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ABSTRACT: We have recently identified FIBCD1 (Fibrinogen C domain containing 1) as a type II transmembrane endocytic receptor located primarily in the intestinal brush border. The ectodomain of FIBCD1 comprises a coiled coil, a polycationic region, and a C-terminal FReD (fibrinogen-related domain) that assembles into disulfide-linked homotetramers. The FIBCD1-FReD binds Ca(2+) dependently to acetylated structures like chitin, N-acetylated carbohydrates, and amino acids. FReDs are present in diverse innate immune pattern recognition proteins including the ficolins and horseshoe crab TL5A. Here, we use chemical cross-linking, combined with analytical ultracentrifugation and electron microscopy of the negatively stained recombinant FIBCD1-FReD to show that it assembles into noncovalent tetramers in the absence of the coiled coil. We use surface plasmon resonance, carbohydrate binding, and pulldown assays combined with site-directed mutagenesis to define the binding site involved in the interaction of FIBCD1 with acetylated structures. We show that mutations of central residues (A432V and H415G) in the hydrophobic funnel (S1) abolish the binding of FIBCD1 to acetylated bovine serum albumin and chitin. The double mutations (D393N/D395A) at the putative calcium-binding site reduce the ability of FIBCD1 to bind ligands. We conclude that the FReDs of FIBCD1 forms noncovalent tetramers and that the acetyl-binding site of FReDs of FIBCD1 is homologous to that of tachylectin 5A and M-ficolin but not to the FReD of L-ficolin. We suggest that the spatial organization of the FIBCD1-FReDs determine the molecular pattern recognition specificity and subsequent biological functions.
Journal of Biological Chemistry 11/2009; 285(2):1229-38. · 4.77 Impact Factor
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ABSTRACT: Collectins are collagenous lectins present in blood, respiratory lining fluid, and other mucosal secretions that play important roles in innate defense against infection. The collectin, surfactant protein D (SP-D), limits infection by viruses and bacteria in the respiratory tract, eye, and female genital tract. Multimeric SP-D has strong antiviral activity and is a potent viral and bacterial agglutinin and opsonin; however, trimers composed of the neck and carbohydrate recognition domain (hSP-D-NCRD) of SP-D lack these activities. We now show that, in contrast, a trimeric neck and CRD construct of bovine serum collectin CL-46 induces aggregation of influenza A virus (IAV) and potently increases IAV uptake by neutrophils. CL-46-NCRD showed calcium-dependent and sugar-sensitive binding to both neutrophils and IAV. Replacement of specific residues of the CRD of human SP-D with those found in bovine serum collectins conferred opsonizing activity. The most effective substitution involved replacement of arginine 343 with valine (hSP-D-NCRD/R343V). hSP-D-NCRD/R343V greatly increased viral uptake by neutrophils and monocytes and also potentiated neutrophil respiratory burst responses. These effects were further increased by cross-linking of hSP-D-NCRD/R343V trimers with MAbs directed against areas of the hSP-D-NCRD not involved in viral binding. Unlike the wild-type human SP-D hSP-D-NCRD, hSP-D-NCRD/R343V also induced viral aggregation. These results indicate that collectins can act as opsonins for IAV even in the absence of the collagen domain or higher order multimerization. This may involve increased affinity of individual CRDs for glycoconjugates displayed on host cells or the viral envelope.
AJP Lung Cellular and Molecular Physiology 10/2009; 298(1):L79-88. · 3.66 Impact Factor
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ABSTRACT: Chitin is a highly acetylated compound and the second most abundant biopolymer in the world next to cellulose. Vertebrates are exposed to chitin both through food ingestion and when infected with parasites, and fungi and chitin modulate the immune response in different directions. We have identified a novel homotetrameric 55-kDa type II transmembrane protein encoded by the FIBCD1 gene and highly expressed in the gastrointestinal tract. The ectodomain of FIBCD1 is characterized by a coiled-coil region, a polycationic region and C-terminal fibrinogen-related domain that by disulfide linkage assembles the protein into tetramers. Functional analysis showed a high-affinity and calcium-dependent binding of acetylated components to the fibrinogen domain, and a function in endocytosis was demonstrated. Screening for ligands revealed that the FIBCD1 is a high-affinity receptor for chitin and chitin fragments. FIBCD1 may play an important role in controlling the exposure of intestine to chitin and chitin fragments, which is of great relevance for the immune defense against parasites and fungi and for immune response modulation.
