Rat Mannose-Binding Protein A Binds CD14

Lipopolysaccharide (LPS) has been known to induce inflammation by interacting with CD14, which serves as a receptor for LPS. Mannose-binding protein (MBP) belongs to the collectin subgroup of the C-type lectin superfamily, along with surfactant proteins SP-A and SP-D. We have recently demonstrated that SP-A modulates LPS-induced cellular responses by interaction with CD14 (H. Sano, H. Sohma, T. Muta, S. Nomura, D. R. Voelker, and Y. Kuroki, J. Immunol. 163:387–395, 2000) and that SP-D also interacts with CD14 (H. Sano, H. Chiba, D. Iwaki, H. Sohma, D. R. Voelker, and Y. Kuroki, J. Biol. Chem. 275:22442–22451, 2000). In this study, we examined whether MBP, a collectin highly homologous to SP-A and SP-D, could bind CD14. Recombinant rat MBP-A bound recombinant human soluble CD14 in a concentration-dependent manner. Its binding was not inhibited in the presence of excess mannose or EDTA. MBP-A bound deglycosylated CD14 treated with N-glycosidase F, neuraminidase, and O-glycosidase, indicating that MBP-A interacts with the peptide portion of CD14. Since LPS was also a ligand for the collectins, we compared the characteristics of binding of MBP-A to LPS with those of binding to CD14. MBP-A bound to lipid A from Salmonella enterica serovar Minnesota and rough LPS (S. enterica serovar Minnesota Re595 and Escherichia coli J5, Rc), but not to smooth LPS (E. coli O26:B6 and O111:B4). Unlike CD14 binding, EDTA and excess mannose attenuated the binding of MBP-A to rough LPS. From these results, we conclude that CD14 is a novel ligand for MBP-A and that MBP-A utilizes a different mechanism for CD14 recognition from that for LPS

Expression and characterization of recombinant chicken mannose binding lectin

Mannose binding lectin (MBL) is a serum collagenous C-type lectin that plays an important role in the innate immune protection against pathogens. Previously, human and mouse studies have demonstrated that MBL binds a broad range of pathogens that results in their neutralization through agglutination, enhanced phagocytosis, and/or complement activation via the lectin pathway. The role of MBL in chicken is not well understood although the MBL concentration in serum seems to correlate with protection against infections. To investigate the role of MBL in chicken further, recombinant chicken MBL (RcMBL) was produced in HeLa R19 cells and purified using mannan affinity chromatography followed by gel filtration. RcMBL was shown to be structurally and functionally similar to native chicken MBL (NcMBL) isolated from serum. RcMBL is expressed as an oligomeric protein (mixture of trimers and oligomerized trimers) with a monomeric mass of 26kDa as determined by mass spectrometry, corresponding to the predicted mass. Glycan array analysis indicated that RcMBL bound most strongly to high-mannose glycans but also glycans with terminal fucose and GlcNac residues. The biological activity of RcMBL was demonstrated via its capacity to agglutinate Salmonella Typhimurium and to inhibit the hemagglutination activity of influenza A virus. The production of a structurally well-characterized and functionally active RcMBL will facilitate detailed studies into the protective role of MBL in innate defense against pathogens in chicken and other avian species

Lack of Association between Mannose Binding Lectin and Antibody Responses after Acellular Pertussis Vaccinations

BACKGROUND: Mannose-binding lectin (MBL) is one of the key molecules in innate immunity and its role in human vaccine responses is poorly known. This study aimed to investigate the possible association of MBL polymorphisms with antibody production after primary and booster vaccinations with acellular pertussis vaccines in infants and adolescents. METHODOLOGY/PRINCIPAL FINDINGS: Five hundred and sixty eight subjects were included in this study. In the adolescent cohort 355 subjects received a dose of diphtheria and tetanus toxoids and acellular pertussis (dTpa) vaccine ten years previously. Follow-up was performed at 3, 5 and 10 years. Infant cohort consisted of 213 subjects, who had received three primary doses of DTaP vaccine at 3, 5, and 12 months of age according to Finnish immunization program. Blood samples were collected before the vaccinations at 2,5 months of age and after the vaccinations at 13 months and 2 years of age. Concentrations of IgG antibodies to pertussis toxin, filamentous hemagglutinin, and pertactin and antibodies to diphtheria and tetanus toxoids were measured by standardized enzyme-linked immunosorbant assay. Single nucleotide polymorphisms of MBL2 gene exon1 (codons 52, 54, 57) were examined. MBL serum concentration was also measured from the adolescent cohort. No association was found with MBL2 exon 1 polymorphisms and antibody responses against vaccine antigens, after primary and booster dTpa vaccination. CONCLUSIONS: This study indicates that MBL polymorphisms do not affect the production and persistence of antibodies after acellular pertussis vaccination. Our finding also suggests that MBL might not be involved in modulating antibody responses to the vaccines made of purified bacterial proteins