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Peter Garred - One of the best experts on this subject based on the ideXlab platform.
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associations between serum l arginine and Ficolins in the early phase of acute ischemic stroke a pilot study
Journal of Stroke & Cerebrovascular Diseases, 2020Co-Authors: Tihamer Molnar, Peter Garred, Dorottya Csuka, Gabriella Pusch, Lajos Nagy, Zsolt IllesAbstract:Abstract Introduction Activation of both the L-arginine and the lectin pathway contributes to the pathophysiology and the outcome of acute ischemic stroke (AIS). However, the interplay between the two systems has not yet been examined. Methods A total of 44 patients with AIS were recruited into this study. Serial measurement of serum L-arginine, asymmetric and symmetric dimethylarginine (ADMA, SDMA), and hsCRP, ficolin-2, ficolin-3, MAP-1, MASP-3 and mannose-binding lectin (MBL) were analyzed within 6 h after onset of stroke and 72 h later. Outcomes were assessed as National Institutes of Health Stroke Scale (NIHSS) worsening by 24 h, poststroke infection, and death by 1 month. Results In the hyperacute stage of AIS, ficolin-3, MAP-1 and MBL were positively correlated with L-arginine within 6 h after onset of symptoms (p Conclusion Early consumption of ficolin-3 is associated with complications such as post-stroke infections. In the hyperacute phase of AIS, the positive correlation between Ficolins and the NO donor L-arginine may reflect the protective role of L-arginine presumably by improving the cerebral microcirculation in a prothrombotic environment induced by complement activation.
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Circulating Ficolin-2 and Ficolin-3 Form Heterocomplexes.
Journal of immunology (Baltimore Md. : 1950), 2020Co-Authors: Ida Jarlhelt, Mikkel-ole Skjoedt, Katrine Pilely, Jytte Bryde Clausen, Rafael Bayarri-olmos, Peter GarredAbstract:The complement system constitutes an important part of the innate immune system. The collectins and the Ficolins are soluble pattern recognition molecules that contribute to complement activation via the lectin pathway. During previous experiments with ficolin-2 and ficolin-3, we have observed that the molecules may interact. We therefore hypothesized the existence of stable ficolin-2/-3 heterocomplexes. We could demonstrate ficolin-2/-3 heterocomplexes in normal human serum and plasma by ELISA using Abs specific for ficolin-2 and ficolin-3. The formation of heteromeric protein complexes were validated by coimmunoprecipitation and Western blot analysis. When recombinant ficolin-2 and recombinant ficolin-3 were mixed, no complexes were formed. However, when coexpressing ficolin-2 and ficolin-3 in Chinese hamster ovary cells, we could detect ficolin-2/-3 heterocomplexes in the supernatant. Furthermore, we measured concentration of the ficolin-2/-3 heterocomplexes in arbitrary units in 94 healthy individuals. We also established the relationship between the concentrations of ficolin-2, ficolin-3, and the ficolin-2/-3 heterocomplexes. We observed that the concentration of the ficolin-2/-3 heterocomplex correlated significantly with ficolin-2 (ρ: 0.24, p < 0.018) and ficolin-3 concentrations (ρ: 0.46, p < 0.0001). In conclusion, we describe a novel protein complex between ficolin-2 and ficolin-3 present in serum and plasma, which might be of additional biological relevance apart from the native ficolin-2 and ficolin-3 molecules.
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The ficolin response to LPS challenge in mice.
Molecular immunology, 2019Co-Authors: Ida Jarlhelt, Ninette Genster, Nikolaj Kirketerp-møller, Mikkel-ole Skjoedt, Peter GarredAbstract:The Ficolins belong to an important family of pattern recognition molecules, which contributes to complement activation via the lectin pathway. How the Ficolins respond to inflammatory stimuli remains only partly understood. In the present study, we investigated the ficolin A and ficolin B expression and protein distribution patterns in a mouse model of LPS-induced inflammation. The time- and tissue-specific expression of ficolin A and B was determined by real time PCR. Furthermore, ficolin protein levels in serum and bone marrow extracts from LPS challenged mice were determined by novel in-house developed sandwich ELISAs. Ficolin A was mainly expressed in liver and spleen. However, our data also suggested that ficolin A is expressed in bone marrow, which is the main site of ficolin B expression. The level of ficolin A and B expression was increased after stimulation with LPS in the investigated tissues. This was followed by a downregulation of expression, causing mRNA levels to return to baseline 24 h post LPS challenge. Protein levels appeared to follow the same pattern as the expression profiles, with an exception of ficolin B levels in serum, which kept increasing for 24 h. Ficolin A was likewise significantly increased in bronchoalveolar lavage fluid from mice infected with the fungi A. fumigatus, pointing towards a similar effect of the Ficolins in non-sterile mouse models of inflammation. The results demonstrate that LPS-induced inflammation can induce a significant ficolin response, suggesting that the murine Ficolins are acute phase reactants with increase in both mRNA expression and protein levels during systemic inflammation.
