Albumin inhibits neutrophil spreading and hydrogen peroxide release by blocking the shedding of CD43 (sialophorin, leukosialin).
Spreading of neutrophils on protein-coated surfaces is a pivotal event in their ability to respond to soluble, physiologic agonists by releasing large amounts of hydrolases and oxidants. Using neutrophils plated on serum-, fibrinogen- or fibronectin-coated surfaces, we investigated the effect of human serum albumin (HSA) on spreading-dependent neutrophil responses. HSA suppressed the respiratory burst of neutrophils in response to tumor necrosis factor-alpha (TNF), complement component C5a or formylated peptide, but not phorbol myristate acetate. HSA was suppressive only if added before the onset of the respiratory burst, and suppression was reversed when HSA was removed. Likewise, HSA selectively and reversibly inhibited TNF-induced cell spreading and the associated fall in cAMP. However, HSA did not hinder TNF-induced cell adherence to the same protein-coated surfaces. We investigated cell surface sialoproteins as modulators of cell spreading and as targets for the anti-spreading action of HSA. Oxidation of the cell surface with periodate followed by reduction with 3H-borohydride and immunoblotting with specific mAbs helped identify the predominant sialoprotein on human neutrophils as CD43 (sialophorin, leukosialin). Treatment of neutrophils with C. perfringens sialidase desialylated CD43, markedly enhanced the ability of the cells to respond to TNF by spreading and undergoing a respiratory burst, and antagonized the ability of HSA to inhibit these responses. TNF-treated, adherent neutrophils shed CD43, and this was blocked by HSA, but not by ovalbumin. Exogenous neutrophil elastase removed CD43 from the neutrophil surface. HSA blocked the actions of both sialidase and elastase on CD43. In contrast, ovalbumin did not block the action of sialidase on CD43, and HSA did not inhibit the ability of sialidase to hydrolyze a synthetic substrate. These results suggested that HSA might bind CD43. In fact, the extracellular portion of CD43 bound to HSA-Sepharose, but not to ovalbumin- or glycylglycine-Sepharose. Finally, two mAbs recognizing different epitopes on CD43 mimicked HSA's inhibitory effects on neutrophil function. Thus, HSA can dissociate attachment of neutrophils from spreading. This dissociation may help neutrophils migrate along a chemotactic gradient, while decreasing their release of oxidants. CD43, a long, rigid molecule with a markedly negative charge, antagonizes neutrophil spreading. HSA appears to inhibit spreading-dependent neutrophil functions by binding to CD43 and interfering with the ability of neutrophils to shed it.
Nathan
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Beatrice and Samuel A. Seaver Laboratory, Department of Medicine, Cornell University Medical College, New York.
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Halbwachs-Mecarelli
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Jin
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D015703:Antigens, CD / Q000187:drug effects / Q000302:isolation & purification / Q000502:physiology*; D002448:Cell Adhesion; D002465:Cell Movement / Q000187:drug effects*; D002846:Chromatography, Affinity; D015936:Complement C5a / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D006801:Humans; D006861:Hydrogen Peroxide / Q000097:blood*; D007700:Kinetics; D051917:Leukosialin; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009439:Neuraminidase / Q000037:antagonists & inhibitors / Q000494:pharmacology; D009504:Neutrophils / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D010196:Pancreatic Elastase / Q000037:antagonists & inhibitors / Q000494:pharmacology; D010504:Periodic Acid / Q000494:pharmacology; D011994:Recombinant Proteins / Q000494:pharmacology; D012709:Serum Albumin / Q000494:pharmacology*; D012795:Sialoglycoproteins / Q000187:drug effects / Q000302:isolation & purification / Q000502:physiology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D015703:Antigens, CD; D051917:Leukosialin; D011994:Recombinant Proteins; C465014:SPN protein, human; D012709:Serum Albumin; D012795:Sialoglycoproteins; D014409:Tumor Necrosis Factor-alpha; D010504:Periodic Acid; D009240:N-Formylmethionine Leucyl-Phenylalanine; D015936:Complement C5a; C009288:metaperiodate; D006861:Hydrogen Peroxide; D009439:Neuraminidase; D010196:Pancreatic Elastase; D013755:Tetradecanoylphorbol Acetate