The Journal of Immunology 09/2009; 183(6):3800-9. · 5.79 Impact Factor
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ABSTRACT: Erythrocytes (E) from a cross-sectional group of 22 outpatients with systemic lupus erythematosus (SLE) and/or mixed connective tissue disease (MCTD), the majority without active disease (n = 14), were analyzed for CR1 antigen expression and capacity to bind complement opsonized, radiolabeled immune complexes (IC). Furthermore, E-bound C3 fragments and the plasma C3d concentration were determined. E-bound C3b/iC3b fragments were not elevated in patients with SLE, whereas E from 11 out of 22 SLE patients had increased C3d levels which correlated with the plasma C3d concentration (Rs 0.73, p < 0.001). E-fixed C3d fragments did not affect the binding of Mab or preopsonized IC to E-CR1 and were not correlated with disease activity or medical treatment. Antigen expression of E-CR1 measured by ELISA or agglutination showed positive correlation with the IC binding capacity of E-CR1 (Rs 0.92 and 0.72 respectively, p < 001). The IC binding capacity of E-CR1 from SLE patients was significantly reduced (p > 0.005), whereas the antigen expression of CR1 (ELISA) on E from the patients did not differ from that of E from healthy donors (p > 0.1). E-CR1 antigen was measured by Mab reacting with an epitope outside the IC-binding site of E-CR1. E-CR1 antigen expression or IC binding showed no correlation either with disease activity or prednisolone treatment. However, 4 og 5 patients with MCTD and 4 of 5 patients receiving Imurel® were found to have low E-CR1 expression and capacity to bind IC. Thus, measurement of antigenic E-CR1 in a cross-sectional group of SLE outpatients by use of Mab reacting with an epitope outside the ligand-binding region of CR1 did not reveal a significantly reduced CR1 expression. However, an assay for CR 1 -mediated IC binding showed a clearly reduced E-CR1 function.
Apmis 08/2009; 98(7‐12):637 - 644. · 1.99 Impact Factor
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ABSTRACT: Surfactant protein-D (SP-D) is a calcium dependent lectin in the innate immune system that facilitates clearance of microbes. The protein is associated with mucosal surfaces, and also found in bronchoalveolar lavage, serum and amniotic fluid. Human SP-D includes trimeric subunits and multimeric assemblies of trimeric subunits, which are stabilized by N-terminal interchain disulfide crosslinks. An N-terminal structural polymorphism (Met11Thr) and associated O-glycosylation are previously shown accompanied by incomplete multimerization and with a relative low proportion of multimeric Thr11 SP-D compared to Met11 SP-D. Multimerization has proven important for enhancement of microbial phagocytosis. In the present study defined multimeric forms of Met11Thr SP-D were isolated from human amniotic fluid. Implementation of ManNAc-affinity chromatography allowed high recovery of natural trimeric SP-D subunits. However, affinity chromatography increased the relative proportion of multimers at the expense of natural trimeric subunits. Multimeric SP-D partially disassembled to form trimeric subunits. The resulting distribution of structural forms was independent of the Met11Thr genotype. Trimeric and multimeric SP-D appeared with distinct patterns of disulphide crosslinking, which partly changed according to interconversion between the structural forms. Solid phase assays demonstrated that trimeric SP-D subunits showed greater binding to LPS and PGN, but lower binding to mannan and LTA, than SP-D multimers. Trimeric SP-D subunits also showed greater binding to endogenous lipoproteins: LDL, oxLDL, and HDL, than multimeric SP-D. In conclusion, purified trimeric and multimeric SP-D represent separate and only partly interconvertible molecular populations with distinct biochemical properties.
Molecular Immunology 08/2009; 46(15):3060-9. · 2.90 Impact Factor
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ABSTRACT: Surfactant protein D (SP-D) is an oligomeric calcium-dependent lectin with important roles in innate host defence against infectious microorganisms. Several studies have shown that patients with inflammatory lung disease have elevated levels of circulating SP-D, and serum SP-D has been suggested to be used as a biomarker for disease e.g. in COPD. We aimed to investigate the variation of circulating SP-D in healthy individuals in and between days for 6 months. In addition, we studied the SP-D response to a standardized physical exercise programme. SP-D was measured in serum using a 5-layered ELISA technique. We found that circulating SP-D remained constant over a 6-month period. However, during the course of one day SP-D varied significantly. Median SP-D peaked at 10 a.m. at 1009 ng/ml (95% CI: 803-1497), subsequently decreasing to nadir at 10 p.m. at 867 ng/ml (95% CI: 650-1148)(P<0.00005). Median pre-exercise level of SP-D was 746 ng/ml (95% CI: 384-2035), and immediately after cessation of physical activity the median SP-D level was 767 ng/ml (95% CI: 367-1885) (P=0.248). Our findings underscore the importance of standardized blood sampling conditions in future studies on the potential role of SP-D as a biomarker. Importantly, stable measures of systemic SP-D over a prolonged period support that SP-D is suitable for biomarker studies.