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Ficolins do not alter host immune responses to lipopolysaccharide-induced inflammation in vivo.
Scientific reports, 2017Co-Authors: Ninette Genster, Jack Cowland, Olga Østrup, Camilla Schjalm, Tom Eirik Mollnes, Peter GarredAbstract:Ficolins are a family of pattern recognition molecules that are capable of activating the lectin pathway of complement. A limited number of reports have demonstrated a protective role of Ficolins in animal models of infection. In addition, an immune modulatory role of Ficolins has been suggested. Yet, the contribution of Ficolins to inflammatory disease processes remains elusive. To address this, we investigated ficolin deficient mice during a lipopolysaccharide (LPS)-induced model of systemic inflammation. Although murine serum ficolin was shown to bind LPS in vitro, there was no difference between wildtype and ficolin deficient mice in morbidity and mortality by LPS-induced inflammation. Moreover, there was no difference between wildtype and ficolin deficient mice in the inflammatory cytokine profiles after LPS challenge. These findings were substantiated by microarray analysis revealing an unaltered spleen transcriptome profile in ficolin deficient mice compared to wildtype mice. Collectively, results from this study demonstrate that Ficolins are not involved in host response to LPS-induced systemic inflammation.
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The Interaction Pattern of Murine Serum Ficolin-A with
2016Co-Authors: Tina Hummelshøj, Lea Munthe-fog, Yuichi Endo, Teizo Fujita, Ying Jie, Thomas Bjarnsholt, Claus Moser, Ole Skjoedt, Luigina Romani, Peter GarredAbstract:The Ficolins are soluble pattern recognition molecules in the lectin pathway of complement, but the spectrum and mode of interaction with pathogens are largely unknown. In this study, we investigated the binding properties of the murine serum ficolin-A towards a panel of different clinical relevant microorganisms (N = 45) and compared the binding profile with human serum ficolin-2 and ficolin-3. Ficolin-A was able to bind Gram-positive bacteria strains including E. faecalis, L. monocytogenes and some S. aureus strains, but not to the investigated S. agalactiae (Group B streptococcus) strains. Regarding Gram-negative bacteria ficolin-A was able to bind to some E. coli and P. aeruginosa strains, but not to the investigated Salmonella strains. Of particular interest ficolin-A bound strongly to the pathogenic E. coli, O157:H7 and O149 strains, but it did not bind to the non-pathogenic E. coli, ATCC 25922 strain. Additionally, ficolin-A was able to bind purified LPS from these pathogenic strains. Furthermore, ficolin-A bound to a clinical isolate of the fungus A. fumigatus. In general ficolin-2 showed similar selective binding spectrum towards pathogenic microorganisms as observed for ficolin-A indicating specific pathophysiological roles of these molecules in host defence. In contrast, ficolin-3 did not bind to any of th
Teizo Fujita - One of the best experts on this subject based on the ideXlab platform.
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The Interaction Pattern of Murine Serum Ficolin-A with
2016Co-Authors: Tina Hummelshøj, Lea Munthe-fog, Yuichi Endo, Teizo Fujita, Ying Jie, Thomas Bjarnsholt, Claus Moser, Ole Skjoedt, Luigina Romani, Peter GarredAbstract:The Ficolins are soluble pattern recognition molecules in the lectin pathway of complement, but the spectrum and mode of interaction with pathogens are largely unknown. In this study, we investigated the binding properties of the murine serum ficolin-A towards a panel of different clinical relevant microorganisms (N = 45) and compared the binding profile with human serum ficolin-2 and ficolin-3. Ficolin-A was able to bind Gram-positive bacteria strains including E. faecalis, L. monocytogenes and some S. aureus strains, but not to the investigated S. agalactiae (Group B streptococcus) strains. Regarding Gram-negative bacteria ficolin-A was able to bind to some E. coli and P. aeruginosa strains, but not to the investigated Salmonella strains. Of particular interest ficolin-A bound strongly to the pathogenic E. coli, O157:H7 and O149 strains, but it did not bind to the non-pathogenic E. coli, ATCC 25922 strain. Additionally, ficolin-A was able to bind purified LPS from these pathogenic strains. Furthermore, ficolin-A bound to a clinical isolate of the fungus A. fumigatus. In general ficolin-2 showed similar selective binding spectrum towards pathogenic microorganisms as observed for ficolin-A indicating specific pathophysiological roles of these molecules in host defence. In contrast, ficolin-3 did not bind to any of th
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New insights into the role of Ficolins in the lectin pathway of innate immunity.
