Identifying genetic susceptibility factors for tuberculosis in Africans: a combined approach using a candidate gene study and a genome-wide screen.
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There is convincing evidence that host genes affect the outcome of infection in human tuberculosis. Two complementary strategies were used to identify the genes involved. A linkage-based genome-wide screen was carried out to locate the positions of genes exerting a major population-wide effect on tuberculosis susceptibility. A candidate-gene-based case-control study was used to examine the effects of genes that may exert a more moderate effect on risk of clinical tuberculosis. The genome screen was conducted in two stages. In the first stage 299 microsatellite markers, spanning all 23 chromosomes, were typed in 92 independent sib-pairs, and seven regions showed some evidence of co-segregation with the disease. These seven regions were examined in a second set of 81 sib-pairs, and markers on chromosomes 15q and Xq showed evidence of linkage to tuberculosis. An X chromosome susceptibility gene may contribute to the excess of males with tuberculosis observed in many populations. The candidate gene approach compared the frequency of polymorphisms in several genes in over 400 subjects with smear-positive pulmonary tuberculosis and 400 ethnically matched healthy controls. Polymorphisms in genes encoding natural-resistance-associated macrophage protein, vitamin D receptor and mannose-binding lectin were associated with tuberculosis. These results suggest that many genes may be involved in determining host susceptibility to tuberculosis, and highlight the importance of using several different study methods to locate them. (+info)
Trypanosoma cruzi-cardiomyocytes: new contributions regarding a better understanding of this interaction.
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The present paper summarizes new approaches regarding the progress done to the understanding of the interaction of Trypanosoma cruzi-cardiomyocytes. Mannose receptors localized at the surface of heart muscle cell are involved in binding and uptake of the parasite. One of the most striking events in the parasite-heart muscle cells interaction is the disruption of the actin cytoskeleton. We have investigated the regulation of the actin mRNA during the cytopathology induced in myocardial cells by the parasite. T. cruzi invasion increases calcium resting levels in cardiomyocytes. We have previously shown that Ca2+ ATPase of the sarcoplasmic reticulum (SERCA) is involved in the invasion of T. cruzi in cardiomyocytes. Treating the cells with thapsigargin, a drug that binds to all SERCA ATPases and causes depletion of intracellular calcium stores, we found a 75% inhibition in the T. cruzi-cardiomyocytes invasion. (+info)
Structural characterisation of the native fetuin-binding protein Scilla campanulata agglutinin: a novel two-domain lectin.
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The three-dimensional structure of a 244-residue, multivalent, fetuin-binding lectin, SCAfet, isolated from bluebell (Scilla campanulata) bulbs, has been solved at 3.3 A resolution by molecular replacement using the coordinates of the 119-residue, mannose-binding lectin, SCAman, also from bluebell bulbs. Unlike most monocot mannose-binding lectins, such as Galanthus nivalis agglutinin from snowdrop bulbs, which fold into a single domain, SCAfet contains two domains with approximately 55% sequence identity, joined by a linker peptide. Both domains are made up of a 12-stranded beta-prism II fold, with three putative carbohydrate-binding sites, one on each subdomain. SCAfet binds to the complex saccharides of various animal glycoproteins but not to simple sugars. (+info)
Mannose receptor-mediated gene transfer into macrophages using novel mannosylated cationic liposomes.
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A novel mannosylated cholesterol derivative, cholesten-5-yloxy-N-(4-((1-imino-2-beta-D-thiomannosyl -ethyl)amino)bu tyl) formamide (Man-C4-Chol), was synthesized in order to perform mannose receptor-mediated gene transfer with liposomes. Plasmid DNA encoding luciferase gene (pCMV-Luc) complexed with liposomes, consisting of a 6:4 mixture of Man-C4-Chol and dioleoylphosphatidylethanolamine (DOPE), showed higher transfection activity than that complexed with 3beta[N-(N', N'-dimethylaminoethane)-carbamoyl]cholesterol (DC-Chol)/DOPE(6:4) and N-[1-(2,3-dioleyloxy)propyl]-N,N,N-trimethylammonium chloride (DOTMA)/DOPE(1:1) liposomes in mouse peritoneal macrophages. The presence of 20 mM mannose significantly inhibited the transfection efficiency of pCMV-Luc complexed with Man-C4-Chol/DC- Chol/DOPE(3:3:4) and Man-C4-Chol/DOPE(6:4) liposomes. High gene expression of pCMV-Luc was observed in the liver after intravenously injecting mice with Man-C4-Chol/DOPE(6:4) liposomes, whereas DC-Chol/DOPE(6:4) liposomes only showed marked expression in the lung. The gene expression with Man-C4-Chol/DOPE(6:4) liposome/ DNA complexes in the liver was observed preferentially in the non-parenchymal cells and was significantly reduced by predosing with mannosylated bovine serum albumin. The gene expression in the liver was greater following intraportal injection. These results suggest that plasmid DNA complexed with mannosylated liposomes exhibits high transfection activity due to recognition by mannose receptors both in vitro and in vivo. Gene Therapy (2000) 7, 292-299. (+info)
