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Sample records for complement lectin pathway

  1. Mitochondria and the lectin pathway of complement.

    PubMed

    Brinkmann, Christel R; Jensen, Lisbeth; Dagns-Hansen, Frederik; Holm, Ida E; Endo, Yuichi; Fujita, Teizo; Thiel, Steffen; Jensenius, Jens C; Degn, Sren E

    2013-03-22

    Mitochondria, the powerhouses of our cells, are remnants of a eubacterial endosymbiont. Notwithstanding the evolutionary time that has passed since the initial endosymbiotic event, mitochondria have retained many hallmarks of their eubacterial origin. Recent studies have indicated that during perturbations of normal homeostasis, such as following acute trauma leading to massive necrosis and release of mitochondria, the immune system might mistake symbiont for enemy and initiate an inappropriate immune response. The innate immune system is the first line of defense against invading microbial pathogens, and as such is the primary suspect in the recognition of mitochondria-derived danger-associated molecular patterns and initiation of an aberrant response. Conversely, innate immune mechanisms are also central to noninflammatory clearance of innocuous agents. Here we investigated the role of a central humoral component of innate immunity, the lectin pathway of complement, in recognition of mitochondria in vitro and in vivo. We found that the soluble pattern recognition molecules, mannan-binding lectin (MBL), L-ficolin, and M-ficolin, were able to recognize mitochondria. Furthermore, MBL in complex with MBL-associated serine protease 2 (MASP-2) was able to activate the lectin pathway and deposit C4 onto mitochondria, suggesting that these molecules are involved either in homeostatic clearance of mitochondria or in induction of untoward inflammatory reactions. We found that following mitochondrial challenge, C3 was consumed in vivo in the absence of overt inflammation, indicating a potential role of complement in noninflammatory clearance of mitochondria. Thus, we report here the first indication of involvement of the lectin pathway in mitochondrial immune handling. PMID:23378531

  2. The Lectin Pathway of Complement and Rheumatic Heart Disease

    PubMed Central

    Beltrame, Marcia Holsbach; Catarino, Sandra Jeremias; Goeldner, Isabela; Boldt, Angelica Beate Winter; de Messias-Reason, Iara Jos

    2014-01-01

    The innate immune system is the first line of host defense against infection and is comprised of humoral and cellular mechanisms that recognize potential pathogens within minutes or hours of entry. The effector components of innate immunity include epithelial barriers, phagocytes, and natural killer cells, as well as cytokines and the complement system. Complement plays an important role in the immediate response against microorganisms, including Streptococcus sp. The lectin pathway is one of three pathways by which the complement system can be activated. This pathway is initiated by the binding of mannose-binding lectin (MBL), collectin 11 (CL-K1), and ficolins (Ficolin-1, Ficolin-2, and Ficolin-3) to microbial surface oligosaccharides and acetylated residues, respectively. Upon binding to target molecules, MBL, CL-K1, and ficolins form complexes with MBL-associated serine proteases 1 and 2 (MASP-1 and MASP-2), which cleave C4 and C2 forming the C3 convertase (C4b2a). Subsequent activation of complement cascade leads to opsonization, phagocytosis, and lysis of target microorganisms through the formation of the membrane-attack complex. In addition, activation of complement may induce several inflammatory effects, such as expression of adhesion molecules, chemotaxis and activation of leukocytes, release of reactive oxygen species, and secretion of cytokines and chemokines. In this chapter, we review the general aspects of the structure, function, and genetic polymorphism of lectin-pathway components and discuss most recent understanding on the role of the lectin pathway in the predisposition and clinical progression of Rheumatic Fever. PMID:25654073

  3. Involvement of the Lectin Pathway of Complement Activation in Antimicrobial Immune Defense during Experimental Septic Peritonitis

    PubMed Central

    Windbichler, Michaela; Echtenacher, Bernd; Hehlgans, Thomas; Jensenius, Jens C.; Schwaeble, Wilhelm; Mnnel, Daniela N.

    2004-01-01

    A critical first line of defense against infection is constituted by the binding of natural antibodies to microbial surfaces, activating the complement system via the classical complement activation pathway. In this function, the classical activation pathway is supported and amplified by two antibody-independent complement activation routes, i.e., the lectin pathway and the alternative pathway. We studied the contribution of the different complement activation pathways in the host defense against experimental polymicrobial peritonitis induced by cecal ligation and puncture by using mice deficient in either C1q or factors B and C2. The C1q-deficient mice lack the classical complement activation pathway. While infection-induced mortality of wild-type mice was 27%, mortality of C1q-deficient mice was increased to 60%. Mice with a deficiency of both factors B and C2 lack complement activation via the classical, the alternative, and the lectin pathways and exhibit a mortality of 92%, indicating a significant contribution of the lectin and alternative pathways of complement activation to survival. For 14 days after infection, mannan-binding lectin (MBL)-dependent activation of C4 was compromised. Serum MBL-A and MBL-C levels were significantly reduced for 1 week, possibly due to consumption. mRNA expression profiles did not lend support for either of the two MBL genes to respond as typical acute-phase genes. Our results demonstrate a long-lasting depletion of MBL-A and MBL-C from serum during microbial infection and underline the importance of both the lectin and the alternative pathways for antimicrobial immune defense. PMID:15322019

  4. The lectin pathway of complement activation contributes to protection from West Nile virus infection.

    PubMed

    Fuchs, Anja; Pinto, Amelia K; Schwaeble, Wilhelm J; Diamond, Michael S

    2011-03-30

    The function of the lectin pathway of complement activation in vivo against West Nile virus (WNV) or many other pathogenic viruses has not been defined. Mice deficient in lectin pathway recognition molecules (mannose binding lectin-A (MBL-A) and mannose binding lectin-C (MBL-C)) or the effector enzyme mannan-binding lectin-associated serine protease-2 (MASP-2), were more vulnerable to WNV infection than wild type mice. Compared with studies of mice deficient in factors of the classical or alternative pathway, MBL-A(-/-) MBL-C(-/-) or MASP-2(-/-) mice showed a less severe course of WNV infection. Indeed, a deficiency in lectin pathway activation did not significantly affect the kinetics of viral spread to the central nervous system (CNS) nor did it profoundly alter generation of adaptive B and T cell immune responses. We conclude that MBL-mediated recognition and lectin pathway activation have important yet subordinate functions in protecting against WNV infection and disease. PMID:21269656

  5. Fibrinogen-specific antibody induces abdominal aortic aneurysm in mice through complement lectin pathway activation

    PubMed Central

    Zhou, Hui-fang; Yan, Huimin; Bertram, Paula; Hu, Ying; Springer, Luke E.; Thompson, Robert W.; Curci, John A.; Hourcade, Dennis E.; Pham, Christine T. N.

    2013-01-01

    Abdominal aortic aneurysm (AAA) is a common vascular disease associated with high mortality rate due to progressive enlargement and eventual rupture. There is currently no established therapy known to alter the rate of aneurysmal expansion. Thus, understanding the processes that initiate and sustain aneurysmal growth is pivotal for the development of medical therapies aimed at halting disease progression. Using an elastase-induced AAA mouse model that recapitulates key features of human AAA, we previously reported that a natural IgG antibody directs alternative pathway complement activation and initiates the inflammatory process that culminates in aneurysmal development. The target of this natural antibody, however, was unknown. Herein we identify a natural IgG that binds to fibrinogen deposited in elastase-perfused aortic tissues, activates the complement lectin pathway (LP), and induces AAA. Moreover, we establish that alterations in the glycosylation patterns of this antibody critically affect its ability to activate the LP in vivo. We find that LP activation precedes the alternative pathway and absence of the LP complement protein mannan-binding lectin abrogates elastase-induced AAA. In human AAA tissues the mouse anti-fibrinogen antibody recognizes epitopes that localize to the same areas that stain positively for mannan-binding lectin, which suggests that the complement LP is engaged in humans as well. Lastly, we demonstrate that circulating antibodies in a subset of AAA patients react against fibrinogen or fibrinogen-associated epitopes in human aneurysmal tissues. Our findings support the concept that an autoimmune process directed at aortic wall self-antigens may play a central role in the immunopathogenesis of AAA. PMID:24167262

  6. Interaction of lectin pathway of complement-activating pattern recognition molecules with mycobacteria.

    PubMed

    Bartlomiejczyk, M A; Swierzko, A S; Brzostek, A; Dziadek, J; Cedzynski, M

    2014-11-01

    We have demonstrated that mannose-binding lectin (MBL) recognizes various slow-growing, pathogenic mycobacteria [Mycobacterium?tuberculosis (MTB), M.?bovis, M.?kansasii, M.?gordonae] as well as non-pathogenic M.?smegmatis. Recognition resulted in activation of the lectin pathway (LP) of complement and an enhancement of phagocytosis (shown for M.?tuberculosis). Although MBL may be considered the main factor activating the LP upon recognition of mycobacteria, involvement of ficolins has also to be considered. Interaction of ficolin-3 with M.?tuberculosis, M.?bovis and M.?kansasii, and ficolin-1 with M.?tuberculosis and M.?bovis was shown for the first time. Binding of recombinant MBL or ficolin-3 to MTB H37 Rv led to the agglutination of bacteria and promoted their phagocytosis, but little effect was apparent with ficolin-1 or ficolin-2. Data from Western blots suggest mannosylated lipoarabinomannan (ManLAM) to be one of the main cell components of slow-growing mycobacteria, involved in LP activation. However, the LP was also activated by other cell fractions. Results presented here supplement considerably the data concerning the ability of complement-activating lectins to interact with mycobacteria. Ficolins (especially ficolin-3) might influence host response to infection and thus have clinical significance, at least as disease modifiers. PMID:25041480

  7. Components of the lectin pathway of complement activation in paediatric patients of intensive care units.

    PubMed

    Świerzko, Anna S; Szala-Poździej, Agnieszka; Kilpatrick, David C; Sobociński, Michał; Chojnacka, Karolina; Sokołowska, Anna; Michalski, Mateusz; Mazerant, Karolina; Jensenius, Jens C; Matsushita, Misao; Krajewski, Wojciech R; Szczapa, Jerzy; Bąk-Romaniszyn, Leokadia; Zeman, Krzysztof; Cedzyński, Maciej

    2016-05-01

    Infections are a major cause of childhood mortality. We investigated components of the lectin pathway of complement activation in the context of sepsis at both genetic and protein levels in neonates, infants and older children. Major components of the lectin pathway and two genes for Toll-like receptors were studied in 87 neonates with confirmed sepsis and compared with 40 babies with infections who did not develop sepsis (disease controls) and 273 infection-free neonatal controls. A second cohort comprised 47 older children with sepsis and 87 controls. Low MBL-conferring genotypes (LXA/O+O/O) were more frequent in sepsis patients than in healthy controls but no significant differences in the frequency of SNPs of other lectin pathway genes (FCN1, FCN2, FCN3, MASP1/3, MASP2) or TLR receptor genes (TLR2, TLR4) were found. One case of primary MASP-2 deficiency was found among healthy pre-terms and one neonate suffering from SIRS was heterozygous for the rare FCN1 gene mutation, +6658 G>A. Generally, sepsis was associated with low serum MBL and low ficolin-2 concentrations on admission. Among neonates, ficolin-1 and MASP-2 levels were elevated in sepsis relative to healthy, but not disease, controls. Unlike neonates, ficolin-3 and MASP-2 levels were lower in older patients than in healthy controls while no difference was found for ficolin-1. With the possible exception of MBL, inherited lectin pathway insufficiencies do not seem to predispose to sepsis, rather changes in protein concentrations reflect alterations in disease course. PMID:26850322

  8. Role of ficolin-A and lectin complement pathway in the innate defense against pathogenic Aspergillus species.

    PubMed

    Bidula, Stefan; Kenawy, Hany; Ali, Youssif M; Sexton, Darren; Schwaeble, Wilhelm J; Schelenz, Silke

    2013-05-01

    Aspergillus species are saprophytic molds causing life-threatening invasive fungal infections in the immunocompromised host. Innate immune recognition, in particular, the mechanisms of opsonization and complement activation, has been reported to be an integral part of the defense against fungi. We have shown that the complement component ficolin-A significantly binds to Aspergillus conidia and hyphae in a concentration-dependent manner and was inhibited by N-acetylglucosamine and N-acetylgalactosamine. Calcium-independent binding to Aspergillus fumigatus and A. terreus was observed, but binding to A. flavus and A. niger was calcium dependent. Ficolin-A binding to conidia was increased under low-pH conditions, and opsonization led to enhanced binding of conidia to A549 airway epithelial cells. In investigations of the lectin pathway of complement activation, ficolin-A-opsonized conidia did not lead to lectin pathway-specific C4 deposition. In contrast, the collectin mannose binding lectin C (MBL-C) but not MBL-A led to efficient lectin pathway activation on A. fumigatus in the absence of ficolin-A. In addition, ficolin-A opsonization led to a modulation of the proinflammatory cytokine interleukin-8. We conclude that ficolin-A may play an important role in the innate defense against Aspergillus by opsonizing conidia, immobilizing this fungus through enhanced adherence to epithelial cells and modulation of inflammation. However, it appears that other immune pattern recognition molecules, i.e., those of the collectin MBL-C, are involved in the Aspergillus-lectin complement pathway activation rather than ficolin-A. PMID:23478320

  9. Role of Ficolin-A and Lectin Complement Pathway in the Innate Defense against Pathogenic Aspergillus Species

    PubMed Central

    Bidula, Stefan; Kenawy, Hany; Ali, Youssif M.; Sexton, Darren; Schwaeble, Wilhelm J.

    2013-01-01

    Aspergillus species are saprophytic molds causing life-threatening invasive fungal infections in the immunocompromised host. Innate immune recognition, in particular, the mechanisms of opsonization and complement activation, has been reported to be an integral part of the defense against fungi. We have shown that the complement component ficolin-A significantly binds to Aspergillus conidia and hyphae in a concentration-dependent manner and was inhibited by N-acetylglucosamine and N-acetylgalactosamine. Calcium-independent binding to Aspergillus fumigatus and A. terreus was observed, but binding to A. flavus and A. niger was calcium dependent. Ficolin-A binding to conidia was increased under low-pH conditions, and opsonization led to enhanced binding of conidia to A549 airway epithelial cells. In investigations of the lectin pathway of complement activation, ficolin-A-opsonized conidia did not lead to lectin pathway-specific C4 deposition. In contrast, the collectin mannose binding lectin C (MBL-C) but not MBL-A led to efficient lectin pathway activation on A. fumigatus in the absence of ficolin-A. In addition, ficolin-A opsonization led to a modulation of the proinflammatory cytokine interleukin-8. We conclude that ficolin-A may play an important role in the innate defense against Aspergillus by opsonizing conidia, immobilizing this fungus through enhanced adherence to epithelial cells and modulation of inflammation. However, it appears that other immune pattern recognition molecules, i.e., those of the collectin MBL-C, are involved in the Aspergillus-lectin complement pathway activation rather than ficolin-A. PMID:23478320

  10. MASP-3 and its association with distinct complexes of the mannan-binding lectin complement activation pathway.

    PubMed

    Dahl, M R; Thiel, S; Matsushita, M; Fujita, T; Willis, A C; Christensen, T; Vorup-Jensen, T; Jensenius, J C

    2001-07-01

    The mannan-binding lectin (MBL) pathway of complement activation is part of the innate immune defense. The binding of MBL to microbial carbohydrates activates the MBL-associated serine proteases (MASPs), which recruit the complement factors, C4 and C2, to generate the C3 convertase or directly activate C3. We present a phylogenetically highly conserved member of the MBL complex, MASP-3, which is generated through alternative splicing of the MASP-1/3 gene. The designation of MASP-3 as a protease is based on homology to known MASPs. Different MBL oligomers were found to have distinct MASP composition and biological activities. MASP-1, MAp19, and direct C3-cleaving activity are associated with smaller oligomers whereas MASP-3 is found together with MASP-2 on larger oligomers. MASP-3 downregulate the C4 and C2 cleaving activity of MASP-2. PMID:11485744

  11. Deletion of wboA Enhances Activation of the Lectin Pathway of Complement in Brucella abortus and Brucella melitensis

    PubMed Central

    Fernandez-Prada, Carmen M.; Nikolich, Mikeljon; Vemulapalli, Ramesh; Sriranganathan, Nammalwar; Boyle, Stephen M.; Schurig, Gerhardt G.; Hadfield, Ted L.; Hoover, David L.