International review of cell and molecular biology, 2015Co-Authors: Yuichi Endo, Misao Matsushita, Teizo FujitaAbstract:In the innate immune system, a variety of recognition molecules provide the first-line host defense to prevent infection and maintain endogenous homeostasis. Ficolin is a soluble recognition molecule, which senses pathogen-associated molecular patterns on microbes and aberrant sugar structures on self-cells. It consists of a collagen-like stalk and a globular fibrinogen-like domain, the latter binding to carbohydrates such as N-acetylglucosamine. Ficolins have been widely identified in animals from higher invertebrates to mammals. In mammals, Ficolins form complexes with mannose-binding lectin-associated serine proteases (MASPs), and ficolin-MASP complexes trigger complement activation via the lectin pathway. Once activated, complement mediates many immune responses including opsonization, phagocytosis, and cytokine production. Although the precise function of each ficolin is still under investigation, accumulating information suggests that Ficolins have a crucial role in host defense by recognizing a variety of microorganisms and interacting with effector proteins.
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Trypanosoma cruzi calreticulin inhibits the complement lectin pathway activation by direct interaction with L-Ficolin.
Molecular Immunology, 2014Co-Authors: Eduardo Sosoniuk, Teizo Fujita, Christine Gaboriaud, Gerardo Vallejos, Hany Kenawy, Nicole Thielens, Wilhelm Schwaeble, Arturo Ferreira, Carolina ValckAbstract:Trypanosoma cruzi, the agent of Chagas' disease, the sixth neglected tropical disease worldwide, infects 10-12 million people in Latin America. Differently from T. cruzi epimastigotes, trypomastigotes are complement-resistant and infective. CRPs, T-DAF, sialic acid and lipases explain at least part of this resistance. In vitro, T. cruzi calreticulin (TcCRT), a chaperone molecule that translocates from the ER to the parasite surface: (a) Inhibits the human classical complement activation, by interacting with C1, (b) As a consequence, an increase in infectivity is evident and, (c) It inhibits angiogenesis and tumor growth. We report here that TcCRT also binds to the L-Ficolin collagenous portion, thus inhibiting approximately between 35 and 64% of the human complement lectin pathway activation, initiated by L-Ficolin, a property not shared by H-Ficolin. While L-Ficolin binds to 60% of trypomastigotes and to 24% of epimastigotes, 50% of the former and 4% of the latter display TcCRT on their surfaces. Altogether, these data indicate that TcCRT is a parasite inhibitory receptor for Ficolins. The resulting evasive activities, together with the TcCRT capacity to inhibit C1, with a concomitant increase in infectivity, may represent T. cruzi strategies to inhibit important arms of the innate immune response.
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Purification, measurement of concentration, and functional complement assay of human Ficolins.
Methods in molecular biology (Clifton N.J.), 2013Co-Authors: Misao Matsushita, Yuichi Endo, Yu Liu, David C. Kilpatrick, Hiroshi Shiraki, Koichiro Tateishi, Mitsushi Tsujimura, Teizo FujitaAbstract:Ficolins constitute a group of lectins involved in innate immunity. L-Ficolin, H-ficolin, and M-ficolin are present in human serum. The human Ficolins differ in carbohydrate-binding specificity, but they have in common the ability to recognize the acetyl group. L-Ficolin and H-ficolin are associated with serine proteases termed MASPs (MBL-associated serine proteases) and their truncated proteins, and the complexes (L/H-ficolin-MASP) activate the lectin pathway of complement upon binding to their ligands. Recombinant M-ficolin is also able to form a complex with MASP, resulting in complement activation. L-Ficolin and H-ficolin can be purified as a complex with MASP from serum by utilizing their binding specificities. These ficolin-MASP complexes have an ability to activate C4. Human Ficolins are quantified by ELISA using specific antibodies or ligands.
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Structural and Functional Overview of the Lectin Complement Pathway: Its Molecular Basis and Physiological Implication
Archivum Immunologiae et Therapiae Experimentalis, 2013Co-Authors: Misao Matsushita, Yuichi Endo, Teizo FujitaAbstract:The complement system is an effector mechanism in immunity. It is activated in three ways, the classical, alternative and lectin pathways. The lectin pathway is initiated by the binding of mannose-binding lectin (MBL) or Ficolins to carbohydrates on the surfaces of pathogens. In humans, MBL and three types of Ficolins (L-ficolin, H-ficolin, and M-ficolin) are present in plasma. Of these lectins, at least, MBL, L-ficolin, and H-ficolin are complexed with three types of MBL-associated serine proteases (MASPs), MASP-1, MASP-2, and MASP-3 and their truncated proteins (MAp44 and sMAP). In the lectin pathway, the lectin–MASP complex (i.e., a complex of lectin, MASPs and their truncated proteins) binds to pathogens, resulting in the activation of C4 and C2 to generate a C3 convertase capable of activating C3. MASP-2 is involved in the activation of C4 and C2. MASP-1 activates C2 and MASP-2. The functions of MASP-3, sMAP, and MAp44 in the lectin pathway remain unknown. MASP-1 and MASP-3 also have a role in the alternative pathway. MBL and Ficolins are able to bind to a variety of pathogens depending on their carbohydrate binding specificity, resulting in the activation of the lectin pathway. Deficiencies of the components of the lectin pathway are associated to susceptibility to infection, indicating an important role of the lectin pathway in innate immunity. The lectin-MASP complex is also involved in innate immunity by activating the coagulation system. Recent findings suggest a crucial role of MASP-3 in development.