Immunobiology 06/2009; 215(4):314-20. · 3.20 Impact Factor
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ABSTRACT: We have cloned and characterized a novel murine transmembrane molecule, mSCART1 belonging to the scavenger receptor cysteine-rich superfamily. The cDNA encodes a polypeptide chain of 989 amino acids, organized as a type I transmembrane protein that contains eight extracellular SRCR domains followed by a transmembrane region and a cytoplasmic domain. The cytoplasmic domain contains two putative src kinase consensus substrate sequences, three additional potential phosphorylation sites, and two potential internalization motifs. Two possible secreted forms that lack the transmembrane region arise by alternative splicing. The murine SCART1 gene maps to chromosome 7 band F5 and the analysis of the genomic organization showed that the gene spans 12.86 kb and contains 14 exons. Quantitative real-time PCR analyses on murine tissues showed high mSCART1 mRNA expression in the lymph node, the trachea, and the lung, and low expression was found in the thymus, the spleen, the skin, and in tissues throughout the gastrointestinal tract. Comparative studies of the domain organization as well as the cytoplasmic domain of mSCART1 with the other members of the SRCR superfamily show that mSCART1 is highly related to the WC1 family of the SRCR superfamily. Finally, a novel human scavenger receptor cysteine-rich molecule with high homology to mSCART1 was identified by searching in the human genomic databases using the mSCART1 cDNA sequence.
Molecular Immunology 04/2009; 46(8-9):1663-72. · 2.90 Impact Factor
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Caroline End,
Floris Bikker,
Marcus Renner,
Gaby Bergmann,
Stefan Lyer,
Stephanie Blaich,
Melanie Hudler,
Burkhard Helmke,
Nikolaus Gassler,
Frank Autschbach, [......],
Peter Schirmacher,
Arie V Nieuw Amerongen,
Philip Rosenstiel,
Christian Sina,
Andre Franke,
Mathias Hafner,
Petra Kioschis,
Stefan Schreiber,
Annemarie Poustka,
Jan Mollenhauer
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ABSTRACT: Deleted in malignant brain tumors 1 (DMBT1) is a secreted glycoprotein displaying a broad bacterial-binding spectrum. Recent functional and genetic studies linked DMBT1 to the suppression of LPS-induced TLR4-mediated NF-kappaB activation and to the pathogenesis of Crohn's disease. Here, we aimed at unraveling the molecular basis of its function in mucosal protection and of its broad pathogen-binding specificity. We report that DMBT1 directly interacts with dextran sulfate sodium (DSS) and carrageenan, a structurally similar sulfated polysaccharide, which is used as a texturizer and thickener in human dietary products. However, binding of DMBT1 does not reduce the cytotoxic effects of these agents to intestinal epithelial cells in vitro. DSS and carrageenan compete for DMBT1-mediated bacterial aggregation via interaction with its bacterial-recognition motif. Competition and ELISA studies identify poly-sulfated and poly-phosphorylated structures as ligands for this recognition motif, such as heparansulfate, LPS, and lipoteichoic acid. Dose-response studies in Dmbt1(-/-) and Dmbt1(+/+) mice utilizing the DSS-induced colitis model demonstrate a differential response only to low but not to high DSS doses. We propose that DMBT1 functions as pattern-recognition molecule for poly-sulfated and poly-phosphorylated ligands providing a molecular basis for its broad bacterial-binding specificity and its inhibitory effects on LPS-induced TLR4-mediated NF-kappaB activation.