    2001-01-01

    Brucella spp. are gram-negative intracellular pathogens that survive and multiply within phagocytic cells of their hosts. Smooth organisms present O polysaccharides (OPS) on their surface. These OPS help the bacteria avoid the bactericidal action of serum. The wboA gene, coding for the enzyme glycosyltransferase, is essential for the synthesis of O chain in Brucella. In this study, the sensitivity to serum of smooth, virulent Brucella melitensis 16M and B. abortus 2308, rough wboA mutants VTRM1, RA1, and WRR51 derived from these two Brucella species, and the B. abortus vaccine strain RB51 was assayed using normal nonimmune human serum (NHS). The deposition of complement components and mannose-binding lectin (MBL) on the bacterial surface was detected by flow cytometry. Rough B. abortus mutants were more sensitive to the bactericidal action of NHS than were rough B. melitensis mutants. Complement components were deposited on smooth strains at a slower rate compared to rough strains. Deposition of iC3b and C5b-9 and bacterial killing occurred when bacteria were treated with C1q-depleted, but not with C2-depleted serum or NHS in the presence of Mg-EGTA. These results indicate that (i) OPS-deficient strains derived from B. melitensis 16M are more resistant to the bactericidal action of NHS than OPS-deficient strains derived from B. abortus 2308, (ii) both the classical and the MBL-mediated pathways are involved in complement deposition and complement-mediated killing of Brucella, and (iii) the alternative pathway is not activated by smooth or rough brucellae. PMID:11401980

  12. Genetically engineered fusion of MAP-1 and factor H domains 1-5 generates a potent dual upstream inhibitor of both the lectin and alternative complement pathways.

    PubMed

    Nordmaj, Mie Anemone; Munthe-Fog, Lea; Hein, Estrid; Skjoedt, Mikkel-Ole; Garred, Peter

    2015-12-01

    Inhibition of the complement cascade has emerged as an option for treatment of a range of diseases. Mannose-binding lectin/ficolin/collectin-associated protein (MAP-1) is a pattern recognition molecule (PRM)-associated inhibitor of the lectin pathway. The central regulator of the alternative pathway (AP) is complement factor H (FH). Our aim was to design a dual upstream inhibitor of both human lectin and APs by fusing MAP-1 with a part of FH. There were 2 different recombinant chimeric proteins comprising full-length human MAP-1 and the first 5 N-terminal domains of human FH designed. The FH domains were orientated either in the N- or C-terminal part of MAP-1. The complement inhibition potential in human serum was assessed. Both chimeric constructs displayed the characteristics of the native molecules and bound to the PRMs with an EC50 of ?2 nM. However, when added to serum diluted 1:4 in a solid-phase functional assay, only the first 5 N-terminal domains of complement FH fused to the C-terminal part of full-length MAP-1 chimeric construct were able to combine inhibition of lectin and AP activation with an half maximal inhibitory concentration of ?100 and 20 nM, respectively. No effect was seen on the classical pathway. Fusion of MAP-1 with FH domains represents a novel therapeutic approach for selective targeting upstream and central complement activation at sites of inflammation.-Nordmaj, M. A., Munthe-Fog, L., Hein, E., Skjoedt, M.-O., Garred, P. Genetically engineered fusion of MAP-1 and factor H domains 1-5 generates a potent dual upstream inhibitor of both the lectin and alternative complement pathways. PMID:26260032

  13. Studies of the binding of ficolin-2 and ficolin-3 from the complement lectin pathway to Leptospira biflexa, Pasteurella pneumotropica and Diarrheagenic Escherichia coli.

    PubMed

    Sahagún-Ruiz, Alfredo; Breda, Leandro Carvalho Dantas; Valencia, Mónica Marcela Castiblanco; Elias, Waldir P; Munthe-Fog, Lea; Garred, Peter; Barbosa, Angela Silva; Isaac, Lourdes

    2015-10-01

    Ficolins recognize pathogen associated molecular patterns and activate the lectin pathway of complement system. However, our knowledge regarding pathogen recognition of human ficolins is still limited. We therefore set out to explore and investigate the possible interactions of the two main serum ficolins, ficolin-2 and ficolin-3 with different Gram-negative bacteria. We used recombinant ficolin molecules and normal human serum, which were detected with anti-ficolin monoclonal antibodies. In addition we investigated the capacity of these pathogens to activate the lectin pathway of complement system. We show for the first time that human ficolin-2 recognizes the nonpathogenic spirochete Leptospira biflexa serovar Patoc, but not the pathogenic Leptospira interrogans serovar Kennewicki strain Fromm. Additionally, human ficolin-2 and ficolin-3 recognize pathogenic Pasteurella pneumotropica, enteropathogenic Escherichia coli (EPEC) serotype O111ab:H2 and enteroaggregative E. coli (EAEC) serogroup O71 but not four enterohemorrhagic E. coli, three EPEC, three EAEC and two nonpathogenic E. coli strains (DH5α and HB101). The lectin pathway was activated by Pasteurella pneumotropica, EPEC O111ab:H2 and EAEC O71 after incubation with C1q depleted human serum. In conclusion, this study provide novel insight in the binding and complement activating capacity of the lectin pathway initiation molecules ficolin-2 and ficolin-3 towards relevant Gram-negative pathogens of pathophysiological relevance. PMID:26074063

  14. Inhibition of the classical and lectin pathway of the complement system by recombinant LAIR-2.

    PubMed

    Olde Nordkamp, Marloes J M; Boross, Peter; Yildiz, Cafer; Jansen, J H Marco; Leusen, Jeanette H W; Wouters, Diana; Urbanus, Rolf T; Hack, C Erik; Meyaard, Linde

    2014-01-01

    Activation of complement may cause severe tissue damage in antibody-mediated allograft rejection and other antibody-mediated clinical conditions; therefore, novel potent complement inhibitors are needed. Previously, we described binding of the inhibitory receptor LAIR-1 and its soluble family member LAIR-2 to collagen. Here, we investigated binding of LAIR-1 and LAIR-2 to the complement proteins C1q and MBL, which both have collagen-like domains, and evaluated the effect of this binding on complement function. We demonstrate specific binding of recombinant LAIR proteins to both C1q and MBL. Surface plasmon resonance experiments showed that LAIR-2-Fc protein bound C1q and MBL with the highest affinity compared to LAIR-2-HIS. We, therefore, hypothesized that LAIR-2-Fc is a potent complement inhibitor. Indeed, LAIR-2-Fc inhibited C4 fixation to IgG or mannan, reduced activation of C4 by aggregated IgG in plasma and inhibited iC3b deposition on cells. Finally, LAIR-2-Fc inhibited complement-mediated lysis of cells sensitized with anti-HLA antibodies in an ex vivo model for antibody-mediated transplant rejection. Thus, LAIR-2-Fc is an effective novel complement inhibitor for the treatment and prevention of antibody-mediated allograft rejection and antibody-mediated clinical conditions. PMID:24192271

  15. The Group B Streptococcus-Secreted Protein CIP Interacts with C4, Preventing C3b Deposition via the Lectin and Classical Complement Pathways.

    PubMed

    Pietrocola, Giampiero; Rindi, Simonetta; Rosini, Roberto; Buccato, Scilla; Speziale, Pietro; Margarit, Immaculada

    2016-01-01

    The group B Streptococcus (GBS) is a leading cause of neonatal invasive disease. GBS bacteria are surrounded by a thick capsular polysaccharide that is a potent inhibitor of complement deposition via the alternative pathway. Several of its surface molecules can however activate the classical and lectin complement pathways, rendering this species still vulnerable to phagocytic killing. In this study we have identified a novel secreted protein named complement interfering protein (CIP) that downregulates complement activation via the classical and lectin pathways, but not the alternative pathway. The CIP protein showed high affinity toward C4b and inhibited its interaction with C2, presumably preventing the formation of the C4bC2a convertase. Addition of recombinant CIP to GBS cip-negative bacteria resulted in decreased deposition of C3b on their surface and in diminished phagocytic killing in a whole-blood assay. Our data reveal a novel strategy exploited by GBS to counteract innate immunity and could be valuable for the development of anti-infective agents against this important pathogen. PMID:26608922

  16. The Group B Streptococcus–Secreted Protein CIP Interacts with C4, Preventing C3b Deposition via the Lectin and Classical Complement Pathways

    PubMed Central

    Pietrocola, Giampiero; Rindi, Simonetta; Rosini, Roberto; Buccato, Scilla

    2016-01-01

    The group B Streptococcus (GBS) is a leading cause of neonatal invasive disease. GBS bacteria are surrounded by a thick capsular polysaccharide that is a potent inhibitor of complement deposition via the alternative pathway. Several of its surface molecules can however activate the classical and lectin complement pathways, rendering this species still vulnerable to phagocytic killing. In this study we have identified a novel secreted protein named complement interfering protein (CIP) that downregulates complement activation via the classical and lectin pathways, but not the alternative pathway. The CIP protein showed high affinity toward C4b and inhibited its interaction with C2, presumably preventing the formation of the C4bC2a convertase. Addition of recombinant CIP to GBS cip-negative bacteria resulted in decreased deposition of C3b on their surface and in diminished phagocytic killing in a whole-blood assay. Our data reveal a novel strategy exploited by GBS to counteract innate immunity and could be valuable for the development of anti-infective agents against this important pathogen. PMID:26608922

  17. Complement Activation by Giardia duodenalis Parasites through the Lectin Pathway Contributes to Mast Cell Responses and Parasite Control.

    PubMed

    Li, Erqiu; Tako, Ernest A; Singer, Steven M

    2016-04-01

    Infection withGiardia duodenalisis one of the most common causes of diarrheal disease in the world. While numerous studies have identified important contributions of adaptive immune responses to parasite control, much less work has examined innate immunity and its connections to the adaptive response during this infection. We explored the role of complement in immunity toGiardiausing mice deficient in mannose-binding lectin (Mbl2) or complement factor 3a receptor (C3aR). Both strains exhibited delayed clearance of parasites and a reduced ability to recruit mast cells in the intestinal submucosa. C3aR-deficient mice had normal production of antiparasite IgA, butex vivoT cell recall responses were impaired. These data suggest that complement is a key factor in the innate recognition ofGiardiaand that recruitment of mast cells and activation of T cell immunity through C3a are important for parasite control. PMID:26831470

  18. H-ficolin binds Aspergillus fumigatus leading to activation of the lectin complement pathway and modulation of lung epithelial immune responses.

    PubMed

    Bidula, Stefan; Sexton, Darren W; Yates, Matthew; Abdolrasouli, Alireza; Shah, Anand; Wallis, Russell; Reed, Anna; Armstrong-James, Darius; Schelenz, Silke

    2015-10-01

    Aspergillus fumigatus is an opportunistic fungal pathogen that typically infects the lungs of immunocompromised patients leading to a high mortality. H-Ficolin, an innate immune opsonin, is produced by type II alveolar epithelial cells and could participate in lung defences against infections. Here, we used the human type II alveolar epithelial cell line, A549, to determine the involvement of H-ficolin in fungal defence. Additionally, we investigated the presence of H-ficolin in bronchoalveolar lavage fluid from transplant patients during pneumonia. H-Ficolin exhibited demonstrable binding to A.fumigatus conidia via l-fucose, d-mannose and N-acetylglucosamine residues in a calcium- and pH-dependent manner. Moreover, recognition led to lectin complement pathway activation and enhanced fungal association with A549 cells. Following recognition, H-ficolin opsonization manifested an increase in interleukin-8 production from A549 cells, which involved activation of the intracellular signalling pathways mitogen-activated protein kinase MAPK kinase 1/2, p38 MAPK and c-Jun N-terminal kinase. Finally, H-ficolin concentrations were significantly higher in bronchoalveolar lavage fluid of patients with lung infections compared with control subjects (n=16; P=000726). Receiver operating characteristics curve analysis further highlighted the potential of H-ficolin as a diagnostic marker for lung infection (area under the curve=077; P<00001). Hence, H-ficolin participates in A.fumigatus defence through the activation of the lectin complement pathway, enhanced fungus-host interactions and modulated immune responses. PMID:26133042

  19. Essential Role for the Lectin Pathway in Collagen Antibody-Induced Arthritis Revealed Through Use of Adenovirus Programming Complement Inhibitor MAp44 Expression

    PubMed Central

    Banda, Nirmal K.; Mehta, Gaurav; Kjaer, Troels R.; Takahashi, Minoru; Schaack, Jerome; Morrison, Thomas E.; Thiel, Steffen; Arend, William P.; Holers, V. Michael

    2014-01-01

    Previous studies using mannose-binding lectin (MBL) and complement C4 deficient mice have suggested that the lectin pathway (LP) is not required for the development of inflammatory arthritis in the collagen antibody-induced arthritis (CAIA) model. MBL, ficolins and collectin-11 are key LP pattern recognition molecules that associate with three serine proteases, MASP-1, MASP-2 and MASP-3, and also with two MBL-associated proteins designated sMAP and MAp44. Recent studies have shown that MAp44, an alternatively spliced product of the MASP-1/3 gene, is a competitive inhibitor of the binding of the recognition molecules to all three MASPs. In these studies we examined the effect of treatment of mice with adenovirus (Ad) programmed to express human MAp44 (AdhMAp44) on the development of CAIA. AdhMAp44 and Ad programming Green fluorescent protein (AdGFP) expression were injected intraperitoneally in C57BL/6 wild-type mice prior to the induction of CAIA. AdhMAp44 significantly reduced the clinical disease activity score (CDA) by 81% compared to mice injected with AdGFP. Similarly, histopathologic injury scores for inflammation, pannus, cartilage and bone damage, as well as C3 deposition in the cartilage and synovium, were significantly reduced by AdhMAp44 pretreatment. Mice treated with AdmMAp44, programming expression of mouse MAp44, also showed significantly decreased CDA and histopathologic injury scores. Additionally, administration of AdhMAp44 significantly diminished the severity of Ross River Virus-induced arthritis, a LP-dependent model. Our study provides conclusive evidence that an intact complement LP is essential to initiate CAIA, and that MAp44 may be an appropriate treatment for inflammatory arthritis. PMID:25070856

  20. Low Invasiveness of Pneumococcal Serotype 11A Is Linked to Ficolin-2 Recognition of O-acetylated Capsule Epitopes and Lectin Complement Pathway Activation

    PubMed Central

    Brady, Allison M.; Calix, Juan J.; Yu, Jigui; Geno, Kimball Aaron; Cutter, Gary R.; Nahm, Moon H.

    2014-01-01

    Background.?The divergent epidemiological behavior of Streptococcus pneumoniae serotypes suggests that serotype-specific features such as capsule O-acetylation influence the propensity of a strain to cause invasive pneumococcal disease (IPD). We hypothesize that innate host factors mediate the observed negative association between IPD and the serotype 11A (ST11A) capsule O-acetyltransferase gene, wcjE. Methods.?We evaluated the ability of ficolin-2, an initiator of the lectin complement pathway that was previously shown to bind ST11A pneumococci, to recognize and mediate complement-dependent opsonophagocytosis of different pneumococcal serotypes. We supplemented findings with an epidemiological meta-analysis comparing invasiveness of the 30 most prevalent pneumococcal serotypes. Results.?Ficolin-2 bound ST11A capsule polysaccharide and other wcjE-containing pneumococcal serotypes, except ST9V and ST20B. Ficolin-2 did not bind wcjE-null serotypes, including the wcjE-null variant of ST11A, ST11E. We observed C1q-independent complement deposition and phagocytic killing of pneumococci expressing ST11A but not those expressing ST11E. Inhibition of ficolin-2 binding abrogated ST11A-associated complement deposition and phagocytosis. In children, invasiveness of ST11A was the lowest among serotypes tested in our meta-analysis, while ST9V was among the highest. Conclusions.?Ficolin-2 mediates serum protection by recognizing specific O-acetylated epitopes of pneumococcal capsule polysaccharides, exemplifying a novel host-microbe interaction that innately offers serotype-specific immunity to IPD. PMID:24683196

  1. The Extracellular Adherence Protein from Staphylococcus aureus Inhibits the Classical and Lectin Pathways of Complement by Blocking Formation of the C3 Pro-Convertase

    PubMed Central

    Garcia, Brandon L.; Ramyar, Kasra X.; Keightley, Andrew; Ruyken, Maartje; Syriga, Maria; Sfyroera, Georgia; Weber, Alexander B.; Zolkiewski, Michal; Ricklin, Daniel; Lambris, John D.; Rooijakkers, Suzan H.M.; Geisbrecht, Brian V.