Christine Gaboriaud - One of the best experts on this subject based on the ideXlab platform.
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Human L-Ficolin Recognizes Phosphocholine Moieties of Pneumococcal Teichoic Acid
Journal of Immunology, 2014Co-Authors: Emilie Vassal-stermann, Evelyne Gout, Monique Lacroix, Lydie Martin, Emmanuelle Laffly, C. M. Pedersen, A. Amoroso, R. R. Schmidt, U. Zahringer, Christine GaboriaudAbstract:Human L-ficolin is a soluble protein of the innate immune system able to sense pathogens through its fibrinogen (FBG) recognition domains and to trigger activation of the lectin complement pathway through associated serine proteases. L-Ficolin has been previously shown to recognize pneumococcal clinical isolates, but its ligands and especially its molecular specificity remain to be identified. Using solid-phase binding assays, serum and recombinant L-Ficolins were shown to interact with serotype 2 pneumococcal strain D39 and its unencapsulated R6 derivative. Incubation of both strains with serum triggered complement activation, as measured by C4b and C3b deposition, which was decreased by using ficolin-depleted serum. Recombinant L-ficolin and its FBG-like recognition domain bound to isolated pneumococcal cell wall extracts, whereas binding to cell walls depleted of teichoic acid (TA) was decreased. Both proteins were also shown to interact with two synthetic TA compounds, each comprising part structures of the complete lipoteichoic acid molecule with two PCho residues. Competition studies and direct interaction measurements by surface plasmon resonance identified PCho as a novel L-ficolin ligand. Structural analysis of complexes of the FBG domain of L-ficolin and PCho revealed that the phosphate moiety interacts with amino acids previously shown to define an acetyl binding site. Consequently, binding of L-ficolin to immobilized acetylated BSA was inhibited by PCho and synthetic TA. Binding of serum L-ficolin to immobilized synthetic TA and PCho-conjugated BSA triggered activation of the lectin complement pathway, thus further supporting the hypothesis of L-ficolin involvement in host antipneumococcal defense.
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Trypanosoma cruzi calreticulin inhibits the complement lectin pathway activation by direct interaction with L-Ficolin.
Molecular Immunology, 2014Co-Authors: Eduardo Sosoniuk, Teizo Fujita, Christine Gaboriaud, Gerardo Vallejos, Hany Kenawy, Nicole Thielens, Wilhelm Schwaeble, Arturo Ferreira, Carolina ValckAbstract:Trypanosoma cruzi, the agent of Chagas' disease, the sixth neglected tropical disease worldwide, infects 10-12 million people in Latin America. Differently from T. cruzi epimastigotes, trypomastigotes are complement-resistant and infective. CRPs, T-DAF, sialic acid and lipases explain at least part of this resistance. In vitro, T. cruzi calreticulin (TcCRT), a chaperone molecule that translocates from the ER to the parasite surface: (a) Inhibits the human classical complement activation, by interacting with C1, (b) As a consequence, an increase in infectivity is evident and, (c) It inhibits angiogenesis and tumor growth. We report here that TcCRT also binds to the L-Ficolin collagenous portion, thus inhibiting approximately between 35 and 64% of the human complement lectin pathway activation, initiated by L-Ficolin, a property not shared by H-Ficolin. While L-Ficolin binds to 60% of trypomastigotes and to 24% of epimastigotes, 50% of the former and 4% of the latter display TcCRT on their surfaces. Altogether, these data indicate that TcCRT is a parasite inhibitory receptor for Ficolins. The resulting evasive activities, together with the TcCRT capacity to inhibit C1, with a concomitant increase in infectivity, may represent T. cruzi strategies to inhibit important arms of the innate immune response.
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Deciphering complement receptor type 1 interactions with recognition proteins of the lectin complement pathway.
