title: Bilirubin impairs bactericidal activity of neutrophils through an antioxidant mechanism in vitro.
abstract: BACKGROUND
  Liver failure accompanied by hyperbilirubinemia after major hepatic resection is profoundly associated with septic complications. Although the immune dysfunction in cholestasis has been intensively investigated, the contribution of increased serum bilirubin to the impaired resistance to bacterial infection remains to be elucidated. Because bilirubin possesses an antioxidant activity, we hypothesized that bilirubin may scavenge reactive oxygen species (ROS) produced by neutrophils and consequently impair neutrophil bacterial killing. To address this, we evaluated the effects of bilirubin on the bactericidal activity of ROS or of neutrophils in vitro.
  
  
  METHODS
  The antioxidant activity of bilirubin was determined using an ROS-sensitive fluorophore, dichlorofluorescin diacetate (DCFH-DA). Bilirubin concentration in the buffer solution was monitored spectorophotometrically after incubation with ROS. The effect of bilirubin on killing of Escherichia coli by ROS or by isolated human neutrophils was determined by counting the viable E. coli after incubation on nutrient agar.
  
  
  RESULTS
  The bilirubin concentration in the buffer solution was decreased by the addition of hydrogen peroxide, especially in the presence of peroxidase or ferrous iron. DCFH-DA oxidation by ROS or activated neutrophils was inhibited by bilirubin in a dose-dependent manner. The bactericidal activity of ROS or of isolated neutrophils was significantly attenuated by bilirubin.
  
  
  CONCLUSIONS
  Bilirubin impairs bactericidal activity of neutrophils through scavenging ROS. Increased levels of serum bilirubin may well be responsible for the impaired bacterial clearance in patients with hyperbilirubinemia.
authors.affiliation: First Department of Surgery, Nagoya University Graduate School of Medicine, Nagoya 466-8550, Japan. tarai@med.nagoya-u.ac.jp
authors.forename: T
authors.initials: T
authors.lastname: Arai
authors.forename: Y
authors.initials: Y
authors.lastname: Yoshikai
authors.forename: J
authors.initials: J
authors.lastname: Kamiya
authors.forename: M
authors.initials: M
authors.lastname: Nagino
authors.forename: K
authors.initials: K
authors.lastname: Uesaka
authors.forename: N
authors.initials: N
authors.lastname: Yuasa
authors.forename: K
authors.initials: K
authors.lastname: Oda
authors.forename: T
authors.initials: T
authors.lastname: Sano
authors.forename: Y
authors.initials: Y
authors.lastname: Nimura
mesh_terms: D000975:Antioxidants / Q000378:metabolism*; D001424:Bacterial Infections / Q000276:immunology / Q000378:metabolism; D001663:Bilirubin / Q000378:metabolism*; D002779:Cholestasis / Q000276:immunology / Q000378:metabolism / Q000382:microbiology; D004926:Escherichia coli / Q000276:immunology*; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D006932:Hyperbilirubinemia / Q000276:immunology* / Q000378:metabolism / Q000382:microbiology; D066298:In Vitro Techniques; D017093:Liver Failure / Q000276:immunology / Q000378:metabolism / Q000382:microbiology; D018375:Neutrophil Activation / Q000502:physiology; D009504:Neutrophils / Q000378:metabolism* / Q000382:microbiology*; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D000975:Antioxidants; D017382:Reactive Oxygen Species; D006861:Hydrogen Peroxide; D001663:Bilirubin