Neutrophil Deactivation by Influenza A Viruses: Mechanisms of Protection After Viral Opsonization With Collectins and Hemagglutination-Inhibiting Antibodies zyxw By Kevan zyxwvutsrqp L. Hartshorn, Kenneth B.M. Reid, Mitchell R. White, Jens C. Jensenius, Shirley M. Morris, Alfred I. Tauber, and Edmond Crouch Bacterial superinfections are a major cause of morbidity and mortality during influenza A virus (IAVI epidemics. Depres- sion of phagocyte functions resutting from attachment of the IAV hemagglutinin (HA) t o cell surface sialo-glycoproteins is a likely contributory cause of these infections. We have proposed that the group of collagenous lectins (termed col- lectins) present in blood and pulmonary surfactant play a role in initial host defense against IAV. We used here several recombinant human surfactant protein D (RhSP-D) prepara- tions to determine the mechanism through which opsoniza- tion of IAV with collectins protects neutrophils against the deactivating effects of IAV on cellular respiratory burst re- sponses in vitro. RhSP-D was markedly more potent than antibodies that inhibited viral hemagglutination activity (anti-HA antibodies) at protecting neutrophils in this assay. Unlike the anti-HA antibodies, RhSP-Dwas protective at con- centrations that minimally inhibited viral hemagglutination ACTERIAL superinfections are a major cause of mor- B bidity and mortality during influenza A virus (IAV) epidemics.’ These superinfections include not only bacterial pneumonia2 and otitis, but also meningococcal meningitis and IAV and parainfluenza viruses are notable among respiratory viruses for being consistently as- sociated with substantial increases in hospital admissions for lower respiratory tract infection in adults.’.’” Although damage to respiratory epithelium is one likely contributor to the development of bacterial superinfection, both IAV and parainfluenza viruses induce another important defect in the host defense barrier by causing phagocyte dysfunction. I‘ ‘I Neutrophils and monocyte/macrophages participate in the early inflammatory response to IAV infection in the airway and both cell types exhibit depressed function in vivo during IAV infection.’* In animal models, this induced functional defect correlates temporally with increased susceptibility to bacterial superinfection. zyxwvutsr l4 Neutrophil functional depression by IAV or parainfluenza virus can be reproduced in IAV itself activates ~ zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHGFEDCBA From the Departments of Medicine and Biochemistry, Boston Uni- versity School of Medicine, Boston, MA; the Department of Biochem- istry, University zyxwvutsrqpo of Oxford, Oxford, UK; the Department of Immunol- ogy, University of Aarhus, Aarhus, Denmark; and the Department of Pathology, Washington University School of Medicine, St LouiA, MO. Submitted May 26, 1995; accepted November 28, 1995. Supported by National Institutes of Health Grants No. A1925.50- 03 and A134897 (K.L.H.) and HL.44015 (E.C.). Address reprint requests to Kevan L. Hartshorn, MD, Boston University School of Medicine, S-301, 80 E Concord St, Boston, MA 02118. The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. section 1734 solely to indicate this fact. zyxwvutsrq 0 1996 by The American Society of Hematology. 0006-497I/96/8708-0045$3.00/0 activity. Two related features of SP-D-the degree of multi- merization and the ability to cause aggregation of IAV parti- cles-were critical determinants of the ability of SP-D t o protect neutrophils against deactivation. Similarly SP-D-in- duced viral aggregate formation resulted in enhanced IAV binding to neutrophils and potentiated the ability of the vi- rus itself to trigger neutrophil respiratory burst responses. In contrast t o the case of IAV-antibody complexes, SP-D-IAV complexes attached t o and activated neutrophils through a neuraminidase-sensitive mechanism (ie, similar t o unopso- nized IAV). These results indicate that collectin-mediated vi- ral aggregation per se may be an important host defense mechanism not only by virtue of reducing the number of infectious viral particles, but also by promoting phagocyte responsiveness. 0 1996 zyxwv by The American Society of Hematology. neutrophils to generate a respiratory burst response that is atypical in that H202 is generated in the absence of detect- able 0; elaboration.’”2’Both IAV-induced respiratory burst activation and IAV-induced functional depression are medi- ated by binding of the viral HA to neutrophil surface sialic acid Given the fact that IAV or the IAV hemag- glutinin (HA) both activate and functionally depress neutro- phils, we have termed the latter phenomenon deactivation. I’ We have previously shown that opsonization of IAV with the mammalian serum collectins mannose-binding protein (MBP)” and conglutinin,*’ as well as the pulmonary surfac- tant collectin, pulmonary surfactant protein D (SP-D),’X in- hibits IAV HA activity and protects neutrophils against the deactivating effects of IAV. The ability of SP-D to protect neutrophils against IAV-induced deactivation is of most in- terest because SP-D present in bronchoalveolar fluids is able to interact with IAV.** We used here several recombinant human SP-D (RhSP-D) preparations to characterize the mechanism through which SP-D alters the interactions of IAV with neutrophils. Opsonization of IAV with anti-HA antibodies also modulated IAV-neutrophil interactions through mechanisms that contrasted in important respects from those of SP-D. MATERIALS AND METHODS Reagents. zyxwv Formyl-methionyl-leucyl-phenylalanine (FMLP), cy- tochalasin B, horseradish peroxidase-type 11, scopoletin, superoxide dismutase (SOD), cytochrome-C, neuraminidase-type X (protease activity, <0.002 U/mg protein), ficol, dextran, sodium citrate, citric acid, and Staph protein A were purchased from Sigma Co (St Louis, MO) and Hypaque was obtained from Winthrop Pharmaceuticals (Des Plaines, IL). Organic solvents were purchased from Fisher Scientific (Fairlawn, NJ). Dulbecco’s phosphate-buffered saline was purchased from Flow Laboratories (Costa Mesa, CA). Phospolipase C was purchased form Boehringer Mannheim (Indianapolis, IN). The monoclonal antibodies (MoAbs) 3G8 (FAb2) and IV3 (FAb) directed respectively against the FcRIII and FcRII subtypes of neu- trophil surface Fc receptors were purchased from Medarex Inc (West Lebanon, NH). RhSP-Ds were expressed in CHO K I Collectin preparations. 3450 Blood, Vol zyxwv 87, No 8 (April 151, 1996: pp 3450-3461 Downloaded from http://ashpublications.org/blood/article-pdf/87/8/3450/621250/3450.pdf by guest on 21 January 2022