European Journal of Immunology 03/2009; 39(3):833-42. · 5.10 Impact Factor
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ABSTRACT: Surfactant protein D (SP-D) mediates clearance of microorganisms and modulates inflammation in response to cytotoxic stimulation. It is present in various epithelia, but also in vascular smooth muscle and endothelial cells. Experiments were designed to determine whether or not SP-D is present in porcine coronary arterial endothelial cells and if so, to investigate the molecular mechanisms underlying this presence. The expression of SP-D, NO synthase, Akt 1/2 and Erk 1/2 proteins was determined in cultures at passages 1 (#1) and 4 (#4). SP-D in primary cells existed in three isoforms (37-38 kDa and 50 kDa). The 37-38 kDa SP-D forms were the dominant isoforms in the porcine endothelium and were prominent at #1 but partially lost at #4. Tumor necrosis factor-alpha (TNF-alpha) significantly augmented the level of SP-D expression at #1 but not at #4. The basal level of 37-38 kDa SP-D isoforms at #1 was reduced by L-NAME, wortmannin and PD 98059. The low basal expression at #4 could be increased by DETA NONOate (donor of NO) or insulin (activator of PI(3)K/Akt). The presence of nitric oxide synthase was reduced while that of Akt 1/2 and Erk 1/2 was increased at #4. In cells both at passages 1 and 4, TNF-alpha downregulated NO synthase and up-regulated p-Erk 1/2 protein. The present findings demonstrate the presence of SP-D in endothelial cells which is NO-, PI(3)K/Akt- and Erk-dependent. They suggest a protective role of SP-D in these cells.
Molecular Immunology 12/2008; 46(6):1050-7. · 2.90 Impact Factor
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Christian Mölleken,
Barbara Sitek,
Corinna Henkel,
Gereon Poschmann,
Bence Sipos,
Sebastian Wiese,
Bettina Warscheid,
Christoph Broelsch,
Markus Reiser,
Scott L Friedman,
Ida Tornøe,
Anders Schlosser,
Günter Klöppel,
Wolff Schmiegel,
Helmut E Meyer, Uffe Holmskov,
Kai Stühler
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ABSTRACT: Hepatic cirrhosis is a life-threatening disease arising from different chronic liver disorders. One major cause for hepatic cirrhosis is chronic hepatitis C. Chronic hepatitis C is characterized by a highly variable clinical course, with at least 20% developing liver cirrhosis within 40 years. Only liver biopsy allows a reliable evaluation of the course of hepatitis C by grading inflammation and staging fibrosis, and thus serum biomarkers for hepatic fibrosis with high sensitivity and specificity are needed. To identify new candidate biomarkers for hepatic fibrosis, we performed a proteomic approach of microdissected cirrhotic septa and liver parenchyma cells. In cirrhotic septa, we detected an increasing expression of cell structure associated proteins, including actin, prolyl 4-hydroxylase, tropomyosin, calponin, transgelin, and human microfibril-associated protein 4 (MFAP-4). Tropomyosin, calponin, and transgelin reflect a contribution of activated stellate cells/myofibroblasts to chronic liver injury. The expression of tropomyosin, transgelin, and MFAP-4, an extracellular matrix associated protein, were further evaluated by immunohistochemistry. Tropomyosin and MFAP-4 demonstrated high serum levels in patients with hepatic cirrhosis of different causes. CONCLUSION: A quantitative analysis of MFAP-4 serum levels in a large number of patients showed MFAP-4 as novel candidate biomarker with high diagnostic accuracy for prediction of nondiseased liver versus cirrhosis [area under receiver operating characteristic curve (AUC) = 0.97, P < 0.0001] as well as stage 0 versus stage 4 fibrosis (AUC = 0.84, P < 0.0001), and stages 0 to 3 versus stage 4 fibrosis (AUC = 0.76, P < 0.0001).
Hepatology 12/2008; 49(4):1257-66. · 11.66 Impact Factor
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ABSTRACT: Collectins are multimeric host defence lectins with trimeric CRDs (carbohydrate-recognition domains) and collagen and N-terminal domains that form higher-order structures composed of four or more trimers. Recombinant trimers composed of only the CRD and adjacent neck domain (termed NCRD) retain binding activity for some ligands and mediate some functional activities. The lung collectin SP-D (surfactant protein D) has strong neutralizing activity for IAVs (influenza A viruses) in vitro and in vivo, however, the NCRD derived from SP-D has weak viral-binding ability and lacks neutralizing activity. Using a panel of mAbs (monoclonal antibodies) directed against the NCRD in the present study we show that mAbs binding near the lectin site inhibit antiviral activity of full-length SP-D, but mAbs which bind other sites on the CRD do not. Two of the non-blocking mAbs significantly increased binding and antiviral activity of NCRDs as assessed by haemagglutination and neuraminidase inhibition and by viral neutralization. mAb-mediated cross-linking also enabled NCRDs to induce viral aggregation and to increase viral uptake by neutrophils and virus-induced respiratory burst responses by these cells. These results show that antiviral activities of SP-D can be reproduced without the N-terminal and collagen domains and that cross-linking of NCRDs is essential for antiviral activity of SP-D with respect to IAV.
