Structural stabilization in tetrameric or polymeric hemoglobin determines its interaction with endogenous antioxidant scavenger pathways.
Hemoglobin (Hb) released into the circulation during hemolysis and chemically modified Hb proposed for use as oxygen therapeutics exert toxic effects that are partially attributable to heme's oxidant activity. Native extracellular Hb is scavenged by haptoglobin (Hp) after alphabeta-subunit dimerization. In the absence of Hp, monocyte/macrophage cell-surface CD163 binds and clears Hb. We evaluated several chemically modified Hbs to establish the role of chemical cross-linking patterns and molecular sizes on binding and clearance by each pathway. We found that Hbs possessing beta-globin cross-linking, irrespective of polymerization, demonstrate increased Hp affinity compared with alpha-globin-stabilized Hbs. These data suggest that Hb alpha-subunit accessibility is critical for Hp binding in the absence of dimerization. beta-Globin chain cross-linked tetramers/polymers displayed strong polyvalent Hp binding with increased viscosity and formation of visible gel matrices. Modified Hb interaction with CD163 and cellular uptake demonstrated an inverse relation with molecular size, irrespective of alpha and beta cross-linking. These findings were confirmed by HO-1 induction and intracellular ferritin accumulation in CD163-expressing HEK293 cells. Based on these results, a rational and systematic approach to HBOC design may be used to optimize interaction with endogenous Hb clearance and detoxification pathways.
Laboratory of Biochemistry and Vascular Biology, Division of Hematology, Center for Biologics Evaluation and Research (CBER), U.S. Food and Drug Administration (FDA), Rockville, Maryland, USA.
Paul W
PW
Buehler
Florence
F
Vallelian
Malgorzata G
MG
Mikolajczyk
Gabriele
G
Schoedon
Thomas
T
Schweizer
Abdu I
AI
Alayash
Dominik J
DJ
Schaer
D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism; D015214:Antigens, Differentiation, Myelomonocytic / Q000235:genetics / Q000378:metabolism; D000975:Antioxidants / Q000378:metabolism*; D002460:Cell Line; D002850:Chromatography, Gel; D019281:Dimerization; D005293:Ferritins / Q000378:metabolism; D005914:Globins / Q000378:metabolism; D006242:Haptoglobins / Q000378:metabolism; D006454:Hemoglobins / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D008954:Models, Biological; D011485:Protein Binding; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism; D051116:Receptors, Scavenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D020349:Surface Plasmon Resonance; D000098902:CD163 Antigen
D015703:Antigens, CD; D015214:Antigens, Differentiation, Myelomonocytic; D000975:Antioxidants; D000098902:CD163 Antigen; D006242:Haptoglobins; D006454:Hemoglobins; D011956:Receptors, Cell Surface; D051116:Receptors, Scavenger; D005914:Globins; D005293:Ferritins