The reaction of hydrogen peroxide with hemoglobin induces extensive alpha-globin crosslinking and impairs the interaction of hemoglobin with endogenous scavenger pathways.
Cell-free hemoglobin (Hb) enhances the oxidation-related toxicity associated with inflammation, ischemia, and hemolytic disorders. Hb is highly vulnerable to oxidative damage, and irreversible structural changes involving iron/heme oxidation, heme-adduct products, and amino acid oxidation have been reported. Specific structural features of Hb, such as unconstrained alpha-chains and molecular size, determine the efficiency of interactions between the endogenous Hb scavengers haptoglobin (Hp) and CD163. Using HPLC, mass spectrometry, and Western blotting, we show that H(2)O(2)-mediated Hb oxidation results in the formation of covalently stabilized globin multimers, with prominent intramolecular crosslinking between alpha-globin chains. These structural alterations are associated with reduced Hp binding, reduced CD163 interaction, and severely impaired endocytosis of oxidized Hb by the Hp-CD163 pathway. As a result, when exposed to oxidized Hb, CD163-positive HEK293 cells and human macrophages do not increase hemeoxygenase-1 (HO-1) expression, the physiological anti-oxidative macrophage response to Hb exposure. Failed Hb clearance, inadequate HO-1 expression, and the subsequent accumulation of oxidatively damaged Hb species might thus contribute to pathologies related to oxidative stress.
Vallelian
Florence
F
Internal Medicine Research Unit, University of Zurich, Zurich, Switzerland.
Pimenova
Tatiana
T
Pereira
Claudia P
CP
Abraham
Bindu
B
Mikolajczyk
Malgorzata G
MG
Schoedon
Gabriele
G
Zenobi
Renato
R
Alayash
Abdu I
AI
Buehler
Paul W
PW
Schaer
Dominik J
DJ
D015703:Antigens, CD / Q000378:metabolism; D015214:Antigens, Differentiation, Myelomonocytic / Q000378:metabolism; D015153:Blotting, Western; D002851:Chromatography, High Pressure Liquid; D006242:Haptoglobins / Q000378:metabolism*; D051547:Heme Oxygenase-1 / Q000378:metabolism; D006454:Hemoglobins / Q000378:metabolism*; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism*; D008264:Macrophages / Q000378:metabolism; D010084:Oxidation-Reduction; D018384:Oxidative Stress / Q000502:physiology*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis; D011956:Receptors, Cell Surface / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D020349:Surface Plasmon Resonance; D055542:alpha-Globins / Q000737:chemistry / Q000378:metabolism*; D000098902:CD163 Antigen
D015703:Antigens, CD; D015214:Antigens, Differentiation, Myelomonocytic; D000098902:CD163 Antigen; D006242:Haptoglobins; D006454:Hemoglobins; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D055542:alpha-Globins; D006861:Hydrogen Peroxide; C496541:HMOX1 protein, human; D051547:Heme Oxygenase-1