    2014-01-01

    The pathogenic bacterium Staphylococcus aureus actively evades many aspects of human innate immunity by expressing a series of small inhibitory proteins. A number of these proteins inhibit the complement system, which labels bacteria for phagocytosis and generates inflammatory chemoattractants. While the majority of staphylococcal complement inhibitors act on the alternative pathway (AP) to block the amplification loop, only a few proteins act on the initial recognition cascades that constitute the classical (CP) and lectin (LP) pathways. We screened a collection of recombinant, secreted staphylococcal proteins to determine if S. aureus produces other molecules that inhibit either the CP and/or LP. Using this approach, we identified the extracellular adherence protein (Eap) as a potent, specific inhibitor of both the CP and LP. We found that Eap blocked CP/LP-dependent activation of C3, but not C4, and that Eap likewise inhibited deposition of C3b on the surface of S. aureus cells. In turn, this significantly diminished the extent of S. aureus opsonophagocytosis and killing by neutrophils. This combination of functional properties suggested that Eap acts specifically at the level of the CP/LP C3 convertase (C4b2a). Indeed, we demonstrated a direct, nanomolar-affinity interaction of Eap with C4b. Eap binding to C4b inhibited binding of both full-length C2 and its C2b fragment, which indicated that Eap disrupts formation of the CP/LP C3 pro-convertase (C4b2). As a whole, our results demonstrate that S. aureus inhibits the two initiation routes of complement by expression of the Eap protein, and thereby define a novel mechanism of immune evasion. PMID:25381436

  2. Complement-mediated neutralization of dengue virus requires mannose-binding lectin.

    PubMed

    Avirutnan, Panisadee; Hauhart, Richard E; Marovich, Mary A; Garred, Peter; Atkinson, John P; Diamond, Michael S

    2011-01-01

    Mannose-binding lectin (MBL) is a key soluble pathogen recognition protein of the innate immune system that binds specific mannose-containing glycans on the surfaces of microbial agents and initiates complement activation via the lectin pathway. Prior studies showed that MBL-dependent activation of the complement cascade neutralized insect cell-derived West Nile virus (WNV) in cell culture and restricted pathogenesis in mice. Here, we investigated the antiviral activity of MBL in infection by dengue virus (DENV), a related flavivirus. Using a panel of nave sera from mouse strains deficient in different complement components, we showed that inhibition of infection by insect cell- and mammalian cell-derived DENV was primarily dependent on the lectin pathway. Human MBL also bound to DENV and neutralized infection of all four DENV serotypes through complement activation-dependent and -independent pathways. Experiments with human serum from nave individuals with inherent variation in the levels of MBL in blood showed a direct correlation between the concentration of MBL and neutralization of DENV; samples with high levels of MBL in blood neutralized DENV more efficiently than those with lower levels. Our studies suggest that allelic variation of MBL in humans may impact complement-dependent control of DENV pathogenesis. IMPORTANCE Dengue virus (DENV) is a mosquito-transmitted virus that causes a spectrum of clinical disease in humans ranging from subclinical infection to dengue hemorrhagic fever and dengue shock syndrome. Four serotypes of DENV exist, and severe illness is usually associated with secondary infection by a different serotype. Here, we show that mannose-binding lectin (MBL), a pattern recognition molecule that initiates the lectin pathway of complement activation, neutralized infection of all four DENV serotypes through complement activation-dependent and -independent pathways. Moreover, we observed a direct correlation with the concentration of MBL in human serum and neutralization of DENV infection. Our studies suggest that common genetic polymorphisms that result in disparate levels and function of MBL in humans may impact DENV infection, pathogenesis, and disease severity. PMID:22167226

  3. A second serine protease associated with mannan-binding lectin that activates complement.

    PubMed

    Thiel, S; Vorup-Jensen, T; Stover, C M; Schwaeble, W; Laursen, S B; Poulsen, K; Willis, A C; Eggleton, P; Hansen, S; Holmskov, U; Reid, K B; Jensenius, J C

    1997-04-01

    The complement system comprises a complex array of enzymes and non-enzymatic proteins that is essential for the operation of the innate as well as the adaptive immune defence. The complement system can be activated in three ways: by the classical pathway which is initiated by antibody-antigen complexes, by the alternative pathway initiated by certain structures on microbial surfaces, and by an antibody-independent pathway that is initiated by the binding of mannan-binding lectin (MBL; first described as mannan-binding protein) to carbohydrates. MBL is structurally related to the complement C1 subcomponent, C1q, and seems to activate the complement system through an associated serine protease known as MASP (ref. 4) or p100 (ref. 5), which is similar to C1r and C1s of the classical pathway. MBL binds to specific carbohydrate structures found on the surface of a range of microorganisms, including bacteria, yeasts, parasitic protozoa and viruses, and exhibits antibacterial activity through killing mediated by the terminal, lytic complement components or by promoting phagocytosis. The level of MBL in plasma is genetically determined, and deficiency is associated with frequent infections in childhood, and possibly also in adults (for review, see ref. 6). We have now identified a new MBL-associated serine protease (MASP-2) which shows a striking homology with the previously reported MASP (MASP-1) and the two C1q-associated serine proteases C1r and C1s. Thus complement activation through MBL, like the classical pathway, involves two serine proteases and may antedate the development of the specific immune system of vertebrates. PMID:9087411

  4. Increased Autoreactivity of the Complement-Activating Molecule Mannan-Binding Lectin in a Type 1 Diabetes Model

    PubMed Central

    Østergaard, Jakob Appel; Ruseva, Marieta Milkova; Malik, Talat Habib; Hoffmann-Petersen, Ingeborg Torp; Pickering, Matthew Caleb; Thiel, Steffen; Hansen, Troels Krarup

    2016-01-01

    Background. Diabetic kidney disease is the leading cause of end-stage renal failure despite intensive treatment of modifiable risk factors. Identification of new drug targets is therefore of paramount importance. The complement system is emerging as a potential new target. The lectin pathway of the complement system, initiated by the carbohydrate-recognition molecule mannan-binding lectin (MBL), is linked to poor kidney prognosis in diabetes. We hypothesized that MBL activates complement upon binding within the diabetic glomerulus. Methods. We investigated this by comparing complement deposition and activation in kidneys from streptozotocin-induced diabetic mice and healthy control mice. Results. After 20 weeks of diabetes, glomerular deposition of MBL was significantly increased. Diabetic animals had 2.0-fold higher (95% CI 1.6–2.5) immunofluorescence intensity from anti-MBL antibodies compared with controls (P < 0.001). Diabetes and control groups did not differ in glomerular immunofluorescence intensity obtained by antibodies against complement factors C4, C3, and C9. However, the circulating complement activation product C3a was increased in diabetes as compared to control mice (P = 0.04). Conclusion. 20 weeks of diabetes increased MBL autoreactivity in the kidney and circulating C3a concentration. Together with previous findings, these results indicate direct effects of MBL within the kidney in diabetes. PMID:26977416

  5. Increased Autoreactivity of the Complement-Activating Molecule Mannan-Binding Lectin in a Type 1 Diabetes Model.

    PubMed

    Østergaard, Jakob Appel; Ruseva, Marieta Milkova; Malik, Talat Habib; Hoffmann-Petersen, Ingeborg Torp; Pickering, Matthew Caleb; Thiel, Steffen; Hansen, Troels Krarup

    2016-01-01

    Background. Diabetic kidney disease is the leading cause of end-stage renal failure despite intensive treatment of modifiable risk factors. Identification of new drug targets is therefore of paramount importance. The complement system is emerging as a potential new target. The lectin pathway of the complement system, initiated by the carbohydrate-recognition molecule mannan-binding lectin (MBL), is linked to poor kidney prognosis in diabetes. We hypothesized that MBL activates complement upon binding within the diabetic glomerulus. Methods. We investigated this by comparing complement deposition and activation in kidneys from streptozotocin-induced diabetic mice and healthy control mice. Results. After 20 weeks of diabetes, glomerular deposition of MBL was significantly increased. Diabetic animals had 2.0-fold higher (95% CI 1.6-2.5) immunofluorescence intensity from anti-MBL antibodies compared with controls (P < 0.001). Diabetes and control groups did not differ in glomerular immunofluorescence intensity obtained by antibodies against complement factors C4, C3, and C9. However, the circulating complement activation product C3a was increased in diabetes as compared to control mice (P = 0.04). Conclusion. 20 weeks of diabetes increased MBL autoreactivity in the kidney and circulating C3a concentration. Together with previous findings, these results indicate direct effects of MBL within the kidney in diabetes. PMID:26977416

  6. Diagnosis of complement alternative pathway disorders.

    PubMed

    Angioi, Andrea; Fervenza, Fernando C; Sethi, Sanjeev; Zhang, Yuzhou; Smith, Richard J; Murray, David; Van Praet, Jens; Pani, Antonello; De Vriese, An S

    2016-02-01

    Kidney diseases resulting from abnormal control of the complement alternative pathway include atypical hemolytic uremic syndrome, C3 glomerulonephritis, and dense-deposit disease, as well as atypical postinfectious glomerulonephritis. Although clinically diverse, they all result from loss of surface or fluid-phase complement control, caused by acquired or genetic defects in the complement alternative pathway. As such, the diagnostic approach is similar and includes a comprehensive biochemical, genetic, and pathologic analysis of the complement pathway. The biochemical test battery includes functional activity measurements of the entire complement pathway, functional and quantitative analysis of individual components and regulators, and quantification of activation products. In patients with a thrombotic microangiopathy, ADAMTS-13 activity should be determined to exclude a thrombotic thrombocytopenic purpura. The spectrum of genes currently known to be involved in the pathogenesis of alternative pathway disorders is rapidly expanding. Pathologic analysis of a kidney biopsy specimen is sophisticated with ad hoc immunofluorescence studies and laser microdissection with mass spectrometry. The identification of the underlying defect in the alternative pathway based on this comprehensive analysis will allow treatment to be directed to the site of dysregulation. PMID:26806831

  7. Lectin Complement Protein Collectin 11 (CL-K1) and Susceptibility to Urinary Schistosomiasis

    PubMed Central

    Antony, Justin S.; Ojurongbe, Olusola; Kremsner, Peter G.; Velavan, Thirumalaisamy P.

    2015-01-01

    Background Urinary Schistosomiasis is a neglected tropical disease endemic in many sub Saharan -African countries. Collectin Kidney 1 (CL-K1, encoded by COLEC11 on chromosome 2p25.3), a member of the vertebrate C-type lectin super family, has recently been identified as pattern-recognition molecule (PRR) of the lectin complement pathway. CL-K1 is preferentially expressed in the kidneys, but also in other organs and it is considered to play a role in host defense to some infectious agents. Schistosome teguments are fucosylated and CL-K1 has, through its collagen-like domain, a high binding affinity to fucose. Methodology/Principal Findings We utilized a Nigerian study group consisting of 167 Schistosoma haematobium infected individuals and 186 matched healthy subjects, and investigated the contribution of CL-K1 deficiency and of COLEC11 polymorphisms to infection phenotype. Higher CL-K1 serum levels were associated with decreased risk of schistosome infection (Pcorr = 0.0004). CL-K1 serum levels were differentially distributed between the COLEC11 genotypes and haplotypes observed. The non-synonymous variant p.R216H was associated with the occurrence of schistosomiasis (OR = 0.44, 95%CI = 0.220.72, Pcorr = 0.0004). The reconstructed COLEC11*TCCA haplotypes were associated with higher CL-K1 serum levels (P = 0.002) and with decreased schistosomiasis (OR = 0.38, 95%CI = 0.230.63, Pcorr = 0.0001). Conclusions In agreement with findings from our earlier published study, our findings support the observation that CL-K1 and their functional variants may be host factors associated with protection in schistosomiasis and may be a useful marker for further investigations. PMID:25807310

  8. A novel IgMH-Ficolin complement pathway to attack allogenic cancer cells in vitro

    PubMed Central

    Lei, Xiaoying; Liu, Chaoxu; Azadzoi, Kazem; Li, Cuiling; Lu, Fan; Xiang, An; Sun, Jianbin; Guo, Yanhai; Zhao, Qingchuan; Yan, Zhen; Yang, Jinghua

    2015-01-01

    The pentameric serum IgMs are critical to immune defense and surveillance through cytotoxicity against microbes and nascent cancer cells. Ficolins, a group of oligomeric lectins with an overall structure similar to C1q and mannose-binding lectin (MBL) participate in microbe infection and apoptotic cell clearance by activating the complement lectin pathway or a primitive opsonophagocytosis. It remains unknown whether serum IgMs interplay with ficolins in cancer immunosurveillance. Here we report a natural cancer killing of different types of cancer cells by sera from a healthy human population mediated by a novel IgMH-ficolin complement activation pathway. We demonstrate for the first time that H-ficolin bound to a subset of IgMs in positive human sera and IgMH-ficolin deposited on cancer cells to activate complement attack in cancer cells. Our data suggest that the IgMH-ficolin -mediated complement activation pathway may be another defensive strategy for human cancer immunosurveillance. PMID:25592840

  9. SALO, a novel classical pathway complement inhibitor from saliva of the sand fly Lutzomyia longipalpis.

    PubMed

    Ferreira, Viviana P; Fazito Vale, Vladimir; Pangburn, Michael K; Abdeladhim, Maha; Ferreira Mendes-Sousa, Antonio; Coutinho-Abreu, Iliano V; Rasouli, Manoochehr; Brandt, Elizabeth A; Meneses, Claudio; Lima, Kolyvan Ferreira; Nascimento Arajo, Ricardo; Horcio Pereira, Marcos; Kotsyfakis, Michalis; Oliveira, Fabiano; Kamhawi, Shaden; Ribeiro, Jose M C; Gontijo, Nelder F; Collin, Nicolas; Valenzuela, Jesus G

    2016-01-01

    Blood-feeding insects inject potent salivary components including complement inhibitors into their host's skin to acquire a blood meal. Sand fly saliva was shown to inhibit the classical pathway of complement; however, the molecular identity of the inhibitor remains unknown. Here, we identified SALO as the classical pathway complement inhibitor. SALO, an 11?kDa protein, has no homology to proteins of any other organism apart from New World sand flies. rSALO anti-complement activity has the same chromatographic properties as the Lu. longipalpis salivary gland homogenate (SGH)counterparts and anti-rSALO antibodies blocked the classical pathway complement activity of rSALO and SGH. Both rSALO and SGH inhibited C4b deposition and cleavage of C4. rSALO, however, did not inhibit the protease activity of C1s nor the enzymatic activity of factor Xa, uPA, thrombin, kallikrein, trypsin and plasmin. Importantly, rSALO did not inhibit the alternative or the lectin pathway of complement. In conclusion our data shows that SALO is a specific classical pathway complement inhibitor present in the saliva of Lu. longipalpis. Importantly, due to its small size and specificity, SALO may offer a therapeutic alternative for complement classical pathway-mediated pathogenic effects in human diseases. PMID:26758086

  10. SALO, a novel classical pathway complement inhibitor from saliva of the sand fly Lutzomyia longipalpis

    PubMed Central

    Ferreira, Viviana P.; Fazito Vale, Vladimir; Pangburn, Michael K.; Abdeladhim, Maha; Ferreira Mendes-Sousa, Antonio; Coutinho-Abreu, Iliano V.; Rasouli, Manoochehr; Brandt, Elizabeth A.; Meneses, Claudio; Lima, Kolyvan Ferreira; Nascimento Araújo, Ricardo; Horácio Pereira, Marcos; Kotsyfakis, Michalis; Oliveira, Fabiano; Kamhawi, Shaden; Ribeiro, Jose M. C.; Gontijo, Nelder F.; Collin, Nicolas; Valenzuela, Jesus G.

    2016-01-01

    Blood-feeding insects inject potent salivary components including complement inhibitors into their host’s skin to acquire a blood meal. Sand fly saliva was shown to inhibit the classical pathway of complement; however, the molecular identity of the inhibitor remains unknown. Here, we identified SALO as the classical pathway complement inhibitor. SALO, an 11 kDa protein, has no homology to proteins of any other organism apart from New World sand flies. rSALO anti-complement activity has the same chromatographic properties as the Lu. longipalpis salivary gland homogenate (SGH)counterparts and anti-rSALO antibodies blocked the classical pathway complement activity of rSALO and SGH. Both rSALO and SGH inhibited C4b deposition and cleavage of C4. rSALO, however, did not inhibit the protease activity of C1s nor the enzymatic activity of factor Xa, uPA, thrombin, kallikrein, trypsin and plasmin. Importantly, rSALO did not inhibit the alternative or the lectin pathway of complement. In conclusion our data shows that SALO is a specific classical pathway complement inhibitor present in the saliva of Lu. longipalpis. Importantly, due to its small size and specificity, SALO may offer a therapeutic alternative for complement classical pathway-mediated pathogenic effects in human diseases. PMID:26758086

  11. Association Study of Mannose-Binding Lectin Levels and Genetic Variants in Lectin Pathway Proteins with Susceptibility to Age-Related Macular Degeneration: A Case-Control Study

    PubMed Central

    Osthoff, Michael; Dean, Melinda M.; Baird, Paul N.; Richardson, Andrea J.; Daniell, Mark; Guymer, Robyn H.; Eisen, Damon P.