Biochemical Journal 07/2008; 412(2):323-9. · 4.90 Impact Factor
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ABSTRACT: Collectin placenta-1 (CL-P1), also known as scavenger receptor with C-type lectin (SRCL), is a type II membrane glycoprotein that shares structural features with both collectins and type A scavenger receptors. CL-P1 was originally cloned from the placenta and found to be associated with endothelial cells. It binds via its lectin domain to desialyated Lewis X containing glycoproteins and it is able to facilitate internalization of bound ligands. Via positively charged residues in the collagen-like region it binds to negatively charged components of microbial membranes. It has previously been proposed that CL-P1 plays a role in the host defense system and in the clearance of glycoproteins from the blood. With the aims of determining the detailed tissue expression of human CL-P1 we expressed CL-P1 recombinantly in both E. coli and CHO cells, and raised monoclonal antibodies against human CL-P1. Three monoclonal antibodies were characterized and used in immunohistochemical analyses of a panel of cryo- and formalin-fixed sections. We find that CL-P1 mainly associates with cytotrophoblasts and syncytiotrophoblasts of the placenta, alveolar macrophages and to a less degree with macrophage-like and stromal cells of the tonsils. By real-time RT-PCR we verified that the placenta is also the main organ of CL-P1 synthesis. The only source of endothelial cells whereto CL-P1 associates are umbilical cord vein endothelial cells (human umbilical vein endothelial cells, HUVEC). In vitro cultured HUVECs express both the CL-P1 mRNA and show anti-CL-P1 immunoreactivity but CL-P1 locates mainly to the cytosol and not to the membrane of these cells. We conclude that CL-P1 is not a common membrane protein on endothelial cells found in normal tissues under steady state conditions.
Molecular Immunology 07/2008; 45(11):3278-88. · 2.90 Impact Factor
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ABSTRACT: Surfactant protein D (SP-D) is a pattern recognition molecule of the collectin family of C-type lectins. It is found in the airways and at mucosal surfaces. SP-D is part of the innate immune system where it neutralizes and leads to elimination of microorganisms. It regulates the functions of other innate immune cells, such as macrophages and neutrophils. It also modulates the adaptive immune response by interacting with antigen-presenting cells and T cells. Monoclonal anti-mouse-SP-D antibodies were raised from SP-D deficient mice using recombinant SP-D as antigen. Ten monoclonal antibodies were characterized and validated for use in sandwich enzyme-linked immunosorbent assay (ELISA). Based on two of these, we established an ELISA that allows for measurements of mouse SP-D in various body fluids. The final ELISA was optimized and calibrated with a standard of purified recombinant mouse SP-D, which was analyzed by quantitative amino acid analysis. The ELISA was validated with respect to parallelism, recovery, intra- and inter-assay variation. The practical working range was estimated to be 1.9-200 ng/ml. We measured SP-D in lung lavage to an average concentration of 435 ng/ml (3-ml lavage), and in mouse vaginal fluid (1-ml lavage) to an average concentration of 94 ng/ml, but could not detect SP-D in the serum or plasma of healthy mice (<3.8 ng/ml). With this ELISA, it will be possible to study the regulation of SP-D in various mouse models and upon various stimuli.
Journal of Immunological Methods 01/2008; 330(1-2):75-85. · 2.20 Impact Factor
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ABSTRACT: In order to investigate the dynamic strength of the interaction between lung surfactant protein D (SP-D) and different sugars, maltose, mannose, glucose, and galactose, we have used an atomic force microscope to monitor the interaction on a single molecule scale. The experiment is performed by measuring the rupture force when the SP-D-sugar bond is subjected to a continuously increasing force. Under these dynamic conditions, SP-D binds strongest to d-mannose and weakest to maltose and d-galactose. These results differ from equilibrium measurements wherein SP-D exhibits preference for maltose. On the basis of this finding, we propose that the binding of the disaccharide maltose to SP-D, which is energetically stronger than the binding of any of the monosacchrides, alters the structure of the binding site in a way that lowers the dynamic strength of the bond. We conclude that determining the strength of a protein-ligand bond under dynamic stress using an atomic force microscope is possibly more relevant for mimicking the actual nonequilibrium physiological situation in the lungs.
