title: The complement-fixing activity of immune complexes containing IgG antibodies of different functional affinities: effects on superoxide production by rabbit neutrophils.
abstract: When neutrophil phagocytes are stimulated by IgG containing immune complexes (IgG-IC), with or without the participation of the complement system, they show a sharp increase in oxygen uptake and begin to release large quantities of superoxide anions (O2-) and hydrogen peroxide (H2O2) into the surrounding medium. The aim of the present investigation was to provide insights into the production and release of O2- by rabbit neutrophils activated with immune complexes (IC) containing IgG antibodies of different functional affinity, opsonized and not opsonized by complement system components. For this purpose, two populations of polyclonal anti-ovalbumin (OVA) IgG antibodies with different functional affinity, 5 x 10(8) M(-1) and 2 x 10(7) M(-1), were prepared. The production of O2- was measured spectrophotometrically by a method using the superoxide dismutase-inhibited reduction of ferricytochrome C to the ferrous form. The activation of complement by different IgG-IC was determined by estimating the total residual haemolytic activity of the alternative and classical pathways in sera treated with different concentrations of anti-OVA IgG/ OVA immune complexes formed at equivalence. The results showed that: 1) antibody functional affinity influenced O2- production and the complement-fixing activity induced by the IC. In general, the higher functional affinity antibodies were more efficient in stimulating the respiratory burst of neutrophils and in activating complement by the classical and alternative pathways than the lower functional affinity antibodies at all IC concentrations tested; 2) complement components incorporated into the immune complex lattice caused an increase in the stimulatory activity of both IgG antibodies to produce O2- (approximately equal to 15% for the IC of IgG with Ka = 5 x 10(8) M(-1) and approximately equal to 7% for the IC of IgG with Ka = 2 x 10(7) M(-1)). This effect was dependent on antibody affinity and concentration; 3) there was a direct relationship between the overall level of complement activation, antibody affinity and superoxide production by neutrophils. Thus, we conclude that antibody affinity influences immune complex lattice formation, modulating its three-dimensional structure and the disposition of Fc fragments interfering with the antibody's biological properties. These results can help understand the precise role of antibody functional affinity in antigen-antibody complex diseases and define the immunochemical characteristics of pathogenic complexes.
authors.affiliation: Department of Biochemistry and Immunology, Faculty of Medicine of Ribeirão Preto, University of São Paulo, Ribeirão Preto, SP, Brazil.
authors.forename: C M O S
authors.initials: CM
authors.lastname: Alves
authors.forename: C M
authors.initials: CM
authors.lastname: Marzocchi-Machado
authors.forename: A E C S
authors.initials: AE
authors.lastname: Azzolini
authors.forename: Y M
authors.initials: YM
authors.lastname: Lucisano-Valim
mesh_terms: D000818:Animals; D000915:Antibody Affinity; D000936:Antigen-Antibody Complex / Q000096:biosynthesis / Q000737:chemistry / Q000276:immunology*; D003167:Complement Activation; D003168:Complement Fixation Tests; D003165:Complement System Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000276:immunology*; D007106:Immune Sera; D007074:Immunoglobulin G / Q000096:biosynthesis / Q000276:immunology* / Q000302:isolation & purification; D009504:Neutrophils / Q000276:immunology*; D010047:Ovalbumin / Q000276:immunology; D011817:Rabbits; D013053:Spectrophotometry; D013481:Superoxides / Q000032:analysis* / Q000276:immunology
chemical_list: D000936:Antigen-Antibody Complex; D007106:Immune Sera; D007074:Immunoglobulin G; D013481:Superoxides; D010047:Ovalbumin; D003165:Complement System Proteins