    2015-01-01

    Background In age-related macular degeneration (AMD) the complement system is thought to be activated by chronic oxidative damage with genetic variants identified in the alternative pathway as susceptibility factors. However, the involvement of the lectin pathway of complement, a key mediator of oxidative damage, is controversial. This study investigated whether mannose-binding lectin (MBL) levels and genetic variants in lectin pathway proteins, are associated with the predisposition to and severity of AMD. Methods MBL levels and single nucleotide polymorphisms (SNPs) in the MBL2 and the ficolin-2 (FCN2) gene were determined in 109 patients with AMD and 109 age- and sex-matched controls. Results MBL expression levels were equally distributed in both cases (early and late AMD) and controls (p>0.05). However, there was a trend towards higher median MBL levels in cases with late AMD compared to cases with early AMD (1.0 vs. 0.4 μg/ml, p = 0.09) and MBL deficiency (<0.5 μg/ml) was encountered less frequently in the late AMD group (35% vs 56%, p = 0.03). FCN2 and MBL2 allele frequencies were similarly distributed in early and late AMD cases compared with controls (p>0.05 for all analyses) as were MBL2 genotypes. Similarly, there was no significant difference in allele frequencies in any SNPs in either the MBL2 or FCN2 gene in cases with early vs. late AMD. Conclusions SNPs of lectin pathway proteins investigated in this study were not associated with AMD or AMD severity. However, MBL levels deserve further study in a larger cohort of early vs. late AMD patients to elucidate any real effect on AMD severity. PMID:26207622

  12. A Metalloproteinase Mirolysin of Tannerella forsythia Inhibits All Pathways of the Complement System.

    PubMed

    Jusko, Monika; Potempa, Jan; Mizgalska, Danuta; Bielecka, Ewa; Ksiazek, Miroslaw; Riesbeck, Kristian; Garred, Peter; Eick, Sigrun; Blom, Anna M

    2015-09-01

    Recent reports focusing on virulence factors of periodontal pathogens implicated proteinases as major determinants of remarkable pathogenicity of these species, with special emphasis on their capacity to modulate complement activity. In particular, bacteria-mediated cleavage of C5 and subsequent release of C5a seems to be an important phenomenon in the manipulation of the local inflammatory response in periodontitis. In this study, we present mirolysin, a novel metalloproteinase secreted by Tannerella forsythia, a well-recognized pathogen strongly associated with periodontitis. Mirolysin exhibited a strong effect on all complement pathways. It inhibited the classical and lectin complement pathways due to efficient degradation of mannose-binding lectin, ficolin-2, ficolin-3, and C4, whereas inhibition of the alternative pathway was caused by degradation of C5. This specificity toward complement largely resembled the activity of a previously characterized metalloproteinase of T. forsythia, karilysin. Interestingly, mirolysin released the biologically active C5a peptide in human plasma and induced migration of neutrophils. Importantly, we demonstrated that combination of mirolysin with karilysin, as well as a cysteine proteinase of another periodontal pathogen, Prevotella intermedia, resulted in a strong synergistic effect on complement. Furthermore, mutant strains of T. forsythia, devoid of either mirolysin or karilysin, showed diminished survival in human serum, providing further evidence for the synergistic inactivation of complement by these metalloproteinases. Taken together, our findings on interactions of mirolysin with complement significantly add to the understanding of immune evasion strategies of T. forsythia and expand the knowledge on molecular mechanisms driving pathogenic events in the infected periodontium. PMID:26209620

  13. Classical and alternative complement pathway activation by pneumococci.

    PubMed Central

    Stephens, C G; Williams, R C; Reed, W P

    1977-01-01

    Sixty-two strains of Streptococcus pneumoniae were studied for their abilities to consume selected components of classical and alternative complement pathways in human sera. The classical pathway was blocked by chelating calcium with ethyleneglycol-bios (beta-aminoethyl ether)-N,N-tetraacetic acid and by removing C4. The alternative pathway was blocked by removing factor B. Each strain's activation of the two pathways was compared with its nonimmune reactivity with the Fc region of immunoglobulin G (IgG). Activation of the classical complement pathway appeared to be independent of such Fc reactivity. Highly Fc-reactive strains, however, were shown to activate the alternative pathway more effectively than did less Fc-reactive strains. Since pneumococcal activation of the alternative pathway requires non-immunospecific IgG, these findings suggest that nonimmune binding of IgG on the pneumococcal surface endows it with complement-activating properties. PMID:19357

  14. Systemic Lupus Erythematosus and Deficiencies of Early Components of the Complement Classical Pathway

    PubMed Central

    Macedo, Ana Catarina Lunz; Isaac, Lourdes

    2016-01-01

    The complement system plays an important role in the innate and acquired immune response against pathogens. It consists of more than 30 proteins found in soluble form or attached to cell membranes. Most complement proteins circulate in inactive forms and can be sequentially activated by the classical, alternative, or lectin pathways. Biological functions, such as opsonization, removal of apoptotic cells, adjuvant function, activation of B lymphocytes, degranulation of mast cells and basophils, and solubilization and clearance of immune complex and cell lysis, are dependent on complement activation. Although the activation of the complement system is important to avoid infections, it also can contribute to the inflammatory response triggered by immune complex deposition in tissues in autoimmune diseases. Paradoxically, the deficiency of early complement proteins from the classical pathway (CP) is strongly associated with development of systemic lupus erythematous (SLE) – mainly C1q deficiency (93%) and C4 deficiency (75%). The aim of this review is to focus on the deficiencies of early components of the CP (C1q, C1r, C1s, C4, and C2) proteins in SLE patients. PMID:26941740

  15. Low mannose-binding lectin complement activation function is associated with predisposition to Legionnaires' disease

    PubMed Central

    Eisen, D P; Stubbs, J; Spilsbury, D; Carnie, J; Leydon, J; Howden, B P

    2007-01-01

    Innate immune system deficiency may predispose to severe infections such as Legionnaires' disease. We have investigated the role of mannose-binding lectin (MBL) deficiency in the Melbourne Aquarium Legionnaires' disease outbreak. Serum samples from patients and controls that were exposed but shown to be uninfected from the Melbourne Aquarium Legionnaires' disease outbreak were tested for MBL function (C4 deposition) and level (mannan-binding). MBL function was lower in Legionnaires' disease cases than in age- and sex-matched uninfected, exposed controls. The frequency of MBL deficiency with C4 deposition < 0·2 U/µl was significantly higher in Legionnaires' disease cases than in controls. This also applied to Legionnaires' disease cases requiring hospital care. There was no difference in MBL mannan-binding levels between Legionnaires' disease patients and controls. There was no significant interval change in MBL function or level after a mean of 46 days. MBL complement activation functional deficiency appears to predispose to Legionnaires' disease. PMID:17425652

  16. Low mannose-binding lectin complement activation function is associated with predisposition to Legionnaires' disease.

    PubMed

    Eisen, D P; Stubbs, J; Spilsbury, D; Carnie, J; Leydon, J; Howden, B P

    2007-07-01

    Innate immune system deficiency may predispose to severe infections such as Legionnaires' disease. We have investigated the role of mannose-binding lectin (MBL) deficiency in the Melbourne Aquarium Legionnaires' disease outbreak. Serum samples from patients and controls that were exposed but shown to be uninfected from the Melbourne Aquarium Legionnaires' disease outbreak were tested for MBL function (C4 deposition) and level (mannan-binding). MBL function was lower in Legionnaires' disease cases than in age- and sex-matched uninfected, exposed controls. The frequency of MBL deficiency with C4 deposition < 0.2 U/microl was significantly higher in Legionnaires' disease cases than in controls. This also applied to Legionnaires' disease cases requiring hospital care. There was no difference in MBL mannan-binding levels between Legionnaires' disease patients and controls. There was no significant interval change in MBL function or level after a mean of 46 days. MBL complement activation functional deficiency appears to predispose to Legionnaires' disease. PMID:17425652

  17. Legionella pneumophila lipopolysaccharide activates the classical complement pathway.

    PubMed Central

    Mintz, C S; Schultz, D R; Arnold, P I; Johnson, W

    1992-01-01

    Legionella pneumophila is a gram-negative bacterium capable of entering and growing in alveolar macrophages and monocytes. Complement and complement receptors are important in the uptake of L. pneumophila by human mononuclear phagocytes. The surface molecules of L. pneumophila that activate the complement system are unknown. To identify these factors, we investigated the effects of L. pneumophila lipopolysaccharide (LPS) on the classical and alternative complement pathways of normal human serum by functional hemolytic assays. Although incubation of LPS in normal human serum at 37 degrees C resulted in the activation of both pathways, complement activation proceeded primarily through the classical pathway. Activation of the classical pathway by LPS was dependent on natural antibodies of the immunoglobulin M class that were present in various quantities in sera from different normal individuals but were absent in an immunoglobulin-deficient serum obtained from an agammaglobulinemic patient. Additional studies using sheep erythrocytes coated with LPS suggested that the antibodies recognized antigenic sites in the carbohydrate portion of LPS. The ability of LPS to interact with the complement system suggests a role for LPS in the uptake of L. pneumophila by mononuclear phagocytes. PMID:1612744

  18. Activated complement classical pathway in a murine model of oxygen-induced retinopathy

    PubMed Central

    Tao, Xue-Ying; Zheng, Shi-Jie; Lei, Bo

    2015-01-01

    AIM To investigate whether the complement system is involved in a murine model of oxygen-induced retinopathy (OIR). METHODS Forty C57BL/6J newborn mice were divided randomly into OIR group and control group. OIR was induced by exposing mice to 75%2% oxygen from postnatal 7d (P7) to P12 and then recovered in room air. For the control group, the litters were raised in room air. At the postnatal 17d (P17), gene expressions of the complement components of the classical pathway (CP), the mannose-binding lectin (MBL) pathway and the alternative pathway (AP) in the retina were determined by quantitative real-time polymerase chain reaction (RT-PCR). Retinal protein expressions of the key components in the CP were examined by Western blotting. RESULTS Whole mounted retina in the OIR mice showed area of central hypoperfusion in both superficial and deep layers and neovascular tufts in the periphery. The expressions of C1qb and C4b genes in the OIR retina were significantly higher than those of the controls. The expression of retinal complement factor B (CFB) gene in OIR mice was significantly lower than those of the controls. However, the expressions of C3 and complement factor H (CFH) genes were higher. The protein synthesis of the key components involved in the CP (C1q, C4 and C3) were also significantly higher in OIR mouse retina. Although MBL-associated serine protease 1 (MASP1) and MASP2 were detected in both the OIR and the control groups, the expressions were weak and the difference between the two groups was not significant. CONCLUSION Our data suggest that the complement system CP is activated during the pathogenesis of murine model of OIR. PMID:25709901

  19. Peptide Inhibitor of Complement C1, a Novel Suppressor of Classical Pathway Activation: Mechanistic Studies and Clinical Potential

    PubMed Central

    Sharp, Julia A.; Whitley, Pamela H.; Cunnion, Kenji M.; Krishna, Neel K.

    2014-01-01

    The classical pathway of complement plays multiple physiological roles including modulating immunological effectors initiated by adaptive immune responses and an essential homeostatic role in the clearance of damaged self-antigens. However, dysregulated classical pathway activation is associated with antibody-initiated, inflammatory diseases processes like cold agglutinin disease, acute intravascular hemolytic transfusion reaction (AIHTR), and acute/hyperacute transplantation rejection. To date, only one putative classical pathway inhibitor, C1 esterase inhibitor (C1-INH), is currently commercially available and its only approved indication is for replacement treatment in hereditary angioedema, which is predominantly a kinin pathway disease. Given the variety of disease conditions in which the classical pathway is implicated, development of therapeutics that specifically inhibits complement initiation represents a major unmet medical need. Our laboratory has identified a peptide that specifically inhibits the classical and lectin pathways of complement. In vitro studies have demonstrated that these peptide inhibitors of complement C1 (PIC1) bind to the collagen-like region of the initiator molecule of the classical pathway, C1q. PIC1 binding to C1q blocks activation of the associated serine proteases (C1sC1rC1rC1s) and subsequent downstream complement activation. Rational design optimization of PIC1 has resulted in the generation of a highly potent derivative of 15 amino acids. PIC1 inhibits classical pathway mediated complement activation in ABO incompatibility in vitro and inhibiting classical pathway activation in vivo in rats. This review will focus on the pre-clinical development of PIC1 and discuss its potential as a therapeutic in antibody-mediated classical pathway disease, specifically AIHTR. PMID:25202312

  20. Neutrophil extracellular traps can activate alternative complement pathways.

    PubMed

    Wang, H; Wang, C; Zhao, M-H; Chen, M

    2015-09-01

    The interaction between neutrophils and activation of alternative complement pathway plays a pivotal role in the pathogenesis of anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). ANCAs activate primed neutrophils to release neutrophil extracellular traps (NETs), which have recently gathered increasing attention in the development of AAV. The relationship between NETs and alternative complement pathway has not been elucidated. The current study aimed to investigate the relationship between NETs and alternative complement pathway. Detection of components of alternative complement pathway on NETs in vitro was assessed by immunostain and confocal microscopy. Complement deposition on NETs were detected after incubation with magnesium salt ethyleneglycol tetraacetic acid (Mg-EGTA)-treated human serum. After incubation of serum with supernatants enriched in ANCA-induced NETs, levels of complement components in supernatants were measured by enzyme-linked immunosorbent assay (ELISA). Complement factor B (Bb) and properdin deposited on NETs in vitro. The deposition of C3b and C5b-9 on NETs incubated with heat-inactivated normal human serum (Hi-NHS) or EGTA-treated Hi-NHS (Mg-EGTA-Hi-NHS) were significantly less than that on NETs incubated with NHS or EGTA-treated NHS (Mg-EGTA-NHS). NETs induced by ANCA could activate the alternative complement cascade in the serum. In the presence of EGTA, C3a, C5a and SC5b-9 concentration decreased from 80042??24481 ng/ml, 768??150 ng/ml, 38215??15975 ng/ml in the supernatants enriched in ANCA induced NETs to 47907??1562 ng/ml, 486??126 ng/ml, 21265??4440 ng/ml in the supernatants of DNase I-degraded NETs (P?complement pathway, and might thus participate in the pathogenesis of AAV. PMID:25963026

  1. Metacyclogenesis of Leishmania (Viannia) braziliensis in vitro: evidence that lentil lectin is a marker of complement resistance and enhanced infectivity.

    PubMed

    Almeida, M C; Cuba, C A; de Sa, C M; Pharoah, M M; Howard, K M; Miles, M A

    1993-01-01

    Axenic culture in modified Grace's medium was used to induce metacyclogenesis of Leishmania (Viannia) braziliensis in vitro. Morphological characteristics, lectin agglutination profiles, susceptibility to complement lysis, and infectivity in vivo were compared between metacyclic promastigotes and promastigotes in mid-log phase growth. Short, arrow-like promastigotes and round, oval promastigotes were defined as putative metacyclic forms on the basis of being highly motile and free swimming, with a small cell body and long flagellum. These forms increased during metacyclogenesis to > 80% whereas long-bodied, slender promastigotes and intermediate slender promastigotes declined progressively. Lentil lectin selectively agglutinated L. braziliensis after the induction of metacyclogenesis, whereas concanavalin A, wheat germ agglutinin and peanut agglutinin similarly agglutinated metacyclic promastigotes and mid-log phase promastigotes. Metacyclic promastigotes survived in 7.5%-20% human serum whereas mid-log phase promastigotes did not. Five hundred metacyclic promastigotes were highly infective to hamsters whereas 500 mid-log phase promastigotes rarely caused any lesion. Specific agglutination by lentil lectin should allow purification of metacyclic organisms for standardization of immunoprotection and challenge experiments. PMID:8236409

  2. Novel Scabies Mite Serpins Inhibit the Three Pathways of the Human Complement System

    PubMed Central

    Mika, Angela; Reynolds, Simone L.; Mohlin, Frida C.; Willis, Charlene; Swe, Pearl M.; Pickering, Darren A.; Halilovic, Vanja; Wijeyewickrema, Lakshmi C.; Pike, Robert N.; Blom, Anna M.; Kemp, David J.; Fischer, Katja

    2012-01-01

    Scabies is a parasitic infestation of the skin by the mite Sarcoptes scabiei that causes significant morbidity worldwide, in particular within socially disadvantaged populations. In order to identify mechanisms that enable the scabies mite to evade human immune defenses, we have studied molecules associated with proteolytic systems in the mite, including two novel scabies mite serine protease inhibitors (SMSs) of the serpin superfamily. Immunohistochemical studies revealed that within mite-infected human skin SMSB4 (54 kDa) and SMSB3 (47 kDa) were both localized in the mite gut and feces. Recombinant purified SMSB3 and SMSB4 did not inhibit mite serine and cysteine proteases, but did inhibit mammalian serine proteases, such as chymotrypsin, albeit inefficiently. Detailed functional analysis revealed that both serpins interfered with all three pathways of the human complement system at different stages of their activation. SMSB4 inhibited mostly the initial and progressing steps of the cascades, while SMSB3 showed the strongest effects at the C9 level in the terminal pathway. Additive effects of both serpins were shown at the C9 level in the lectin pathway. Both SMSs were able to interfere with complement factors without protease function. A range of binding assays showed direct binding between SMSB4 and seven complement proteins (C1, properdin, MBL, C4, C3, C6 and C8), while significant binding of SMSB3 occurred exclusively to complement factors without protease function (C4, C3, C8). Direct binding was observed between SMSB4 and the complement proteases C1s and C1r. However no complex formation was observed between either mite serpin and the complement serine proteases C1r, C1s, MASP-1, MASP-2 and MASP-3. No catalytic inhibition by either serpin was observed for any of these enzymes. In summary, the SMSs were acting at several levels mediating overall inhibition of the complement system and thus we propose that they may protect scabies mites from complement-mediated gut damage. PMID:22792350