Biochemistry 11/2007; 46(43):12231-7. · 3.42 Impact Factor
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Marcus Renner,
Gaby Bergmann,
Inge Krebs,
Caroline End,
Stefan Lyer,
Frank Hilberg,
Burkhard Helmke,
Nikolaus Gassler,
Frank Autschbach,
Floris Bikker, [......], Uffe Holmskov,
Vito Annese,
Anna Latiano,
Peter Schirmacher,
Arie V Nieuw Amerongen,
Mauro D'Amato,
Petra Kioschis,
Mathias Hafner,
Annemarie Poustka,
Jan Mollenhauer
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ABSTRACT: Impaired mucosal defense plays an important role in the pathogenesis of Crohn's disease (CD), one of the main subtypes of inflammatory bowel disease (IBD). Deleted in malignant brain tumors 1 (DMBT1) is a secreted scavenger receptor cysteine-rich protein with predominant expression in the intestine and has been proposed to exert possible functions in regenerative processes and pathogen defense. Here, we aimed at analyzing the role of DMBT1 in IBD.
We studied DMBT1 expression in IBD and normal tissues by quantitative reverse transcription-polymerase chain reaction, immunohistochemistry, and mRNA in situ hybridization. Genetic polymorphisms within DMBT1 were analyzed in an Italian IBD case-control sample. Dmbt1(-/-) mice were generated, characterized, and analyzed for their susceptibility to dextran sulfate sodium-induced colitis.
DMBT1 levels correlate with disease activity in inflamed IBD tissues. A highly significant fraction of the patients with IBD displayed up-regulation of DMBT1 specifically in the intestinal epithelial surface cells and Paneth cells. A deletion allele of DMBT1 with a reduced number of scavenger receptor cysteine-rich domain coding exons is associated with an increased risk of CD (P = .00056; odds ratio, 1.75) but not for ulcerative colitis. Dmbt1(-/-) mice display enhanced susceptibility to dextran sulfate sodium-induced colitis and elevated Tnf, Il6, and Nod2 expression levels during inflammation.
DMBT1 may play a role in intestinal mucosal protection and prevention of inflammation. Impaired DMBT1 function may contribute to the pathogenesis of CD.
Gastroenterology 11/2007; 133(5):1499-509. · 11.68 Impact Factor
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ABSTRACT: Development of dementia, including Alzheimer's disease (AD), is associated with lipid dysregulation and inflammation. As the host defense lectin surfactant protein D (SP-D) has multiple effects in lipid homeostasis and inflammation, the correlation between SP-D concentrations and development of dementia was investigated. A total of 418 non-demented persons were included in the study and cognitively re-examined after 3 years, while survival was followed for 11 years. Serum SP-D concentrations were measured at baseline. Logistic regression analysis controlling for age, gender, smoking status, and CRP showed that Odds Ratio for developing dementia was 2.62 (1.12-6.15) with an SP-D concentration in the highest quartile compared to the other quartiles. The risk of AD was 2.55 (0.95-6.90). Cox regression controlling for the same variables showed that hazard ratio of death was 1.43 (1.06-1.92) in the highest quartile. SP-D concentration thus correlates to development of dementia as well as to augmented mortality.
Clinical Immunology 07/2007; 123(3):333-7. · 4.05 Impact Factor
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ABSTRACT: Trefoil factors (TTFs) are small, compact proteins coexpressed with mucins in the gastrointestinal tract. Three trefoil factors are known in mammals: TFF1, TFF2, and TFF3. They are implicated to play diverse roles in maintenance and repair of the gastrointestinal channel. We compared the expression pattern of the three trefoil factors analyzing mRNA from a panel of 20 human tissues by conventional reverse transcriptase (RT) PCR and, in addition, by real-time PCR. These findings were supported by immunohistochemical analysis of paraffin-embedded human tissues using rabbit polyclonal antibodies raised against these factors. TFF1 showed highest expression in the stomach and colon, whereas TFF2 and TFF3 showed highest expression in stomach and colon, respectively. All three TFFs were found in the ducts of pancreas. Whereas TFF2 was found to be restricted to these two tissues, the structurally more closely related TFF1 and TFF3 showed a more general tissue distribution and were found to colocalize on an array of mucosal surfaces. This is the first thorough parallel description of the tissue distribution of TFFs in normal tissues, and it provides a baseline for similar analysis in diseased tissues.
