Antibody-mediated activation of the classical pathway of complement may compensate for mannose-binding lectin deficiency (original) (raw)
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Functional characterization of the lectin pathway of complement in human serum
Molecular Immunology
Mannan-binding lectin (MBL) is a major initiator of the lectin pathway (LP) of complement. Polymorphisms in exon 1 of the MBL gene are associated with impaired MBL function and infections. Functional assays to assess the activity of the classical pathway (CP) and the alternative pathway (AP) of complement in serum are broadly used in patient diagnostics. We have now developed a functional LP assay that enables the specific quantification of autologous MBL-dependent complement activation in human serum. Complement activation was assessed by ELISA using coated mannan to assess the LP and coated IgM to assess the CP. Normal human serum (NHS) contains IgG, IgA and IgM antibodies against mannan, as shown by ELISA. These antibodies are likely to induce CP activation. Using C1q-blocking and MBL-blocking mAb, it was confirmed that both the LP and the CP contribute to complement activation by mannan. In order to quantify LP activity without interference of the CP, LP activity was measured in...
Relative Roles of Complement Factor 3 and Mannose-Binding Lectin in Host Defense against Infection
Infection and Immunity, 2005
Staphylococcus aureus is a major cause of severe nosocomial and community-acquired infections. Phagocytes and humoral molecules, including complement, have been proposed to cooperate in host defense against gram-positive bacteria. Circumstantial evidence indicates a role for complement, but this has not been formally defined. Complement activation is initiated by the classical, alternative, or lectin pathway, with the latter requiring mannose-binding lectin (MBL, also known as mannose-binding protein). MBL is an oligomeric serum protein that recognizes carbohydrates decorating a broad range of infectious agents, including S. aureus. We previously reported that MBL null mice were highly susceptible to S. aureus infection, confirming that MBL plays a key role in first-line host defense. In this study, we evaluated the relative roles of C3 and MBL against S. aureus infection by generating MBL × C3 null mice to compare with C3 single null mice. C3 deficiency alone significantly reduced ...
International Journal of Immunogenetics, 2007
Mannose-binding lectin (MBL) mediates activation of the complement system via the lectin pathway. Two forms of MBL, MBL-A and MBL-C, were characterized in rodents, rabbits, bovine and rhesus monkeys, whereas only one form was identified in humans, chimpanzees and chickens. The two forms are encoded by two distinct genes named MBL1 and MBL2, which have been identified in many species including the pig. In this report, we studied the two porcine genes MBL1 and MBL2. The porcine MBL genes had higher identities to bovine rather than primate and rodent sequences. Both genes were assigned to chromosome 14 by radiation hybrid panel and linkage mapping. Both MBL genes were highly expressed in liver. MBL1 was also found to be expressed in the lung, testis and brain, whereas low expression of MBL2 was detected in the testis and kidney. New single nucleotide polymorphisms of porcine MBL2 gene were found and genotyped in an experimental F2 pig population, together with a previously reported SNP of MBL1. MBL1 genotypes differed in C3c serum concentration, i.e. in vivo complement activity, at P < 0.1. Correspondingly, linkage analysis revealed a quantitative trait locus for C3c serum level close to the position of the MBL genes. The study thus promotes the porcine MBL genes as functional and positional candidate gene for complement activity.
Human IgA Activates the Complement System Via the Mannan-Binding Lectin Pathway1
2000
The recently identified lectin pathway of the complement system, initiated by binding of mannan-binding lectin (MBL) to its ligands, is a key component of innate immunity. MBL-deficient individuals show an increased susceptibility for infections, espe- cially of the mucosal system. We examined whether IgA, an important mediator of mucosal immunity, activates the complement system via the lectin pathway. Our results
Complement Receptor 1/Cd35 Is a Receptor for Mannan-Binding Lectin
Journal of Experimental Medicine, 2000
Mannan-binding lectin (MBL), a member of the collectin family, is known to have opsonic function, although identification of its cellular receptor has been elusive. Complement C1q, which is homologous to MBL, binds to complement receptor 1 (CR1/CD35), and thus we investigated whether CR1 also functions as the MBL receptor. Radioiodinated MBL bound to recombinant soluble CR1 (sCR1) that had been immobilized on plastic with an apparent equilibrium dissociation constant of 5 nM. N -acetyl-D -glucosamine did not inhibit sCR1-MBL binding, indicating that the carbohydrate binding site of MBL is not involved in binding CR1. C1q inhibited MBL binding to immobilized sCR1, suggesting that MBL and C1q might bind to the same or adjacent sites on CR1. MBL binding to polymorphonuclear leukocytes (PMNs) was associated positively with changes in CR1 expression induced by phorbol myristate acetate. Finally, CR1 mediated the adhesion of human erythrocytes to immobilized MBL and functioned as a phagocytic receptor on PMNs for MBL-immunoglobulin G opsonized bacteria. Thus, MBL binds to both recombinant sCR1 and cellular CR1, which supports the role of CR1 as a cellular receptor for the collectin MBL. 1 Abbreviations used in this paper: CR1, complement receptor 1; CRD, carbohydrate recognition domain; C1qRp, C1q receptor of phagocytosis; LHR, long homologous repeat; MASP, MBL-associated serine protease; MBL, mannan-binding lectin; MFC, mean fluorescent channel; sCR1, soluble CR1; TBS, Tris-buffered saline.
Journal of Immunological Methods, 2009
Mannose-binding lectin (MBL), L-ficolin and H-ficolin are human serum lectins, all of which form complexes with MBL-associated serine proteases (MASP). The lectin-MASP complexes bind to the surface of microbes, leading to activation of the lectin pathway of complement. Enzyme-linked immunosorbent assays (ELISA) of the lectin pathway activity reported so far determined the activity via either MBL or L-ficolin, but an assay of activity via plural host defense lectins has not been established. To measure the lectin pathway activation mediated by plural lectins simultaneously, we developed an ELISA system in which N-acetylglucosamine-pentamer conjugated to dipalmitoylphosphatidylethanolamine (GN5-DPPE) was employed as a ligand for the lectins. In our ELISA system, both purified MBL and L-ficolin isolated from serum diluted in a buffer containing high ionic NaCl bound to GN5-DPPE and activated C4. Purified H-ficolin was not capable of binding to GN5-DPPE. MBL and L-ficolin in MBL-sufficient serum also bound to GN5-DPPE and activated C4. Mannose and N-acetylgalactosamine inhibited binding of MBL and L-ficolin to GN5-DPPE, respectively. MBL-deficient serum that had been depleted of L-ficolin did not exhibit C4 activation, but addition of both or either purified MBL and/or L-ficolin to the serum restored the activation in a dose-dependent manner. Thus, C4 cleaving activity could be evaluated with the coexistence of MBL and L-ficolin in vitro. In conclusion, we propose a novel method using GN5-DPPE for investigating the MBL-and L-ficolin-dependent lectin pathway and anticipate that this method will be useful in innate immunity and clinical research.