  3. The alternative complement pathway regulates pathological angiogenesis in the retina.

    PubMed

    Sweigard, J Harry; Yanai, Ryoji; Gaissert, Philipp; Saint-Geniez, Magali; Kataoka, Keiko; Thanos, Aristomenis; Stahl, Gregory L; Lambris, John D; Connor, Kip M

    2014-07-01

    A defining feature in proliferative retinopathies is the formation of pathological neovessels. In these diseases, the balance between neovessel formation and regression determines blindness, making the modulation of neovessel growth highly desirable. The role of the immune system in these retinopathies is of increasing interest, but it is not completely understood. We investigated the role of the alternative complement pathway during the formation and resolution of aberrant neovascularization. We used alternative complement pathway-deficient (Fb(-/-)) mice and age- and strain-matched control mice to assess neovessel development and regression in an oxygen-induced retinopathy (OIR) mouse model. In the control mice, we found increased transcription of Fb after OIR treatment. In the Fb(-/-) mice, we prepared retinal flatmounts and identified an increased number of neovessels, peaking at postnatal day 17 (P17; P=0.001). Subjecting human umbilical vein endothelial cells (HUVECs) to low oxygen, mimicking a characteristic of neovessels, decreased the expression of the complement inhibitor Cd55. Finally, using laser capture microdissection (LCM) to isolate the neovessels after OIR, we found decreased expression of Cd55 (P=0.005). Together, our data implicate the alternative complement pathway in facilitating neovessel clearance by down-regulating the complement inhibitor Cd55 specifically on neovessels, allowing for their targeted removal while leaving the established vasculature intact.-Sweigard, J. H., Yanai, R., Gaissert, P., Saint-Geniez, M., Kataoka, K., Thanos, A., Stahl, G. L., Lambris, J. D., Connor, K. M. The alternative complement pathway regulates pathological angiogenesis in the retina. PMID:24668752

  4. The alternative complement pathway regulates pathological angiogenesis in the retina

    PubMed Central

    Sweigard, J. Harry; Yanai, Ryoji; Gaissert, Philipp; Saint-Geniez, Magali; Kataoka, Keiko; Thanos, Aristomenis; Stahl, Gregory L.; Lambris, John D.; Connor, Kip M.

    2014-01-01

    A defining feature in proliferative retinopathies is the formation of pathological neovessels. In these diseases, the balance between neovessel formation and regression determines blindness, making the modulation of neovessel growth highly desirable. The role of the immune system in these retinopathies is of increasing interest, but it is not completely understood. We investigated the role of the alternative complement pathway during the formation and resolution of aberrant neovascularization. We used alternative complement pathway–deficient (Fb−/−) mice and age- and strain-matched control mice to assess neovessel development and regression in an oxygen-induced retinopathy (OIR) mouse model. In the control mice, we found increased transcription of Fb after OIR treatment. In the Fb−/− mice, we prepared retinal flatmounts and identified an increased number of neovessels, peaking at postnatal day 17 (P17; P=0.001). Subjecting human umbilical vein endothelial cells (HUVECs) to low oxygen, mimicking a characteristic of neovessels, decreased the expression of the complement inhibitor Cd55. Finally, using laser capture microdissection (LCM) to isolate the neovessels after OIR, we found decreased expression of Cd55 (P=0.005). Together, our data implicate the alternative complement pathway in facilitating neovessel clearance by down-regulating the complement inhibitor Cd55 specifically on neovessels, allowing for their targeted removal while leaving the established vasculature intact.—Sweigard, J. H., Yanai, R., Gaissert, P., Saint-Geniez, M., Kataoka, K., Thanos, A., Stahl, G. L., Lambris, J. D., Connor, K. M. The alternative complement pathway regulates pathological angiogenesis in the retina. PMID:24668752

  5. Differential complement activation pathways promote C3b deposition on native and acetylated LDL thereby inducing lipoprotein binding to the complement receptor 1.

    PubMed

    Klop, Boudewijn; van der Pol, Pieter; van Bruggen, Robin; Wang, Yanan; de Vries, Marijke A; van Santen, Selvetta; O'Flynn, Joseph; van de Geijn, Gert-Jan M; Njo, Tjin L; Janssen, Hans W; de Man, Peter; Jukema, J Wouter; Rabelink, Ton J; Rensen, Patrick C N; van Kooten, Cees; Cabezas, Manuel Castro

    2014-12-19

    Lipoproteins can induce complement activation resulting in opsonization and binding of these complexes to complement receptors. We investigated the binding of opsonized native LDL and acetylated LDL (acLDL) to the complement receptor 1 (CR1). Binding of complement factors C3b, IgM, C1q, mannose-binding lectin (MBL), and properdin to LDL and acLDL were investigated by ELISA. Subsequent binding of opsonized LDL and acLDL to CR1 on CR1-transfected Chinese Hamster Ovarian cells (CHO-CR1) was tested by flow cytometry. Both native LDL and acLDL induced complement activation with subsequent C3b opsonization upon incubation with normal human serum. Opsonized LDL and acLDL bound to CR1. Binding to CHO-CR1 was reduced by EDTA, whereas MgEGTA only reduced the binding of opsonized LDL, but not of acLDL suggesting involvement of the alternative pathway in the binding of acLDL to CR1. In vitro incubations showed that LDL bound C1q, whereas acLDL bound to C1q, IgM, and properdin. MBL did neither bind to LDL nor to acLDL. The relevance of these findings was demonstrated by the fact that ex vivo up-regulation of CR1 on leukocytes was accompanied by a concomitant increased binding of apolipoprotein B-containing lipoproteins to leukocytes without changes in LDL-receptor expression. In conclusion, CR1 is able to bind opsonized native LDL and acLDL. Binding of LDL to CR1 is mediated via the classical pathway, whereas binding of acLDL is mediated via both the classical and alternative pathways. Binding of lipoproteins to CR1 may be of clinical relevance due to the ubiquitous cellular distribution of CR1. PMID:25349208

  6. Differential Complement Activation Pathways Promote C3b Deposition on Native and Acetylated LDL thereby Inducing Lipoprotein Binding to the Complement Receptor 1

    PubMed Central

    Klop, Boudewijn; van der Pol, Pieter; van Bruggen, Robin; Wang, Yanan; de Vries, Marijke A.; van Santen, Selvetta; O'Flynn, Joseph; van de Geijn, Gert-Jan M.; Njo, Tjin L.; Janssen, Hans W.; de Man, Peter; Jukema, J. Wouter; Rabelink, Ton J.; Rensen, Patrick C. N.; van Kooten, Cees; Cabezas, Manuel Castro

    2014-01-01

    Lipoproteins can induce complement activation resulting in opsonization and binding of these complexes to complement receptors. We investigated the binding of opsonized native LDL and acetylated LDL (acLDL) to the complement receptor 1 (CR1). Binding of complement factors C3b, IgM, C1q, mannose-binding lectin (MBL), and properdin to LDL and acLDL were investigated by ELISA. Subsequent binding of opsonized LDL and acLDL to CR1 on CR1-transfected Chinese Hamster Ovarian cells (CHO-CR1) was tested by flow cytometry. Both native LDL and acLDL induced complement activation with subsequent C3b opsonization upon incubation with normal human serum. Opsonized LDL and acLDL bound to CR1. Binding to CHO-CR1 was reduced by EDTA, whereas MgEGTA only reduced the binding of opsonized LDL, but not of acLDL suggesting involvement of the alternative pathway in the binding of acLDL to CR1. In vitro incubations showed that LDL bound C1q, whereas acLDL bound to C1q, IgM, and properdin. MBL did neither bind to LDL nor to acLDL. The relevance of these findings was demonstrated by the fact that ex vivo up-regulation of CR1 on leukocytes was accompanied by a concomitant increased binding of apolipoprotein B-containing lipoproteins to leukocytes without changes in LDL-receptor expression. In conclusion, CR1 is able to bind opsonized native LDL and acLDL. Binding of LDL to CR1 is mediated via the classical pathway, whereas binding of acLDL is mediated via both the classical and alternative pathways. Binding of lipoproteins to CR1 may be of clinical relevance due to the ubiquitous cellular distribution of CR1. PMID:25349208

  7. Eosinophil granule cationic proteins regulate the classical pathway of complement.

    PubMed Central

    Weiler, J M; Edens, R E; Bell, C S; Gleich, G J

    1995-01-01

    Major basic protein, the primary constituent of eosinophil granules, regulates the alternative and classical pathways of complement. Major basic protein and other eosinophil granule cationic proteins, which are important in mediating tissue damage in allergic disease, regulate the alternative pathway by interfering with C3b interaction with factor B to assemble an alternative pathway C3 convertase. In the present study, eosinophil peroxidase, eosinophil cationic protein and eosinophil-derived neurotoxin, as well as major basic protein, were examined for capacity to regulate the classical pathway. Eosinophil peroxidase, eosinophil cationic protein and major basic protein inhibited formation of cell-bound classical pathway C3 convertase (EAC1,4b,2a), causing 50% inhibition of complement-mediated lysis at about 0.19, 0.75 and 0.5 micrograms/10(7) cellular intermediates, respectively. Eosinophil-derived neurotoxin had no activity on this pathway of complement. The eosinophil granule proteins were examined for activity on the formation of the membrane attack complex. Major basic protein and eosinophil cationic protein had no activity on terminal lysis. In contrast, eosinophil peroxidase inhibited lysis of EAC1,4b,2a,3b,5b, but had only minimal activity on later events in complement lysis. These polycations were then examined to determine the site(s) at which they regulated the early classical pathway. Eosinophil granule polycationic proteins: (1) reduced the Zmax at all time points but had only minimal effect on the Tmax during the formation of the classical pathway C3 convertase (EAC1,4b,2a); (2) inhibited formation of EAC1,4b,2a proportional to C4 but independent of C2 concentration; (3) inhibited fluid phase formation of C1,4b,2a, as reflected by a decrease in C1-induced consumption of C2 over time; and (4) inhibited C1 activity over time without a direct effect on either C4 or C2. These observations suggest that polycations regulate the early classical pathway by interfering with C1 and may exert this activity in vivo. PMID:7750997

  8. Complement

    MedlinePLUS

    ... the suspected disease are done first. C3 and C4 are the complement components measured most often. A ... normal levels of the complement proteins C3 and C4 . Complement activity varies throughout the body. For example, ...

  9. A metalloproteinase karilysin present in the majority of Tannerella forsythia isolates inhibits all pathways of the complement system1

    PubMed Central

    Jusko, Monika; Potempa, Jan; Karim, Abdulkarim Y.; Ksiazek, Miroslaw; Riesbeck, Kristian; Garred, Peter; Eick, Sigrun; Blom, Anna M.

    2012-01-01

    Tannerella forsythia is a poorly studied pathogen despite being one of the main causes of periodontitis, which is an inflammatory disease of the supporting structures of the teeth. We found that despite being recognized by all complement pathways T. forsythia is resistant to killing by human complement, which is present at up to 70% of serum concentration in gingival crevicular fluid. Incubation of human serum with karilysin, a metalloproteinase of T. forsythia, resulted in a decrease in bactericidal activity of the serum. T. forsythia strains expressing karilysin at higher levels were more resistant than low expressing strain. Furthermore, the low expressing strain was significantly more opsonized with C3b and membrane attack complex from serum compared to the other strains. The high expressing strain was more resistant to killing in human blood. The protective effect of karilysin against serum bactericidal activity was attributable to its ability to inhibit complement at several stages. The classical and lectin complement pathways were inhibited due to the efficient degradation of mannose-binding lectin, ficolin-2, ficolin-3 and C4 by karilysin, while inhibition of the terminal pathway was caused by degradation of C5. Interestingly, karilysin was able to release biologically active C5a peptide in human plasma and induce migration of neutrophils. Importantly, we detected the karilysin gene in over 90% of gingival crevicular fluid samples containing T. forsythia obtained from patients with periodontitis. Taken together, the newly characterized karilysin appears to be an important virulence factor of T. forsythia and might have several important implications for immune evasion. PMID:22287711

  10. Activation of the alternative complement pathway by Naegleria fowleri.

    PubMed Central

    Holbrook, T W; Boackle, R J; Parker, B W; Vesely, J

    1980-01-01

    Naegleria fowleri amoebae were lysed by adult fresh human serum, and their multiplication was inhibited in culture medium supplemented with 10% fresh human serum. Heat inactivation (56 degrees C, 30 min) of serum abrogated these lytic and inhibitory effects. Absorption of human serum with amoebae failed to reduce immunoglobulin levels, and no specific antibody was detected in untreated or treated sera by counterimmunoelectrophoresis. Conversion of C3 and C3i occurred after incubation of n. fowleri with serum which had been treated with ethylene glycol-bis(beta-aminoethyl ether)-N,N-tetraacetic acid, indicating activation of complement via the alternative pathway. Images Fig. 1 PMID:7439979

  11. An amphioxus gC1q protein binds human IgG and initiates the classical pathway: Implications for a C1q-mediated complement system in the basal chordate.

    PubMed

    Gao, Zhan; Li, Mengyang; Ma, Jie; Zhang, Shicui

    2014-12-01

    The origin of the classical complement pathway remains open during chordate evolution. A C1q-like member, BjC1q, was identified in the basal chordate amphioxus. It is predominantly expressed in the hepatic caecum, hindgut, and notochord, and is significantly upregulated following challenge with bacteria or lipoteichoic acid and LPS. Recombinant BjC1q and its globular head domain specifically interact with lipoteichoic acid and LPS, but BjC1q displays little lectin activity. Moreover, rBjC1q can assemble to form the high molecular weight oligomers necessary for binding to proteases C1r/C1s and for complement activation, and binds human C1r/C1s/mannan-binding lectin-associated serine protease-2 as well as amphioxus serine proteases involved in the cleavage of C4/C2, and C3 activation. Importantly, rBjC1q binds with human IgG as well as an amphioxus Ig domain containing protein, resulting in the activation of the classical complement pathway. This is the first report showing that a C1q-like protein in invertebrates is able to initiate classical pathway, raising the possibility that amphioxus possesses a C1q-mediated complement system. It also suggests a new scenario for the emergence of the classical complement pathway, in contrast to the proposal that the lectin pathway evolved into the classical pathway. PMID:25174509

  12. Characterization of mannose binding lectin from channel catfish Ictalurus punctatus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Mannose-binding lectin (MBL) is an important component of innate immunity capable of activating the lectin pathway of the complement system. A MBL gene was isolated from channel catfish (Ictalurus punctatus). The deduced protein contains a canonical collagen-like domain, a carbohydrate recognition d...

