (IL-8) or lipopolysaccharide (LPS). Morphine secretion was determined by a morphine-specific ELISA. mu opioid receptor expression was assessed with flow cytometry. Serum morphine concentrations of septic patients were determined with a morphine-specific ELISA and morphine identity was confirmed in human neutrophils and serum of septic patients by mass spectrometry analysis. The effects of the concentration of morphine found in serum of septic patients on LPS-induced release of IL-8 by human neutrophils were tested.
  
  
  RESULTS
  We confirmed the presence of morphine in human neutrophil extracts and showed its colocalisation with lactoferrin within the secondary granules of neutrophils. Morphine secretion was quantified in the supernatant of activated human polymorphonuclear neutrophils in the presence and absence of Ca(2+). LPS and IL-8 were able to induce a significant release of morphine only in presence of Ca(2+). LPS treatment increased mu opioid receptor expression on neutrophils. Low concentration of morphine (8 nM) significantly inhibited the release of IL-8 from neutrophils when coincubated with LPS. This effect was reversed by naloxone. Patients with sepsis, severe sepsis and septic shock had significant higher circulating morphine levels compared to patients with systemic inflammatory response syndrome and healthy controls. Mass spectrometry analysis showed that endogenous morphine from serum of patient with sepsis was identical to poppy-derived morphine.
  
  
  CONCLUSIONS
  Our results indicate that morphine concentrations are increased significantly in the serum of patients with systemic infection and that morphine is, at least in part, secreted from neutrophils during sepsis. Morphine concentrations equivalent to those found in the serum of septic patients significantly inhibited LPS-induced IL-8 secretion in neutrophils.
authors.affiliation: Inserm, U575, Physiopathologie du Système Nerveux and Nociception and Pain Department, Institut des Neurosciences Cellulaires et Intégratives, Centre National de la Recherche Scientifique, Université de Strasbourg, Strasbourg, France.
authors.forename: Elise
authors.initials: E
authors.lastname: Glattard
authors.forename: Ingeborg D
authors.initials: ID
authors.lastname: Welters
authors.forename: Thomas
authors.initials: T
authors.lastname: Lavaux
authors.forename: Arnaud H
authors.initials: AH
authors.lastname: Muller
authors.forename: Alexis
authors.initials: A
authors.lastname: Laux
authors.forename: Dan
authors.initials: D
authors.lastname: Zhang
authors.forename: Alexander R
authors.initials: AR
authors.lastname: Schmidt
authors.forename: François
authors.initials: F
authors.lastname: Delalande
authors.forename: Benoît-Joseph
authors.initials: BJ
authors.lastname: Laventie
authors.forename: Sylvie
authors.initials: S
authors.lastname: Dirrig-Grosch
authors.forename: Didier A
authors.initials: DA
authors.lastname: Colin
authors.forename: Alain
authors.initials: A
authors.lastname: Van Dorsselaer
authors.forename: Dominique
authors.initials: D
authors.lastname: Aunis
authors.forename: Marie-Hélène
authors.initials: MH
authors.lastname: Metz-Boutigue
authors.forename: Francis
authors.initials: F
authors.lastname: Schneider
authors.forename: Yannick
authors.initials: Y
authors.lastname: Goumon
mesh_terms: D015153:Blotting, Western; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D006801:Humans; D007150:Immunohistochemistry; D016209:Interleukin-8 / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology; D008213:Lymphocyte Activation / Q000187:drug effects; D018613:Microscopy, Confocal; D009020:Morphine / Q000097:blood*; D009504:Neutrophils / Q000187:drug effects / Q000502:physiology*; D017450:Receptors, Opioid, mu / Q000378:metabolism; D018805:Sepsis / Q000097:blood*; D053719:Tandem Mass Spectrometry
chemical_list: D016209:Interleukin-8; D008070:Lipopolysaccharides; D017450:Receptors, Opioid, mu; D009020:Morphine