Journal of Immunology, 2013Co-Authors: Mickaël Jacquet, Evelyne Gout, Monique Lacroix, Nicole Thielens, Christine Gaboriaud, Sarah Ancelet, Véronique RossiAbstract:Complement receptor type 1 (CR1) is a membrane receptor expressed on a wide range of cells. It is involved in immune complex clearance, phagocytosis, and complement regulation. Its ectodomain is composed of 30 complement control protein (CCP) modules, organized into four long homologous repeats (A-D). In addition to its main ligands C3b and C4b, CR1 was reported to interact with C1q and mannan-binding lectin (MBL) likely through its C-terminal region (CCP22-30). To decipher the interaction of human CR1 with the recognition proteins of the lectin complement pathway, a recombinant fragment encompassing CCP22-30 was expressed in eukaryotic cells, and its interaction with human MBL and Ficolins was investigated using surface plasmon resonance spectroscopy. MBL and L-ficolin were shown to interact with immobilized soluble CR1 and CR1 CCP22-30 with apparent dissociation constants in the nanomolar range, indicative of high affinity. The binding site for CR1 was located at or near the MBL-associated serine protease (MASP) binding site in the collagen stalks of MBL and L-ficolin, as shown by competition experiments with MASP-3. Accordingly, the mutation of an MBL conserved lysine residue essential for MASP binding (K55) abolished binding to soluble CR1 and CCP22-30. The CR1 binding site for MBL/Ficolins was mapped to CCP24-25 of long homologous repeat D using deletion mutants. In conclusion, we show that Ficolins are new CR1 ligands and propose that MBL/L-ficolin binding involves major ionic interactions between conserved lysine residues of their collagen stalks and surface exposed acidic residues located in CR1 CCP24 and/or CCP25.
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carbohydrate recognition properties of human Ficolins glycan array screening reveals the sialic acid binding specificity of m ficolin
Journal of Biological Chemistry, 2010Co-Authors: Evelyne Gout, Monique Lacroix, Gérard J Arlaud, Lydie Martin, Jean-yves Cesbron, V Garlatti, David F Smith, Chantal Dumestreperard, Thomas Lunardi, Christine GaboriaudAbstract:Ficolins are oligomeric innate immune recognition proteins consisting of a collagen-like region and a fibrinogen-like recognition domain that bind to pathogen- and apoptotic cell-associated molecular patterns. To investigate their carbohydrate binding specificities, serum-derived L-ficolin and recombinant H- and M-Ficolins were fluorescently labeled, and their carbohydrate binding ability was analyzed by glycan array screening. L-ficolin preferentially recognized disulfated N-acetyllactosamine and tri- and tetrasaccharides containing terminal galactose or N-acetylglucosamine. Binding was sensitive to the position and orientation of the bond between N-acetyllactosamine and the adjacent carbohydrate. No significant binding of H-ficolin to any of the 377 glycans probed could be detected, providing further evidence for its poor lectin activity. M-ficolin bound preferentially to 9-O-acetylated 2-6-linked sialic acid derivatives and to various glycans containing sialic acid engaged in a 2-3 linkage. To further investigate the structural basis of sialic acid recognition by M-ficolin, point mutants were produced in which three residues of the fibrinogen domain were replaced by their counterparts in L-ficolin. Mutations G221F and A256V inhibited binding to the 9-O-acetylated sialic acid derivatives, whereas Y271F abolished interaction with all sialic acid-containing glycans. The crystal structure of the Y271F mutant fibrinogen domain was solved, showing that the mutation does not alter the structure of the ligand binding pocket. These analyses reveal novel ficolin ligands such as sulfated N-acetyllactosamine (L-ficolin) and gangliosides (M-ficolin) and provide precise insights into the sialic acid binding specificity of M-ficolin, emphasizing the essential role of Tyr(271) in this respect.
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Structural insights into the recognition properties of human Ficolins.
Journal of innate immunity, 2009Co-Authors: V Garlatti, Evelyne Gout, Monique Lacroix, Gérard J Arlaud, Nicole Thielens, Lydie Martin, Christine GaboriaudAbstract:Innate immunity relies upon the ability of a variety of recognition molecules to sense pathogens through conserved molecular signatures that are often carbohydrates. Ficolins are oligomeric proteins assembled from collagen-like stalks and fibrinogen-like domains that have the ability to sense these molecular patterns on both pathogens and apoptotic cell surfaces. Three Ficolins, termed L, H and M, have been identified in humans. They differ in their localization and concentration in extracellular fluids, their mode of secretion and their recognition properties. From a structural point of view, Ficolins are assembled from basal trimeric subunits comprising a collagen-like triple helix and a globular domain composed of 3 fibrinogen-like domains. The globular domains are responsible for sensing danger signals whereas the collagen-like stalks provide a link with immune effectors. This review mainly focuses on the structure and recognition properties of the 3 human Ficolins, as revealed by recent crystallographic analysis of their recognition domains. The ligand binding sites have been identified in the 3 Ficolins and their recognition mechanisms have been characterized at the atomic level. In the case of M-ficolin, a structural transition at acidic pH disables the binding pocket, and thus likely participates in the functional cycle of this protein.
Misao Matsushita - One of the best experts on this subject based on the ideXlab platform.