  13. Identification of Novel Pathways in Plant Lectin-Induced Cancer Cell Apoptosis.

    PubMed

    Shi, Zheng; Sun, Rong; Yu, Tian; Liu, Rong; Cheng, Li-Jia; Bao, Jin-Ku; Zou, Liang; Tang, Yong

    2016-01-01

    Plant lectins have been investigated to elucidate their complicated mechanisms due to their remarkable anticancer activities. Although plant lectins seems promising as a potential anticancer agent for further preclinical and clinical uses, further research is still urgently needed and should include more focus on molecular mechanisms. Herein, a Nave Bayesian model was developed to predict the protein-protein interaction (PPI), and thus construct the global human PPI network. Moreover, multiple sources of biological data, such as smallest shared biological process (SSBP), domain-domain interaction (DDI), gene co-expression profiles and cross-species interolog mapping were integrated to build the core apoptotic PPI network. In addition, we further modified it into a plant lectin-induced apoptotic cell death context. Then, we identified 22 apoptotic hub proteins in mesothelioma cells according to their different microarray expressions. Subsequently, we used combinational methods to predict microRNAs (miRNAs) which could negatively regulate the abovementioned hub proteins. Together, we demonstrated the ability of our Nave Bayesian model-based network for identifying novel plant lectin-treated cancer cell apoptotic pathways. These findings may provide new clues concerning plant lectins as potential apoptotic inducers for cancer drug discovery. PMID:26867193

  14. Identification of Novel Pathways in Plant Lectin-Induced Cancer Cell Apoptosis

    PubMed Central

    Shi, Zheng; Sun, Rong; Yu, Tian; Liu, Rong; Cheng, Li-Jia; Bao, Jin-Ku; Zou, Liang; Tang, Yong

    2016-01-01

    Plant lectins have been investigated to elucidate their complicated mechanisms due to their remarkable anticancer activities. Although plant lectins seems promising as a potential anticancer agent for further preclinical and clinical uses, further research is still urgently needed and should include more focus on molecular mechanisms. Herein, a Naïve Bayesian model was developed to predict the protein-protein interaction (PPI), and thus construct the global human PPI network. Moreover, multiple sources of biological data, such as smallest shared biological process (SSBP), domain-domain interaction (DDI), gene co-expression profiles and cross-species interolog mapping were integrated to build the core apoptotic PPI network. In addition, we further modified it into a plant lectin-induced apoptotic cell death context. Then, we identified 22 apoptotic hub proteins in mesothelioma cells according to their different microarray expressions. Subsequently, we used combinational methods to predict microRNAs (miRNAs) which could negatively regulate the abovementioned hub proteins. Together, we demonstrated the ability of our Naïve Bayesian model-based network for identifying novel plant lectin-treated cancer cell apoptotic pathways. These findings may provide new clues concerning plant lectins as potential apoptotic inducers for cancer drug discovery. PMID:26867193

  15. Complementation.

    PubMed Central

    Yook, Karen

    2005-01-01

    Mutations in many genes can result in a similar phenotype. Finding a number of mutants with the same phenotype tells you little about how many genes you are dealing with, and how mutable those genes are until you can assign those mutations to genetic loci. The genetic assay for gene assignment is called the complementation test. The simplicity and robustness of this test makes it a fundamental genetic tool for gene assignment. However, there are occasional unexpected outcomes from this test that bear explanation. This chapter reviews the complementation test and its various outcomes, highlighting relatively rare but nonetheless interesting exceptions such as intragenic complementation and non-allelic non-complementation. PMID:18023121

  16. The mannan-binding lectin pathway and lung disease in cystic fibrosis--disfunction of mannan-binding lectin-associated serine protease 2 (MASP-2) may be a major modifier.

    PubMed

    Olesen, H V; Jensenius, J C; Steffensen, R; Thiel, S; Schitz, P O

    2006-12-01

    The lectin pathway of complement activation is initiated by mannan-binding lectin (MBL) or the ficolins through the common MBL-associated serine protease-2 (MASP-2). Deficiency of MBL has been associated with poorer outcome in cystic fibrosis (CF). We investigated the MBL pathway further by analysis of the MASP-2 deficiency mutation (D105G) as well as MBL-2 genotypes. Concentrations and genotypes of MASP-2 and MBL in 109 CF patients were correlated to lung function and chronic infections. We describe the first CF patient homozygous for the mutation, a girl with extremely severe lung disease with no other precipitating factors. We suspect total MASP-2 dysfunction to be a major modifier of CF lung disease. However, heterozygosity for the D105G mutation of MASP-2 had no correlation to MBL pathway function or poor lung function. Lung function was higher in the MBL deficiency determining genotypes (XA/YO+YO/YO) than in the other genotypes. PMID:17045845

  17. Regulation of the alternative pathway of complement modulates injury and immunity in a chronic model of dextran sulphate sodium-induced colitis.

    PubMed

    Elvington, M; Schepp-Berglind, J; Tomlinson, S

    2015-03-01

    The role of complement in inflammatory bowel disease (IBD) has been studied primarily using acute models, and it is unclear how complement affects processes in more relevant chronic models of IBD in which modulation of adaptive immunity and development of fibrosis have pathogenic roles. Using mice deficient in C1q/mannose-binding lectin (MBL) or C3, we demonstrated an important role for these opsonins and/or the classical pathway C3 convertase in providing protection against mucosal injury and infection in a model of chronic dextran sulphate sodium (DSS)-induced colitis. In contrast, deficiency of the alternative pathway (fB(-/-) mice) had significantly less impact on injury profiles. Consequently, the effect of a targeted inhibitor of the alternative pathway was investigated in a therapeutic protocol. Following the establishment of colitis, mice were treated with CR2-fH during subsequent periods of DSS treatment and acute injury (modelling relapse). CR2-fH significantly reduced complement activation, inflammation and injury in the colon, and additionally reduced fibrosis. Alternative pathway inhibition also altered the immune response in the chronic state in terms of reducing numbers of B cells, macrophages and mature dendritic cells in the lamina propria. This study indicates an important role for the alternative pathway of complement in the pathogenesis and the shaping of an immune response in chronic DSS-induced colitis, and supports further investigation into the use of targeted alternative pathway inhibition for the treatment of IBD. PMID:25293413

  18. Time-course analysis of C3a and C5a quantifies the coupling between the upper and terminal Complement pathways in vitro.

    PubMed

    Morad, Hassan O J; Belete, Samuel C; Read, Thomas; Shaw, Andrew M

    2015-12-01

    An in vitro zymosan-activation of the Complement system, through the lectin and alternative pathways, was performed in pooled human serum over a 24h time-course. Activation was quantitatively monitored by measuring the concentration of the upper Complement pathway fragment, C3a and the terminal pathway fragment, C5a. Upper Complement showed a maximum activation of 39% and the time-to-maximum activation reduced 8-fold, as a highly non-linear function of the zymosan dose. The C3a:C5a molar ratio rose to a maximum of 1100:1, before terminal pathway activation was initiated; indicating a flux threshold. This threshold appears to be exceeded once more than 31% of C3 molecules are activated. Above this threshold, significant activation of terminal pathway was observed; reducing the molar ratio to 17:1. The C5a/C3a molar ratio was used to determine the terminal pathway activation relative to total Complement activation and ranged from 0.1-0.8%. This depicts upper Complement activation to be 49-fold larger than terminal activation, a figure consistent with the observed density of the membrane attack complex in the membrane of cells. Our results thus indicate that the relative activity of opsonisation is ~50-fold greater than membrane attack complex formation, in vitro, in the pooled serum phenotype. The results suggest a potential clinical application, where an in vitro analysis of a patient on admission, or prior to a surgical procedure, would indicate their upper Complement activation capacity, with activation of C3 measured thereafter, or post-operatively. A patient with an exhausted upper Complement capacity may be vulnerable to infections and complications, such as sepsis. PMID:26391915

  19. Molecular defects in the mannose binding lectin pathway in dermatological disease: Case report and literature review

    PubMed Central

    2010-01-01

    Mannose-binding lectin (MBL) and the Mannose-binding lectin-associated serine proteases (MASPs) are an essential aspect of innate immune responses that probably play an important but understudied role in cutaneous function. The MBL-MASP pathway appears to exert its primary role by assisting in the clearance of apoptotic skin cells (thus preventing accumulation and a subsequent autoimmune response) and promoting opsonophagocytosis of invading pathogens, limiting their dissemination. Deficiencies of the pathway have been described and are associated with infectious, autoimmune and vascular complications. However, the role of this pathway in dermatological disease is essentially unexplored. We describe 6 patients presenting with recurrent inflammatory and/or infectious skin conditions who also demonstrated severely low MBL levels. One patient also had a defect in the MASP2 gene. Genotype analysis revealed specific point mutations in the MBL2 promoter in all 6 patients and a variant MASP-2 gene in one patient. Five patients presented recurrent pustular skin infections (cellulitis, folliculitis and cutaneous abscess). A case of Grover's disease and one forme fruste of Behcet's syndrome (orogenital ulcers) were also observed. The patients responded to antimicrobial therapy, although in some, recurrence of infection was the rule. It appears that MBL deficiency may contribute to recurrent skin infections and to certain forms of inflammatory skin disease. The mechanisms may relate to the role of this pathway in innate immunity, removal of apoptotic cells and in immune complexes. Further study of MBL pathway defects in dermatological disease is required. PMID:20338057

  20. Activation of the human terminal complement pathway in atherosclerosis.

    PubMed

    Niculescu, F; Rus, H G; Vlaicu, R

    1987-11-01

    The presence of the terminal C5b-9 complement complex in tissues indicates that complement activation has occurred in situ with subsequent membrane damage, tissue injury, and inflammatory response mediation. The terminal C5b-9 neoantigens of the complement system, S protein C3c, C3d, and apolipoprotein B deposits were localized in 20 aortic fibrous plaques, 12 aortic intimal thickenings, 8 aortic fatty streak intimae, 14 coronary fibrous plaques, 5 coronary intimal thickenings, and 8 femoral fibrous plaques, using an indirect and double-staining immunoperoxidase technique. The specific granular deposits were present from the early to the advanced stages of atherosclerosis in relation to the degree of fibrosis and necrosis. The different double-staining localization of C5b-9 and S protein may suggest local assembly of the complex as a consequence of complement activation and may sustain its role in the chronic progression of atherosclerosis. PMID:2444373

  1. Complement pathway biomarkers and age-related macular degeneration.

    PubMed

    Gemenetzi, M; Lotery, A J

    2016-01-01

    In the age-related macular degeneration (AMD) 'inflammation model', local inflammation plus complement activation contributes to the pathogenesis and progression of the disease. Multiple genetic associations have now been established correlating the risk of development or progression of AMD. Stratifying patients by their AMD genetic profile may facilitate future AMD therapeutic trials resulting in meaningful clinical trial end points with smaller sample sizes and study duration. PMID:26493033

  2. C-Type Lectin-Like Receptors of the Dectin-1 Cluster: Ligands and Signaling Pathways

    PubMed Central

    Plato, Anthony; Willment, Janet A.

    2013-01-01

    Innate immunity is constructed around genetically encoded receptors that survey the intracellular and extracellular environments for signs of invading microorganisms. These receptors recognise the invader and through complex intracellular networks of molecular signaling, they destroy the threat whilst instructing effective adaptive immune responses. Many of these receptors, like the Toll-like receptors in particular, are well-known for their ability to mediate downstream responses upon recognition of exogenous or endogenous ligands; however, the emerging family known as the C-type lectin-like receptors contains many members that have a huge impact on immune and homeostatic regulation. Of particular interest here are the C-type lectin-like receptors that make up the Dectin-1 cluster and their intracellular signaling motifs that mediate their functions. In this review, we aim to draw together current knowledge of ligands, motifs and signaling pathways, present downstream of Dectin-1 cluster receptors, and discuss how these dictate their role within biological systems. PMID:23570314

  3. Clinical hypothermia temperatures increase complement activation and cell destruction via the classical pathway

    PubMed Central

    2014-01-01

    Background Therapeutic hypothermia is a treatment modality that is increasingly used to improve clinical neurological outcomes for ischemia-reperfusion injury-mediated diseases. Antibody-initiated classical complement pathway activation has been shown to contribute to ischemia-reperfusion injury in multiple disease processes. However, how therapeutic hypothermia affects complement activation is unknown. Our goal was to measure the independent effect of temperature on complement activation, and more specifically, examine the relationship between clinical hypothermia temperatures (3133C), and complement activation. Methods Antibody-sensitized erythrocytes were used to assay complement activation at temperatures ranging from 0-41C. Individual complement pathway components were assayed by ELISA, Western blot, and quantitative dot blot. Peptide Inhibitor of complement C1 (PIC1) was used to specifically inhibit activation of C1. Results Antibody-initiated complement activation resulting in eukaryotic cell lysis was increased by 2-fold at 31C compared with 37C. Antibody-initiated complement activation in human serum increased as temperature decreased from 37C until dramatically decreasing at 13C. Quantitation of individual complement components showed significantly increased activation of C4, C3, and C5 at clinical hypothermia temperatures. In contrast, C1s activation by heat-aggregated IgG decreased at therapeutic hypothermia temperatures consistent with decreased enzymatic activity at lower temperatures. However, C1q binding to antibody-coated erythrocytes increased at lower temperatures, suggesting that increased classical complement pathway activation is mediated by increased C1 binding at therapeutic hypothermia temperatures. PIC1 inhibited hypothermia-enhanced complement-mediated cell lysis at 31C by up to 60% (P?=?0.001) in a dose dependent manner. Conclusions In summary, therapeutic hypothermia temperatures increased antibody-initiated complement activation and eukaryotic cell destruction suggesting that the benefits of therapeutic hypothermia may be mediated via other mechanisms. Antibody-initiated complement activation has been shown to contribute to ischemia-reperfusion injury in several animal models, suggesting that for diseases with this mechanism hypothermia-enhanced complement activation may partially attenuate the benefits of therapeutic hypothermia. PMID:24962100

  4. Functional Characterization of a Ficolin-mediated Complement Pathway in Amphioxus*

    PubMed Central

    Huang, Huiqing; Huang, Shengfeng; Yu, Yingcai; Yuan, Shaochun; Li, Rui; Wang, Xin; Zhao, Hongchen; Yu, Yanhong; Li, Jun; Yang, Manyi; Xu, Liqun; Chen, Shangwu; Xu, Anlong

    2011-01-01

    The ficolin-mediated complement pathway plays an important role in vertebrate immunity, but it is not clear whether this pathway exists in invertebrates. Here we identified homologs of ficolin pathway components from the cephalochordate amphioxus and investigated whether they had been co-opted into a functional ficolin pathway. Four of these homologs, ficolin FCN1, serine protease MASP1 and MASP3, and complement component C3, were highly expressed in mucosal tissues and gonads, and were significantly up-regulated following bacterial infection. Recombinant FCN1 could induce hemagglutination, discriminate among sugar components, and specifically recognize and aggregate several bacteria (especially Gram-positive strains) without showing bactericidal activity. This suggested that FCN1 is a dedicated pattern-recognition receptor. Recombinant serine protease MASP1/3 formed complexes with recombinant FCN1 and facilitated the activation of native C3 protein in amphioxus humoral fluid, in which C3 acted as an immune effector. We conclude that amphioxus have developed a functional ficolin-complement pathway. Because ficolin pathway components have not been reported in non-chordate species, our findings supported the idea that this pathway may represent a chordate-specific innovation in the evolution of the complement system. PMID:21832079

  5. Activation of the alternate complement pathway by peptidoglycan from streptococcal cell wall.

    PubMed

    Greenblatt, J; Boackle, R J; Schwab, J H

    1978-01-01

    Activation of the alternate complement pathway in human serum by several bacterial components was compared. Peptidoglycan from group A streptococcal cell walls was the most active material, on a weight basis, followed by cell walls, protoplast membranes, and whole cells. The group-specific carbohydrate was inactive. Treatment of peptidoglycan with low concentrations of lysozyme or short periods of sonic treatment enhanced complement activation. High concentrations of lysozyme or extended sonic treatment of peptidoglycan destroyed or greatly reduced the capacity to activate complement. Lysozyme treatment of group A streptococcal cell walls or lipopolysaccharide had no measurable effect. Activation of the alternate complement pathway by group D streptococcal cell walls was destroyed by lysozyme. Activity of peptidoglycan was not inhibited by N-acetyl glucosamine, N-acetyl muramic acid, or D-alanine-D-alanine. Conversion of C3 and factored B by peptidoglycan was shown to occur by immunoelectrophoresis and crossed immunoelectrophoresis. PMID:415005

  6. The complement system in human cardiometabolic disease.

    PubMed

    Hertle, E; Stehouwer, C D A; van Greevenbroek, M M J

    2014-10-01

    The complement system has been implicated in obesity, fatty liver, diabetes and cardiovascular disease (CVD). Complement factors are produced in adipose tissue and appear to be involved in adipose tissue metabolism and local inflammation. Thereby complement links adipose tissue inflammation to systemic metabolic derangements, such as low-grade inflammation, insulin resistance and dyslipidaemia. Furthermore, complement has been implicated in pathophysiological mechanisms of diet- and alcohol induced liver damage, hyperglycaemia, endothelial dysfunction, atherosclerosis and fibrinolysis. In this review, we summarize current evidence on the role of the complement system in several processes of human cardiometabolic disease. C3 is the central component in complement activation, and has most widely been studied in humans. C3 concentrations are associated with insulin resistance, liver dysfunction, risk of the metabolic syndrome, type 2 diabetes and CVD. C3 can be activated by the classical, the lectin and the alternative pathway of complement activation; and downstream activation of C3 activates the terminal pathway. Complement may also be activated via extrinsic proteases of the coagulation, fibrinolysis and the kinin systems. Studies on the different complement activation pathways in human cardiometabolic disease are limited, but available evidence suggests that they may have distinct roles in processes underlying cardiometabolic disease. The lectin pathway appeared beneficial in some studies on type 2 diabetes and CVD, while factors of the classical and the alternative pathway were related to unfavourable cardiometabolic traits. The terminal complement pathway was also implicated in insulin resistance and liver disease, and appears to have a prominent role in acute and advanced CVD. The available human data suggest a complex and potentially causal role for the complement system in human cardiometabolic disease. Further, preferably longitudinal studies are needed to disentangle which aspects of the complement system and complement activation affect the different processes in human cardiometabolic disease. PMID:25017306