Macrophage-induced lung inflammation contributes substantially to respiratory failure during Pneumocystis carinii pneumonia. We isolated a P. carinii cell wall fraction rich in glucan carbohydrate, which potently induces TNF-alpha and macrophage-inflammatory protein-2 generation from alveolar macrophages. Instillation of this purified P. carinii carbohydrate cell wall fraction into healthy rodents is accompanied by substantial increases in whole lung TNF-alpha generation and is associated with neutrophilic infiltration of the lungs. Digestion of the P. carinii cell wall isolate with zymolyase, a preparation containing predominantly beta-1,3 glucanase, substantially reduces the ability of this P. carinii cell wall fraction to activate alveolar macrophages, thus suggesting that beta-glucan components of the P. carinii cell wall largely mediate TNF-alpha release. Furthermore, the soluble carbohydrate beta-glucan receptor antagonists laminariheptaose and laminarin also substantially reduce the ability of the P. carinii cell wall isolate to stimulate macrophage-inflammatory activation. In contrast, soluble alpha-mannan, a preparation that antagonizes macrophage mannose receptors, had minimal effect on TNF-alpha release induced by the P. carinii cell wall fraction. P. carinii beta-glucan-induced TNF-alpha release from alveolar macrophages was also inhibited by both dexamethasone and pentoxifylline, two pharmacological agents with potential activity in controlling P. carinii-induced lung inflammation. These data demonstrate that P. carinii beta-glucan cell wall components can directly stimulate alveolar macrophages to release proinflammatory cytokines mainly through interaction with cognate beta-glucan receptors on the phagocyte. (+info)
Crystal structure of the cysteine-rich domain of mannose receptor complexed with a sulfated carbohydrate ligand.
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The macrophage and epithelial cell mannose receptor (MR) binds carbohydrates on foreign and host molecules. Two portions of MR recognize carbohydrates: tandemly arranged C-type lectin domains facilitate carbohydrate-dependent macrophage uptake of infectious organisms, and the NH(2)-terminal cysteine-rich domain (Cys-MR) binds to sulfated glycoproteins including pituitary hormones. To elucidate the mechanism of sulfated carbohydrate recognition, we determined crystal structures of Cys-MR alone and complexed with 4-sulfated-N-acetylgalactosamine at 1.7 and 2.2 A resolution, respectively. Cys-MR folds into an approximately three-fold symmetric beta-trefoil shape resembling fibroblast growth factor. The sulfate portions of 4-sulfated-N-acetylgalactosamine and an unidentified ligand found in the native crystals bind in a neutral pocket in the third lobe. We use the structures to rationalize the carbohydrate binding specificities of Cys-MR and compare the recognition properties of Cys-MR with other beta-trefoil proteins. (+info)
The cysteine-rich domain of the macrophage mannose receptor is a multispecific lectin that recognizes chondroitin sulfates A and B and sulfated oligosaccharides of blood group Lewis(a) and Lewis(x) types in addition to the sulfated N-glycans of lutropin.
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The mannose receptor (MR) is an endocytic protein on macrophages and dendritic cells, as well as on hepatic endothelial, kidney mesangial, tracheal smooth muscle, and retinal pigment epithelial cells. The extracellular portion contains two types of carbohydrate-recognition domain (CRD): eight membrane-proximal C-type CRDs and a membrane-distal cysteine-rich domain (Cys-MR). The former bind mannose-, N-acetylglucosamine-, and fucose-terminating oligosaccharides, and may be important in innate immunity towards microbial pathogens, and in antigen trapping for processing and presentation in adaptive immunity. Cys-MR binds to the sulfated carbohydrate chains of pituitary hormones and may have a role in hormonal clearance. A second feature of Cys-MR is binding to macrophages in marginal zones of the spleen, and to B cell areas in germinal centers which may help direct MR-bearing cells toward germinal centers during the immune response. Here we describe two novel classes of carbohydrate ligand for Cys-MR: chondroitin-4 sulfate chains of the type found on proteoglycans produced by cells of the immune system, and sulfated blood group chains. We further demonstrate that Cys-MR interacts with cells in the spleen via the binding site for sulfated carbohydrates. Our data suggest that the three classes of sulfated carbohydrate ligands may variously regulate the trafficking and function of MR-bearing cells. (+info)
Structure of a C-type carbohydrate recognition domain from the macrophage mannose receptor.
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The mannose receptor of macrophages and liver endothelium mediates clearance of pathogenic organisms and potentially harmful glycoconjugates. The extracellular portion of the receptor includes eight C-type carbohydrate recognition domains (CRDs), of which one, CRD-4, shows detectable binding to monosaccharide ligands. We have determined the crystal structure of CRD-4. Although the basic C-type lectin fold is preserved, a loop extends away from the core of the domain to form a domain-swapped dimer in the crystal. Of the two Ca(2+) sites, only the principal site known to mediate carbohydrate binding in other C-type lectins is occupied. This site is altered in a way that makes sugar binding impossible in the mode observed in other C-type lectins. The structure is likely to represent an endosomal form of the domain formed when Ca(2+) is lost from the auxiliary calcium site. The structure suggests a mechanism for endosomal ligand release in which the auxiliary calcium site serves as a pH sensor. Acid pH-induced removal of this Ca(2+) results in conformational rearrangements of the receptor, rendering it unable to bind carbohydrate ligands. (+info)