title: Bilirubin oxidation products, oxidative stress, and intracerebral hemorrhage.
abstract: Hematoma and perihematomal regions after intracerebral hemorrhage (ICH) are biochemically active environments known to undergo potent oxidizing reactions. We report facile production of bilirubin oxidation products (BOXes) via hemoglobin/Fenton reaction under conditions approximating putative in vivo conditions seen following ICH. Using a mixture of human hemoglobin, physiological buffers, unconjugated solubilized bilirubin, and molecular oxygen and/or hydrogen peroxide, we generated BOXes, confirmed by spectral signature consistent with known BOXes mixtures produced by independent chemical synthesis, as well as HPLC-MS of BOX A and BOX B. Kinetics are straightforward and uncomplicated, having initial rates around 0.002 microM bilirubin per microM hemoglobin per second under normal experimental conditions. In hematomas from porcine ICH model, we observed significant production of BOXes, malondialdehyde, and superoxide dismutase, indicating a potent oxidizing environment. BOX concentrations increased from 0.084 +/- 0.01 in fresh blood to 22.24 +/- 4.28 in hematoma at 72h, and were 11.22 +/- 1.90 in adjacent white matter (nmol/g). Similar chemical and analytical results are seen in ICH in vivo, indicating the hematoma is undergoing similar potent oxidations. This is the first report of BOXes production using a well-defined biological reaction and in vivo model of same. Following ICH, amounts of unconjugated bilirubin in hematoma can be substantial, as can levels of iron and hemoglobin. Oxidation of unconjugated bilirubin to yield bioactive molecules, such as BOXes, is an important discovery, expanding the role of bilirubin in pathological processes seen after ICH.
authors.affiliation: Department of Neurology, University of Cincinnati, Cincinnati, OH 45267-0536, USA. Joseph.clark@uc.edu
authors.forename: J F
authors.initials: JF
authors.lastname: Clark
authors.forename: M
authors.initials: M
authors.lastname: Loftspring
authors.forename: W L
authors.initials: WL
authors.lastname: Wurster
authors.forename: S
authors.initials: S
authors.lastname: Beiler
authors.forename: C
authors.initials: C
authors.lastname: Beiler
authors.forename: K R
authors.initials: KR
authors.lastname: Wagner
authors.forename: G J
authors.initials: GJ
authors.lastname: Pyne-Geithman
mesh_terms: D000818:Animals; D001663:Bilirubin / Q000737:chemistry / Q000378:metabolism*; D001929:Brain Edema / Q000097:blood / Q000209:etiology / Q000378:metabolism; D002543:Cerebral Hemorrhage / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology*; D004195:Disease Models, Animal; D006406:Hematoma / Q000378:metabolism / Q000473:pathology; D006454:Hemoglobins / Q000378:metabolism; D008315:Malondialdehyde / Q000378:metabolism*; D010084:Oxidation-Reduction; D018384:Oxidative Stress / Q000502:physiology*; D013482:Superoxide Dismutase / Q000378:metabolism; D013552:Swine; D013997:Time Factors
chemical_list: D006454:Hemoglobins; D008315:Malondialdehyde; D013482:Superoxide Dismutase; D001663:Bilirubin