  7. Current Understanding of the Role of Complement in IgA Nephropathy.

    PubMed

    Maillard, Nicolas; Wyatt, Robert J; Julian, Bruce A; Kiryluk, Krzysztof; Gharavi, Ali; Fremeaux-Bacchi, Veronique; Novak, Jan

    2015-07-01

    Complement activation has a role in the pathogenesis of IgA nephropathy, an autoimmune disease mediated by pathogenic immune complexes consisting of galactose-deficient IgA1 bound by antiglycan antibodies. Of three complement-activation pathways, the alternative and lectin pathways are involved in IgA nephropathy. IgA1 can activate both pathways in vitro, and pathway components are present in the mesangial immunodeposits, including properdin and factor H in the alternative pathway and mannan-binding lectin, mannan-binding lectin-associated serine proteases 1 and 2, and C4d in the lectin pathway. Genome-wide association studies identified deletion of complement factor H-related genes 1 and 3 as protective against the disease. Because the corresponding gene products compete with factor H in the regulation of the alternative pathway, it has been hypothesized that the absence of these genes could lead to more potent inhibition of complement by factor H. Complement activation can take place directly on IgA1-containing immune complexes in circulation and/or after their deposition in the mesangium. Notably, complement factors and their fragments may serve as biomarkers of IgA nephropathy in serum, urine, or renal tissue. A better understanding of the role of complement in IgA nephropathy may provide potential targets and rationale for development of complement-targeting therapy of the disease. PMID:25694468

  8. Alternative Complement Pathway Deficiency Ameliorates Chronic Smoke-Induced Functional and Morphological Ocular Injury

    PubMed Central

    Woodell, Alex; Coughlin, Beth; Kunchithapautham, Kannan; Casey, Sarah; Williamson, Tucker; Ferrell, W. Drew; Atkinson, Carl; Jones, Bryan W.; Rohrer, Bärbel

    2013-01-01

    Background Age-related macular degeneration (AMD), a complex disease involving genetic variants and environmental insults, is among the leading causes of blindness in Western populations. Genetic and histologic evidence implicate the complement system in AMD pathogenesis; and smoking is the major environmental risk factor associated with increased disease risk. Although previous studies have demonstrated that cigarette smoke exposure (CE) causes retinal pigment epithelium (RPE) defects in mice, and smoking leads to complement activation in patients, it is unknown whether complement activation is causative in the development of CE pathology; and if so, which complement pathway is required. Methods Mice were exposed to cigarette smoke or clean, filtered air for 6 months. The effects of CE were analyzed in wildtype (WT) mice or mice without a functional complement alternative pathway (AP; CFB−/−) using molecular, histological, electrophysiological, and behavioral outcomes. Results CE in WT mice exhibited a significant reduction in function of both rods and cones as determined by electroretinography and contrast sensitivity measurements, concomitant with a thinning of the nuclear layers as measured by SD-OCT imaging and histology. Gene expression analyses suggested that alterations in both photoreceptors and RPE/choroid might contribute to the observed loss of function, and visualization of complement C3d deposition implies the RPE/Bruch's membrane (BrM) complex as the target of AP activity. RPE/BrM alterations include an increase in mitochondrial size concomitant with an apical shift in mitochondrial distribution within the RPE and a thickening of BrM. CFB−/− mice were protected from developing these CE-mediated alterations. Conclusions Taken together, these findings provide clear evidence that ocular pathology generated in CE mice is dependent on complement activation and requires the AP. Identifying animal models with RPE/BrM damage and verifying which aspects of pathology are dependent upon complement activation is essential for developing novel complement-based treatment approaches for the treatment of AMD. PMID:23825688

  9. Glomeruli of Dense Deposit Disease contain components of the alternative and terminal complement pathway

    PubMed Central

    Sethi, Sanjeev; Gamez, Jeffrey D.; Vrana, Julie A.; Theis, Jason D.; Bergen, H. Robert; Zipfel, Peter F.; Dogan, Ahmet; Smith, Richard J. H.

    2009-01-01

    Dense Deposit Disease (DDD), or membranoproliferative glomerulonephritis type II, is a rare renal disease characterized by dense deposits in the mesangium and along the glomerular basement membranes that can be seen by electron microscopy. Although these deposits contain complement factor C3, as determined by immunofluorescence microscopy, their precise composition remains unknown. To address this question, we used mass spectrometry to identify the proteins in laser microdissected glomeruli isolated from paraffin-embedded tissue of eight confirmed cases of DDD. Compared to glomeruli from five control patients, we found that all of the glomeruli from patients with DDD contain components of the alternative pathway and terminal complement complex. Factor C9 was uniformly present as well as the two fluid-phase regulators of terminal complement complex clusterin and vitronectin. In contrast, in nine patients with immune complexmediated membranoproliferative glomerulonephritis, glomerular samples contained mainly immunoglobulins and complement factors C3 and C4. Our study shows that in addition to fluid-phase dysregulation of the alternative pathway, soluble components of the terminal complement complex contribute to glomerular lesions found in DDD. PMID:19177158

  10. Activation of the alternative complement pathway in canine normal serum by Paracoccidioides brasiliensis

    PubMed Central

    Bianchini, A.A.C.; Petroni, T.F.; Fedatto, P.F.; Bianchini, R.R.; Venancio, E.J.; Itano, E.N.; Ono, M.A.

    2009-01-01

    The dimorphic fungus Paracoccidioides brasiliensis is the etiological agent of paracoccidioidomycosis, a human granulomatous disease. Recently the first case of natural disease in dogs was reported. The complement system is an important effector component of humoral immunity against infectious agents. Therefore, the aim of this study was to evaluate the activation of the dog alternative complement pathway by P. brasiliensis. Initially, the ability of erythrocytes of guinea pig, rabbit, sheep, chicken and swine to activate the dog alternative pathway was evaluated. The guinea pig erythrocytes showed the greatest capacity to activate dog alternative pathway. The alternative (AH50) hemolytic activity was evaluated in 27 serum samples from healthy dogs and the mean values were 87.2 AH50/ml. No significant differences were observed in relation to sex and age. The alternative pathway activation by P. brasiliensis was higher in serum samples from adult dogs when compared to puppies and aged dogs (p ? 0.05). This is the first report of dog alternative complement pathway activation by P. brasiliensis and suggests that it may play a protective role in canine paracoccidioidomycosis. PMID:24031350

  11. Complement activation by the alternative pathway and macrophage enzyme secretion in the pathogenesis of chronic inflammation.

    PubMed Central

    Schorlemmer, H U; Bitter-Suermann, D; Allison, A C

    1977-01-01

    A number of stimuli known to induce acid hydrolase secretion from cultured macrophages were examined for their ability to activate C3 via the alternative pathway of the complement system. Loss of haemolytically active C3 was checked in normal and C4-deficient guinea-pig serum. For comparison the interactions of cultured macrophages with other agents well known as potent activators of the alternative pathway of the complement system have been investigated. As judged by their activity in these assays, group A streptococcal cell walls, different carrageenan preparations, dental plaque and Actinomyces viscosus were all capable of initiating the alternative pathway but differed with respect to their potency and their ability to inhibit C3 turnover at high concentrations. Zymosan, some carrageenans, polyanethol sulphonate, and Corynebacterium parvum all induce the release of hydrolytic enzymes from macrophages in culture, even in the absence of serum in the medium. The release is time- and dose-dependent and is not associated with loss of the cytoplasmic enzyme lactate dehydrogenase or any other sign of cell death. The parallelism between the capacity of several agents to activate the complement system via the alternative pathway and to induce inflammatory responses in vivo and selective lysosoma enzyme secretion from cultures of macrophages is discussed. PMID:328387

  12. Complement Pathway is Frequently Altered in Endometriosis and Endometriosis-Associated Ovarian Cancer

    PubMed Central

    Suryawanshi, Swati; Huang, Xin; Elishaev, Esther; Budiu, Raluca A.; Zhang, Lixin; Kim, SungHwan; Donnellan, Nicole; Mantia-Smaldone, Gina; Ma, Tianzhou; Tseng, George; Lee, Ted; Mansuria, Suketu; Edwards, Robert; Vlad, Anda M.

    2014-01-01

    Purpose Mechanisms of immune dysregulation associated with advanced tumors are relatively well understood. Much less is known about the role of immune effectors against cancer precursor lesions. Endometrioid and clear cell ovarian tumors partly derive from endometriosis, a commonly diagnosed chronic inflammatory disease. We performed here a comprehensive immune gene expression analysis of pelvic inflammation in endometriosis and endometriosis-associated ovarian cancer (EAOC). Experimental design RNA was extracted from 120 paraffin tissue blocks comprising of normal endometrium (n=32), benign endometriosis (n=30), atypical endometriosis (n=15) and EAOC (n=43). Serous tumors (n=15) were included as non-endometriosis associated controls. The immune microenvironment was profiled using Nanostring and the nCounter GX Human Immunology Kit, comprising probes for a total of 511 immune genes. Results One third of the endometriosis patients revealed a tumor-like inflammation profile, suggesting that cancerlike immune signatures may develop earlier, in patients classified as clinically benign. Gene expression analyses revealed the complement pathway as most prominently involved in both endometriosis and EAOC. Complement proteins are abundantly present in epithelial cells in both benign and malignant lesions. Mechanistic studies in ovarian surface epithelial (OSE) cells from mice with conditional (Cre-loxP) mutations show intrinsic production of complement in epithelia and demonstrate an early link between Kras- and Pten-driven pathways and complement upregulation. Downregulation of complement in these cells interferes with cell proliferation. Conclusions These findings reveal new characteristics of inflammation in precursor lesions and point to previously unknown roles of complement in endometriosis and EAOC. PMID:25294912

  13. Alternative Pathway of Complement in Children with Diarrhea-Associated Hemolytic Uremic Syndrome

    PubMed Central

    Thurman, Joshua M.; Marians, Russell; Emlen, Woodruff; Wood, Susan; Smith, Christopher; Akana, Hillary; Holers, V. Michael; Lesser, Martin; Kline, Myriam; Hoffman, Cathy; Christen, Erica

    2009-01-01

    Background and objectives: Diarrhea-associated hemolytic uremic syndrome (D+HUS) is a common cause of acute kidney injury in children. Mutations in alternative pathway (AP) complement regulatory proteins have been identified in severe cases of thrombotic microangiopathy, but the role of the AP in D+HUS has not been studied. Therefore, we determined whether plasma levels of markers of activation of the AP are increased in D+HUS and are biomarkers of the severity of renal injury that predict the need for dialysis. Design, setting, participants, & measurements: Patients were randomly selected from among participants in the HUS-SYNSORB Pk trial. Plasma samples were collected on days 1, 4, 7, and 10 after enrollment and day 28 after discharge from the hospital. Levels of two complement pathway products, Bb and SC5b-9, were determined by ELISA. Results: Seventeen children (6 boys and 11 girls; age, 5.4 ± 3.5 yr) were studied. Eight (47%) required dialysis support, and two had serious extrarenal events. On the day of enrollment, plasma levels of Bb and SC5b-9 were significantly increased in all patients compared with healthy controls (P < 0.01). The elevated concentrations normalized by day 28 after discharge. Circulating levels of complement pathway fragments did not correlate with severity of renal injury or occurrence of complications. Conclusions: Patients with acute-onset D+HUS manifest activation of the AP of complement that is temporally related to the onset of disease and that resolves within 1 mo. Therapies to inhibit the AP of complement may be useful in attenuating the severity of renal injury and extrarenal complications. PMID:19820137

  14. The role of complement in membranous nephropathy

    PubMed Central

    Ma, Hong; Sandor, Dana G.; Beck, Laurence H.

    2013-01-01

    Membranous nephropathy (MN) describes a histopathological pattern of injury marked by glomerular subepithelial immune deposits and collectively represents one of the most common causes of adult nephrotic syndrome. Studies in Heymann nephritis, an experimental model of MN, have established a paradigm in which these deposits locally activate complement to cause podocyte injury, culminating in cytoskeletal reorganization, loss of slit diaphragms, and proteinuria. There is much circumstantial evidence for a prominent role of complement in human MN, as C3 and C5b-9 are consistently found within immune deposits. Secondary MN often exhibits the additional presence of C1q, implicating the classical pathway of complement activation. Primary MN, however, is IgG4-predominant and IgG4 is considered incapable of binding C1q and activating the complement pathway. Recent studies have identified the M-type phospholipase A2 receptor (PLA2R) as the major target antigen in primary MN. Early evidence hints that IgG4 anti-PLA2R autoantibodies can bind mannan-binding lectin and activate the lectin complement pathway. The identification of anti-PLA2R antibodies as likely participants in the pathogenesis of disease will allow focused investigation into the role of complement in MN. Definitive therapy for MN is immunosuppression, although future therapeutic agents that specifically target complement activation may represent an effective temporizing measure to forestall further glomerular injury. PMID:24161038

  15. Differential ability to resist to complement lysis and invade host cells mediated by MBL in R4 and 860 strains of Trypanosoma cruzi.

    PubMed

    Evans-Osses, Ingrid; Mojoli, Andres; Beltrame, Marcia Holsbach; da Costa, Denise Endo; DaRocha, Wanderson Duarte; Velavan, Thirumalaisamy P; de Messias-Reason, Iara; Ramirez, Marcel Ivan

    2014-03-18

    To produce an infection Trypanosoma cruzi must evade lysis by the complement system. During early stages of infection, the lectin pathway plays an important role in host defense and can be activated by binding of mannan-binding lectin (MBL) to carbohydrates on the surface of pathogens. We hypothesized that MBL has a dual role during parasite-host cell interaction as lectin complement pathway activator and as binding molecule to invade the host cell. We used two polarized strains of T. cruzi, R4 (susceptible) and 860 (resistant) strains, to investigate the role of MBL in complement-mediated lysis. Interestingly R4, but not 860 metacyclic strain, markedly increases the invasion of host cells, suggesting that MBL drives the invasion process while the parasite deactivates the Lectin complement pathway. PMID:24560788

  16. Exogenous expression of marine lectins DlFBL and SpRBL induces cancer cell apoptosis possibly through PRMT5-E2F-1 pathway

    PubMed Central

    Wu, Liqin; Yang, Xinyan; Duan, Xuemei; Cui, Lianzhen; Li, Gongchu

    2014-01-01

    Lectins are widely existed in marine bioresources, and some purified marine lectins were found toxic to cancer cells. In this report, genes encoding Dicentrarchus labrax fucose-binding lectin (DlFBL) and Strongylocentrotus purpuratus rhamnose-binding lectin (SpRBL) were inserted into an adenovirus vector to form Ad.FLAG-DlFBL and Ad.FLAG-SpRBL, which elicited significant in vitro suppressive effect on a variety of cancer cells. Anti-apoptosis factors Bcl-2 and XIAP were determined to be downregulated by Ad.FLAG-DlFBL and Ad.FLAG-SpRBL. Subcellular localization studies showed that DlFBL but not SpRBL widely distributed in membrane systems. Both DlFBL and SpRBL were shown associated with protein arginine methyltransferase 5 (PRMT5), and PRMT5-E2F-1 pathway was suggested to be responsible for the DlFBL and SpRBL induced apoptosis. Further investigations revealed that PRMT5 acted as a common binding target for various exogenous lectin and non-lectin proteins, suggesting a role of PRMT5 as a barrier for foreign gene invasion. The cellular response to exogenous lectins may provide insights into a novel way for cancer gene therapy. PMID:24675921

  17. A single-CRD C-type lectin from oyster Crassostrea gigas mediates immune recognition and pathogen elimination with a potential role in the activation of complement system.