Journal of Histochemistry and Cytochemistry 06/2007; 55(5):505-13. · 2.72 Impact Factor
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ABSTRACT: The lectin pathway of the complement system is activated when Mannan-binding lectin (MBL) in complex with MASP-2 binds microorganisms. Polymorphisms in both genes are responsible for low serum levels, which associate with increased risk of infection and autoimmune disease. The present study includes 1215 MBL measurements and 1214 MASP-2 activity measurements in healthy Danish adult twins. Total MASP-2 activity was estimated by C4 cleaving activity of samples diluted in an excess of MBL. Twin-twin correlations were higher in monozygotic (MZ) than in dizygotic (DZ) twins for both traits. Heritabilites of MBL levels and MASP-2 activity were estimated using structural equation modeling allowing assessment of the contribution of common genes affecting both traits. The estimated heritability was 0.77 [95% CI 0.64;0.91] for MBL levels and 0.75 [95% CI 0.59;0.81] for MASP-2 activity with the presence of additive genetic factors, shared environmental factors, and non-shared environmental factors. The genetic correlation, i.e., common genetic factors affecting MBL and MASP-2 activity was estimated to r(g) = 0.34 [0.25;0.42]. The data indicate a strong genetic influence for the serum levels of MBL and for MASP-2 activity with a significant genetic correlation between the two traits.
Genetic Epidemiology 02/2007; 31(1):31-41. · 3.44 Impact Factor
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ABSTRACT: Surfactant proteins A (SP-A) and D (SP-D) have been implicated in pulmonary innate immunity. The proteins are host defense lectins, belonging to the collectin family which also includes mannan-binding lectin (MBL). SP-A and SP-D are pattern-recognition molecules with the lectin domains binding preferentially to sugars on a broad spectrum of pathogen surfaces and thereby facilitating immune functions including viral neutralization, clearance of bacteria, fungi and apoptotic and necrotic cells, modulation of allergic reactions, and resolution of inflammation. SP-A and SP-D can interact with receptor molecules present on immune cells leading to enhanced microbial clearance and modulation of inflammation. SP-A and SP-D also modulate the functions of cells of the adaptive immune system including dendritic cells and T cells. Studies on SP-A and SP-D polymorphisms and protein levels in bronchoalveolar lavage and blood have indicated associations with a multitude of pulmonary inflammatory diseases. In addition, accumulating evidence in mouse models of infection and inflammation indicates that recombinant forms of the surfactant proteins are biologically active in vivo and may have therapeutic potential in controlling pulmonary inflammatory disease. The presence of the surfactant collectins, especially SP-D, in non-pulmonary tissues, such as the gastrointestinal tract and genital organs, suggest additional actions located to other mucosal surfaces. The aim of this review is to summarize studies on genetic polymorphisms, structural variants, and serum levels of human SP-A and SP-D and their associations with human pulmonary disease.
Immunobiology 02/2007; 212(4-5):381-416. · 3.20 Impact Factor
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ABSTRACT: The severe acute respiratory syndrome coronavirus (SARS-CoV) infects host cells with its surface glycosylated spike-protein (S-protein). Here we expressed the SARS-CoV S-protein to investigate its interactions with innate immune mechanisms in the lung. The purified S-protein was detected as a 210 kDa glycosylated protein. It was not secreted in the presence of tunicamycin and was detected as a 130 kDa protein in the cell lysate. The purified S-protein bound to Vero but not 293T cells and was itself recognized by lung surfactant protein D (SP-D), a collectin found in the lung alveoli. The binding required Ca(2+) and was inhibited by maltose. The serum collectin, mannan-binding lectin (MBL), exhibited no detectable binding to the purified S-protein. S-protein binds and activates macrophages but not dendritic cells (DCs). It suggests that SARS-CoV interacts with innate immune mechanisms in the lung through its S-protein and regulates pulmonary inflammation.
Immunobiology 02/2007; 212(3):201-11. · 3.20 Impact Factor