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chapter 5 Ficolins
The Complement FactsBook (Second Edition), 2018Co-Authors: Misao MatsushitaAbstract:The human Ficolins are homooligomers. Each subunit is composed of an N-terminal region including two cysteine residues, a collagen-like domain, a neck region and a fibrinogen-like domain. Electron microscopy of the whole proteins of human ficolin-2 and ficolin-3 show bouquet-like images, similar to those first described for C1q. All human Ficolins recognise N-acetylated carbohydrates and compounds such as N-acetylglucosamine (GlcNAc) in common, although the spectrum of ligands for each ficolin is slightly different. Human ficolin-1 binds sialic acid, and human ficolin-3 recognises d-fucose and galactose. Ficolin-2 exhibits broad binding specificities due to it having four distinct binding sites in the fibrinogen-like domain that allow it to bind various carbohydrates, including β-(1,3)-d-glucan and peptideglycan, and sulphated carbohydrates, including heparin, while ficolin-1 and ficolin-3 have only one binding site. In addition to carbohydrates, ficolin-1 binds to tectonin9, and ficolin-2 binds elastin, corticosteroid11 and DNA. Each ficolin is able to bind to a spectrum of bacteria. Ficolin-1 and ficolin-2 interact with C-reactive protein, boosting both the lectin and classical pathway. A low concentration of ficolin-1 is also present in the serum with the mean level of 1 μg/mL. Ficolin-2 and ficolin-3 are expressed in the liver and are secreted into the serum. Two cases of ficolin-3 deficiency due to homozygosity for a FCN3 frameshift mutation (c.349delC, rs28357092) lacking the entire fibrinogen-like in exon 5 leading to the abnormal ficolin-3 lacking the entire fibrinogen-like domain have been reported.
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Chapter 5 – Ficolins
The Complement FactsBook, 2018Co-Authors: Misao MatsushitaAbstract:The human Ficolins are homooligomers. Each subunit is composed of an N-terminal region including two cysteine residues, a collagen-like domain, a neck region and a fibrinogen-like domain. Electron microscopy of the whole proteins of human ficolin-2 and ficolin-3 show bouquet-like images, similar to those first described for C1q. All human Ficolins recognise N-acetylated carbohydrates and compounds such as N-acetylglucosamine (GlcNAc) in common, although the spectrum of ligands for each ficolin is slightly different. Human ficolin-1 binds sialic acid, and human ficolin-3 recognises d-fucose and galactose. Ficolin-2 exhibits broad binding specificities due to it having four distinct binding sites in the fibrinogen-like domain that allow it to bind various carbohydrates, including β-(1,3)-d-glucan and peptideglycan, and sulphated carbohydrates, including heparin, while ficolin-1 and ficolin-3 have only one binding site. In addition to carbohydrates, ficolin-1 binds to tectonin9, and ficolin-2 binds elastin, corticosteroid11 and DNA. Each ficolin is able to bind to a spectrum of bacteria. Ficolin-1 and ficolin-2 interact with C-reactive protein, boosting both the lectin and classical pathway. A low concentration of ficolin-1 is also present in the serum with the mean level of 1 μg/mL. Ficolin-2 and ficolin-3 are expressed in the liver and are secreted into the serum. Two cases of ficolin-3 deficiency due to homozygosity for a FCN3 frameshift mutation (c.349delC, rs28357092) lacking the entire fibrinogen-like in exon 5 leading to the abnormal ficolin-3 lacking the entire fibrinogen-like domain have been reported.
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New insights into the role of Ficolins in the lectin pathway of innate immunity.
International review of cell and molecular biology, 2015Co-Authors: Yuichi Endo, Misao Matsushita, Teizo FujitaAbstract:In the innate immune system, a variety of recognition molecules provide the first-line host defense to prevent infection and maintain endogenous homeostasis. Ficolin is a soluble recognition molecule, which senses pathogen-associated molecular patterns on microbes and aberrant sugar structures on self-cells. It consists of a collagen-like stalk and a globular fibrinogen-like domain, the latter binding to carbohydrates such as N-acetylglucosamine. Ficolins have been widely identified in animals from higher invertebrates to mammals. In mammals, Ficolins form complexes with mannose-binding lectin-associated serine proteases (MASPs), and ficolin-MASP complexes trigger complement activation via the lectin pathway. Once activated, complement mediates many immune responses including opsonization, phagocytosis, and cytokine production. Although the precise function of each ficolin is still under investigation, accumulating information suggests that Ficolins have a crucial role in host defense by recognizing a variety of microorganisms and interacting with effector proteins.
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Serum ficolin-2 in hospitalised patients with community-acquired pneumonia.
Inflammation, 2014Co-Authors: James D. Chalmers, Misao Matsushita, David C. Kilpatrick, Gilly B. Fleming, Julia Rutherford, Adam T. HillAbstract:Mannose binding lectin (MBL) and Ficolins contribute to host defence through activation of the lectin pathway of complement. In this study, serum levels of ficolin-2 and MBL were determined in 276 patients with community-acquired pneumonia (CAP). MBL deficiency and ficolin-2 insufficiency were defined using previously validated cut-offs. No differences were observed in MBL or ficolin-2 between patients and controls. MBL-deficient patients (
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Purification, measurement of concentration, and functional complement assay of human Ficolins.
Methods in molecular biology (Clifton N.J.), 2013Co-Authors: Misao Matsushita, Yuichi Endo, Yu Liu, David C. Kilpatrick, Hiroshi Shiraki, Koichiro Tateishi, Mitsushi Tsujimura, Teizo FujitaAbstract:Ficolins constitute a group of lectins involved in innate immunity. L-Ficolin, H-ficolin, and M-ficolin are present in human serum. The human Ficolins differ in carbohydrate-binding specificity, but they have in common the ability to recognize the acetyl group. L-Ficolin and H-ficolin are associated with serine proteases termed MASPs (MBL-associated serine proteases) and their truncated proteins, and the complexes (L/H-ficolin-MASP) activate the lectin pathway of complement upon binding to their ligands. Recombinant M-ficolin is also able to form a complex with MASP, resulting in complement activation. L-Ficolin and H-ficolin can be purified as a complex with MASP from serum by utilizing their binding specificities. These ficolin-MASP complexes have an ability to activate C4. Human Ficolins are quantified by ELISA using specific antibodies or ligands.
Mikkel-ole Skjoedt - One of the best experts on this subject based on the ideXlab platform.
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Circulating Ficolin-2 and Ficolin-3 Form Heterocomplexes.
Journal of immunology (Baltimore Md. : 1950), 2020Co-Authors: Ida Jarlhelt, Mikkel-ole Skjoedt, Katrine Pilely, Jytte Bryde Clausen, Rafael Bayarri-olmos, Peter GarredAbstract:The complement system constitutes an important part of the innate immune system. The collectins and the Ficolins are soluble pattern recognition molecules that contribute to complement activation via the lectin pathway. During previous experiments with ficolin-2 and ficolin-3, we have observed that the molecules may interact. We therefore hypothesized the existence of stable ficolin-2/-3 heterocomplexes. We could demonstrate ficolin-2/-3 heterocomplexes in normal human serum and plasma by ELISA using Abs specific for ficolin-2 and ficolin-3. The formation of heteromeric protein complexes were validated by coimmunoprecipitation and Western blot analysis. When recombinant ficolin-2 and recombinant ficolin-3 were mixed, no complexes were formed. However, when coexpressing ficolin-2 and ficolin-3 in Chinese hamster ovary cells, we could detect ficolin-2/-3 heterocomplexes in the supernatant. Furthermore, we measured concentration of the ficolin-2/-3 heterocomplexes in arbitrary units in 94 healthy individuals. We also established the relationship between the concentrations of ficolin-2, ficolin-3, and the ficolin-2/-3 heterocomplexes. We observed that the concentration of the ficolin-2/-3 heterocomplex correlated significantly with ficolin-2 (ρ: 0.24, p < 0.018) and ficolin-3 concentrations (ρ: 0.46, p < 0.0001). In conclusion, we describe a novel protein complex between ficolin-2 and ficolin-3 present in serum and plasma, which might be of additional biological relevance apart from the native ficolin-2 and ficolin-3 molecules.
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The ficolin response to LPS challenge in mice.
Molecular immunology, 2019Co-Authors: Ida Jarlhelt, Ninette Genster, Nikolaj Kirketerp-møller, Mikkel-ole Skjoedt, Peter GarredAbstract:The Ficolins belong to an important family of pattern recognition molecules, which contributes to complement activation via the lectin pathway. How the Ficolins respond to inflammatory stimuli remains only partly understood. In the present study, we investigated the ficolin A and ficolin B expression and protein distribution patterns in a mouse model of LPS-induced inflammation. The time- and tissue-specific expression of ficolin A and B was determined by real time PCR. Furthermore, ficolin protein levels in serum and bone marrow extracts from LPS challenged mice were determined by novel in-house developed sandwich ELISAs. Ficolin A was mainly expressed in liver and spleen. However, our data also suggested that ficolin A is expressed in bone marrow, which is the main site of ficolin B expression. The level of ficolin A and B expression was increased after stimulation with LPS in the investigated tissues. This was followed by a downregulation of expression, causing mRNA levels to return to baseline 24 h post LPS challenge. Protein levels appeared to follow the same pattern as the expression profiles, with an exception of ficolin B levels in serum, which kept increasing for 24 h. Ficolin A was likewise significantly increased in bronchoalveolar lavage fluid from mice infected with the fungi A. fumigatus, pointing towards a similar effect of the Ficolins in non-sterile mouse models of inflammation. The results demonstrate that LPS-induced inflammation can induce a significant ficolin response, suggesting that the murine Ficolins are acute phase reactants with increase in both mRNA expression and protein levels during systemic inflammation.
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Ficolins and the lectin pathway of complement in patients with systemic lupus erythematosus
Molecular Immunology, 2015Co-Authors: Estrid Hein, Mikkel-ole Skjoedt, Louise Nielsen, Christoffer Tandrup Nielsen, Søren Jacobsen, Lea Munthefog, Peter GarredAbstract:The complement system plays a pathophysiological role in systemic lupus erythematosus (SLE). This study aims to investigate whether an association exists between the Ficolins that are part of the lectin complement pathway and SLE. EDTA plasma samples from 68 Danish SLE patients and 29 healthy donors were included in the study. Plasma concentrations of Ficolin-1, -2, and -3 were determined in specific sandwich ELISAs. Lectin pathway activity via Ficolin-3 was measured in ELISA on acetylated bovine serum albumin (acBSA) and measured as Ficolin-3 binding and deposition of C4, C3 and the terminal complement complex (TCC). SLE patients had increased levels of Ficolin-3, 21.6μg/ml as compared to 17.0μg/ml in healthy controls (P=0.0098). The Ficolin-1 plasma concentration was negatively correlated with SLE Disease Activity Index (SLEDAI) (Rho=-0.29, P=0.015) and positively correlated to the [Systemic Lupus International Collaborating Clinics (SLICC)/American College of Rheumatology (ACR) Damage Index] (SDI) (Rho=0.27, P=0.026). The Ficolin-1 concentration was also associated with the occurrence of arterial (P=0.0053) but not venous thrombosis (P=0.42). Finally, deposition of C4, C3 and TCC in the Ficolin-3 pathway were all correlated to SLEDAI, respectively (P<0.0076). The Ficolin-1 association to SLEDAI and SDI as well as arterial thrombosis shown in this study suggests that Ficolin-1 may be a potential new biomarker for patients with SLE. Furthermore, Ficolin-3 mediated complement activation may be valuable in monitoring disease activity in SLE patients due to the high sensitivity for complement consumption in the assay independent of the Ficolin-3 concentration.
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Ficolins and the lectin pathway of complement in patients with systemic lupus erythematosus.
Molecular immunology, 2014Co-Authors: Estrid Hein, Lea Munthe-fog, Mikkel-ole Skjoedt, Louise Nielsen, Christoffer Tandrup Nielsen, Søren Jacobsen, Peter GarredAbstract:The complement system plays a pathophysiological role in systemic lupus erythematosus (SLE). This study aims to investigate whether an association exists between the Ficolins that are part of the lectin complement pathway and SLE. EDTA plasma samples from 68 Danish SLE patients and 29 healthy donors were included in the study. Plasma concentrations of Ficolin-1, -2, and -3 were determined in specific sandwich ELISAs. Lectin pathway activity via Ficolin-3 was measured in ELISA on acetylated bovine serum albumin (acBSA) and measured as Ficolin-3 binding and deposition of C4, C3 and the terminal complement complex (TCC). SLE patients had increased levels of Ficolin-3, 21.6μg/ml as compared to 17.0μg/ml in healthy controls (P=0.0098). The Ficolin-1 plasma concentration was negatively correlated with SLE Disease Activity Index (SLEDAI) (Rho=-0.29, P=0.015) and positively correlated to the [Systemic Lupus International Collaborating Clinics (SLICC)/American College of Rheumatology (ACR) Damage Index] (SDI) (Rho=0.27, P=0.026). The Ficolin-1 concentration was also associated with the occurrence of arterial (P=0.0053) but not venous thrombosis (P=0.42). Finally, deposition of C4, C3 and TCC in the Ficolin-3 pathway were all correlated to SLEDAI, respectively (P
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collectin 11 masp complex formation triggers activation of the lectin complement pathway the fifth lectin pathway initiation complex
Journal of Innate Immunity, 2013Co-Authors: Ying Jie, Mikkel-ole Skjoedt, Peter GarredAbstract:Collectins and Ficolins are important in the clearance of endogenous and exogenous danger materials. A new human collectin-11 was recently identified in low concentration in serum in complex with mannose-binding lectin (MBL)/ficolin-associated serine proteases. Collectin-11 binds to carbohydrate residues present on various microorganisms. Thus, we hypothesized that collectin-11 could be a novel initiation molecule in the lectin pathway of complement. We can show that collectin-11 associates with all the known MBL-associated serine proteases (MASP-1, MASP-2 and MASP-3) as well as the lectin complement pathway regulator MAP-1. Furthermore, we found that complex formation between recombinant collectin-11 and recombinant MASP-2 on Candida albicans leads to deposition of C4b. Native collectin-11 in serum mediated complement activation and deposition of C4b and C3b, and formation of the terminal complement complex on C. albicans. Moreover, spiking collectin-11-depleted serum, which did not mediate complement activation, with recombinant collectin-11 restored the complement activation capability. These results define collectin-11 as the fifth recognition molecule in the lectin complement pathway in addition to MBL, ficolin-1, ficolin-2 and ficolin-3.