    PubMed

    Li, Hui; Zhang, Huan; Jiang, Shuai; Wang, Weilin; Xin, Lusheng; Wang, Hao; Wang, Lingling; Song, Linsheng

    2015-06-01

    C-type lectins (CTLs), serving as pattern recognition receptors (PRRs), are a superfamily of Ca(2+)-dependent carbohydrate-recognition proteins that participate in nonself-recognition and pathogen elimination. In the present study, a single carbohydrate-recognition domain (CRD) CTL was identified from oyster Crassostrea gigas (designated as CgCLec-2). There was only one CRD within the deduced amino acid sequence of CgCLec-2 consisting of 129 amino acid residues. A conserved EPN (Glu246-Pro247-Asn248) motif was found in Ca(2+)-binding site 2 of CgCLec-2. The CgCLec-2 mRNA could be detected in all the examined tissues at different expression levels in oysters. The mRNA expression of CgCLec-2 in hemocytes was up-regulated significantly at 6h post Vibrio splendidus challenge. The recombinant CgCLec-2 (rCgCLec-2) could bind various Pathogen-Associated Molecular Patterns (PAMPs), including lipopolysaccharide, mannan and peptidoglycan, and displayed strong binding abilities to Vibrio anguillarum, V. splendidus and Yarrowiali polytica and week binding ability to Staphylococcus aureus. It could also enhance the phagocytic activity of oyster hemocytes to V. splendidus and exhibited growth suppression activity against gram-positive bacteria S. aureus but no effect on gram-negative bacteria V. splendidus. Furthermore, the interaction between rCgCLec-2 and rCgMASPL-1 was confirmed by GST Pull down. The results suggested that CgCLec-2 served as not only a PRR in immune recognition but also a regulatory factor in pathogen elimination, and played a potential role in the activation of complement system. PMID:25800112

  18. Edwardsiella tarda evades serum killing by preventing complement activation via the alternative pathway.

    PubMed

    Li, Mo-fei; Sun, Li; Li, Jun

    2015-04-01

    Edwardsiella tarda is a Gram-negative bacterium with a broad host range that includes a wide variety of farmed fish as well as humans. E. tarda has long been known to be able to survive in host serum, but the relevant mechanism is unclear. In this study, we investigated the fundamental question, i.e. whether E. tarda activated serum complement or not. We found that (i) when incubated with flounder serum, E. tarda exhibited a high survival rate (87.6%), which was slightly but significantly reduced in the presence of Mg(2+); (ii) E. tarda-incubated serum possessed strong hemolytic activity and bactericidal activity, (iii) compared to the serum incubated with a complement-sensitive laboratory Escherichia coli strain, E. tarda-incubated serum exhibited much less chemotactic activity, (iv) in contrast to the serum incubated with live E. tarda, the serum incubated with heat-inactivated E. tarda exhibited no apparent hemolytic capacity. Taken together, these results indicate for the first time that E. tarda circumvents serum attack by preventing, to a large extent, complement activation via the alternative pathway, and that heat-labile surface structures likely play an essential role in the complement evasion of E. tarda. PMID:25575477

  19. Activation of the alternative pathway of complement during the acute phase of typical haemolytic uraemic syndrome.

    PubMed

    Ferraris, J R; Ferraris, V; Acquier, A B; Sorroche, P B; Saez, M S; Ginaca, A; Mendez, C F

    2015-07-01

    Haemolytic uraemic syndrome (HUS) is characterized by haemolytic anaemia, thrombocytopenia and acute renal failure. We studied the activation state of classical and alternative pathways of complement during the acute phase of Shiga toxin-associated HUS by performing a prospective study of 18 patients and 17 age-matched healthy controls to evaluate C3, C3c, C4, C4d, Bb and SC5b-9 levels. SC5b-9 levels were increased significantly in all patients at admission compared to healthy and end-stage renal disease controls, but were significantly higher in patients presenting with oliguria compared to those with preserved diuresis. C3 and C4 levels were elevated significantly at admission in the non-oliguric group when compared to controls. No significant differences were found for C4d values, whereas factor Bb was elevated in all patients and significantly higher in oliguric patients when compared to both controls and non-oliguric individuals. A positive and significant association was detected when Bb formation was plotted as a function of plasma SC5b-9 at admission. Bb levels declined rapidly during the first week, with values not significantly different from controls by days 3 and 5 for non-oligurics and oligurics, respectively. Our data demonstrate the activation of the alternative pathway of complement during the acute phase of Stx-associated HUS. This finding suggests that complement activation may represent an important trigger for the cell damage that occurs during the syndrome. PMID:25677399

  20. Restriction of the alternative pathway of human complement by intact Trypanosoma brucei subsp. gambiense.

    PubMed Central

    Devine, D V; Falk, R J; Balber, A E

    1986-01-01

    We studied the interaction of African trypanosomes with human complement. Bloodstream forms of Trypanosoma brucei subsp. gambiense isolated from mice activated the alternative pathway of complement during a 30-min incubation in vitro. In human serum, all cells remained intact and motile during this period. C3 was detected on the surface by a direct binding assay with a monoclonal antibody which recognizes C3b and iC3b. C3 deposition could also be detected by this radioimmunoassay when parasites were incubated with purified C3. Such C3 binding was enhanced by factor B, factor D, and magnesium. Surface deposition of factor B was demonstrated both by flow immunofluorescence analysis and binding of radiolabeled factor B. C3 binding and factor B binding were inhibitable by EDTA but not by ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N' -tetraacetic acid (EGTA). The inhibited binding could be restored by addition of magnesium. No human immunoglobulin G or mouse immunoglobulin was detected on the trypanosome surface. By flow cytometry, neither human C5 nor polymerized C9 was detected on trypanosomes incubated in serum, although this assay was able to detect C5 and C9 on the surface of complement-treated human erythrocytes. Using a radioimmunoassay which measures C5b-9 in serum, we found that there was no generation of SC5b-9 in serum which had been incubated with trypanosomes. We concluded that, although trypanosomes activate the alternative pathway of complement, they are not lysed, because the cascade does not continue beyond the establishment of C3 convertase. PMID:3633873

  1. Identification of peptidic inhibitors of the alternative complement pathway based on Staphylococcus aureus SCIN proteins.

    PubMed

    Summers, Brady J; Garcia, Brandon L; Woehl, Jordan L; Ramyar, Kasra X; Yao, Xiaolan; Geisbrecht, Brian V

    2015-10-01

    The complement system plays a central role in a number of human inflammatory diseases, and there is a significant need for development of complement-directed therapies. The discovery of an arsenal of anti-complement proteins secreted by the pathogen Staphylococcus aureus brought with it the potential for harnessing the powerful inhibitory properties of these molecules. One such family of inhibitors, the SCINs, interact with a functional "hot-spot" on the surface of C3b. SCINs not only stabilize an inactive form of the alternative pathway (AP) C3 convertase (C3bBb), but also overlap the C3b binding site of complement factors B and H. Here we determined that a conserved Arg residue in SCINs is critical for function of full-length SCIN proteins. Despite this, we also found SCIN-specific differences in the contributions of other residues found at the C3b contact site, which suggested that a more diverse repertoire of residues might be able to recognize this region of C3b. To investigate this possibility, we conducted a phage display screen aimed at identifying SCIN-competitive 12-mer peptides. In total, seven unique sequences were identified and all exhibited direct C3b binding. A subset of these specifically inhibited the AP in assays of complement function. The mechanism of AP inhibition by these peptides was probed through surface plasmon resonance approaches, which revealed that six of the seven peptides disrupted C3bBb formation by interfering with factor B/C3b binding. To our knowledge this study has identified the first small molecules that retain inhibitory properties of larger staphylococcal immune evasion proteins. PMID:26052070

  2. Complement activity is associated with disease severity in multifocal motor neuropathy

    PubMed Central

    Vlam, Lotte; Cats, Elisabeth A.; Harschnitz, Oliver; Jansen, Marc D.; Piepers, Sanne; Veldink, Jan Herman; Franssen, Hessel; Stork, Abraham C.J.; Heezius, Erik; Rooijakkers, Suzan H.M.; Herpers, Bjorn L.; van Strijp, Jos A.; van den Berg, Leonard H.

    2015-01-01

    Objective: To investigate whether high innate activity of the classical and lectin pathways of complement is associated with multifocal motor neuropathy (MMN) and whether levels of innate complement activity or the potential of anti-GM1 antibodies to activate the complement system correlate with disease severity. Methods: We performed a case-control study including 79 patients with MMN and 79 matched healthy controls. Muscle weakness was documented with Medical Research Council scale sum score and axonal loss with nerve conduction studies. Activity of the classical and lectin pathways of complement was assessed by ELISA. We also determined serum mannose-binding lectin (MBL) concentrations and polymorphisms in the MBL gene (MBL2) and quantified complement-activating properties of anti-GM1 IgM antibodies by ELISA. Results: Activity of the classical and lectin pathways, MBL2 genotypes, and serum MBL concentrations did not differ between patients and controls. Complement activation by anti-GM1 IgM antibodies was exclusively mediated through the classical pathway and correlated with antibody titers (p < 0.001). Logistic regression analysis showed that both high innate activity of the classical pathway of complement and high complement-activating capacity of anti-GM1 IgM antibodies were significantly associated with more severe muscle weakness and axonal loss. Conclusion: High innate activity of the classical pathway of complement and efficient complement-activating properties of anti-GM1 IgM antibodies are determinants of disease severity in patients with MMN. These findings underline the importance of anti-GM1 antibodymediated complement activation in the pathogenesis and clinical course of MMN. PMID:26161430

  3. Complement in animal development: unexpected roles of a highly conserved pathway.

    PubMed

    Leslie, Jonathan D; Mayor, Roberto

    2013-02-01

    The complement pathway is most famous for its role in immunity, orchestrating an exquisitely refined system for immune surveillance. At its core lies a cascade of proteolytic events that ultimately serve to recognise microbes, infected cells or debris and target them for elimination. Mounting evidence has shown that a number of the proteolytic intermediaries in this cascade have, in themselves, other functions in the body, signalling through receptors to drive events that appear to be unrelated to immune surveillance. It seems, then, that the complement system not only functions as an immunological effector, but also has cell-cell signalling properties that are utilised by a number of non-immunological processes. In this review we examine a number of these processes in the context of animal development, all of which share a requirement for precise control of cell behaviour in time and space. As we will see, the scope of the complement system's function is indeed much greater than we might have imagined only a few years ago. PMID:23665279

  4. The alternative complement pathway is longitudinally associated with adverse cardiovascular outcomes. The CODAM study.

    PubMed

    Hertle, Elisabeth; Arts, Ilja C W; van der Kallen, Carla J H; Feskens, Edith J M; Schalkwijk, Casper G; Stehouwer, Coen D A; van Greevenbroek, Marleen M J

    2016-01-27

    The alternative pathway of complement activation is highly reactive and can be activated spontaneously in the vasculature. Activation may contribute to vascular damage and development of cardiovascular disease (CVD). We aimed to investigate functional components of the alternative pathway in cardiovascular risk. We studied 573 individuals who were followed-up for seven years. At baseline, we measured the enhancer properdin; the rate-limiting protease factor D (FD); and a marker of systemic activation, Bb. Using generalised estimating equations, we investigated their longitudinal associations with cardiovascular events (CVE, N=89), CVD (N=159), low-grade inflammation (LGI), endothelial dysfunction (ED) and carotid intima-media thickness (cIMT). Furthermore, we investigated associations with incident CVE (N=39) and CVD (N=73) in 342 participants free of CVD at baseline. CVE included myocardial infarction, stroke, cardiac angioplasty and/or cardiac bypass. CVD additionally included ischaemia on an electrocardiogram and/or ankle-brachial index complement pathway contributes to processes of vascular damage, and that in particular a high potential to enhance alternative pathway activation may promote unfavourable cardiovascular outcomes in humans. PMID:26446431

  5. Borrelia burgdorferi BBK32 Inhibits the Classical Pathway by Blocking Activation of the C1 Complement Complex

    PubMed Central

    Wager, Beau; Höök, Magnus; Skare, Jon T.

    2016-01-01

    Pathogens that traffic in blood, lymphatics, or interstitial fluids must adopt strategies to evade innate immune defenses, notably the complement system. Through recruitment of host regulators of complement to their surface, many pathogens are able to escape complement-mediated attack. The Lyme disease spirochete, Borrelia burgdorferi, produces a number of surface proteins that bind to factor H related molecules, which function as the dominant negative regulator of the alternative pathway of complement. Relatively less is known about how B. burgdorferi evades the classical pathway of complement despite the observation that some sensu lato strains are sensitive to classical pathway activation. Here we report that the borrelial lipoprotein BBK32 potently and specifically inhibits the classical pathway by binding with high affinity to the initiating C1 complex of complement. In addition, B. burgdorferi cells that produce BBK32 on their surface bind to both C1 and C1r and a serum sensitive derivative of B. burgdorferi is protected from killing via the classical pathway in a BBK32-dependent manner. Subsequent biochemical and biophysical approaches localized the anti-complement activity of BBK32 to its globular C-terminal domain. Mechanistic studies reveal that BBK32 acts by entrapping C1 in its zymogen form by binding and inhibiting the C1 subcomponent, C1r, which serves as the initiating serine protease of the classical pathway. To our knowledge this is the first report of a spirochetal protein acting as a direct inhibitor of the classical pathway and is the only example of a biomolecule capable of specifically and noncovalently inhibiting C1/C1r. By identifying a unique mode of complement evasion this study greatly enhances our understanding of how pathogens subvert and potentially manipulate host innate immune systems. PMID:26808924

  6. Borrelia burgdorferi BBK32 Inhibits the Classical Pathway by Blocking Activation of the C1 Complement Complex.

    PubMed

    Garcia, Brandon L; Zhi, Hui; Wager, Beau; Höök, Magnus; Skare, Jon T

    2016-01-01

    Pathogens that traffic in blood, lymphatics, or interstitial fluids must adopt strategies to evade innate immune defenses, notably the complement system. Through recruitment of host regulators of complement to their surface, many pathogens are able to escape complement-mediated attack. The Lyme disease spirochete, Borrelia burgdorferi, produces a number of surface proteins that bind to factor H related molecules, which function as the dominant negative regulator of the alternative pathway of complement. Relatively less is known about how B. burgdorferi evades the classical pathway of complement despite the observation that some sensu lato strains are sensitive to classical pathway activation. Here we report that the borrelial lipoprotein BBK32 potently and specifically inhibits the classical pathway by binding with high affinity to the initiating C1 complex of complement. In addition, B. burgdorferi cells that produce BBK32 on their surface bind to both C1 and C1r and a serum sensitive derivative of B. burgdorferi is protected from killing via the classical pathway in a BBK32-dependent manner. Subsequent biochemical and biophysical approaches localized the anti-complement activity of BBK32 to its globular C-terminal domain. Mechanistic studies reveal that BBK32 acts by entrapping C1 in its zymogen form by binding and inhibiting the C1 subcomponent, C1r, which serves as the initiating serine protease of the classical pathway. To our knowledge this is the first report of a spirochetal protein acting as a direct inhibitor of the classical pathway and is the only example of a biomolecule capable of specifically and noncovalently inhibiting C1/C1r. By identifying a unique mode of complement evasion this study greatly enhances our understanding of how pathogens subvert and potentially manipulate host innate immune systems. PMID:26808924

  7. A zebrafish model for uremic toxicity: role of the complement pathway.

    PubMed

    Berman, Nathaniel; Lectura, Melisa; Thurman, Joshua M; Reinecke, James; Raff, Amanda C; Melamed, Michal L; Quan, Zhe; Evans, Todd; Meyer, Timothy W; Hostetter, Thomas H

    2013-01-01

    Many organic solutes accumulate in end-stage renal disease (ESRD) and some are poorly removed with urea-based prescriptions for hemodialysis. However, their toxicities have been difficult to assess. We have employed an animal model, the zebrafish embryo, to test the toxicity of uremic serum compared to control. Serum was obtained from stable ESRD patients predialysis or from normal subjects. Zebrafish embryos 24 h postfertilization were exposed to experimental media at a water:human serum ratio of 3:1. Those exposed to serum from uremic subjects had significantly reduced survival at 8 h (19 18 vs. 94 6%, p < 0.05, uremic serum vs. control, respectively). Embryos exposed to serum from ESRD subjects fractionated at 50 kDa showed significantly greater toxicity with the larger molecular weight fraction (83 11 vs. 7 17% survival, p < 0.05, <50 vs. >50 kDa, respectively). Heating serum abrogated its toxicity. EDTA, a potent inhibitor of complement by virtue of calcium chelation, reduced the toxicity of uremic serum compared to untreated uremic serum (96 5 vs. 28 20% survival, p < 0.016, chelated vs. nonchelated serum, respectively). Anti-factor B, a specific inhibitor of the alternative complement pathway, reduced the toxicity of uremic serum, compared to untreated uremic serum (98 6 vs. 3 9% survival, p < 0.016, anti-factor B treated vs. nontreated, respectively). Uremic serum is thus more toxic to zebrafish embryos than normal serum. Furthermore, this toxicity is associated with a fraction of large size, is inactivated by heat, and is reduced by both specific and nonspecific inhibitors of complement activation. Together these data lend support to the hypothesis that at least some uremic toxicities may be mediated by complement. PMID:23689420

  8. Coding polymorphisms in the genes of the alternative complement pathway and abdominal aortic aneurysm.

    PubMed

    Bradley, D T; Badger, S A; Bown, M J; Sayers, R D; Hughes, A E

    2011-06-01

    Variants in the genes of the alternative complement pathway are associated with risk of numerous inflammatory diseases. Abdominal aortic aneurysm is associated with inflammation and is a common cause of illness and death among European populations. This study tested 49 single nucleotide polymorphisms, including common putatively functional polymorphisms, in the genes of the alternative complement cascade (CFH, CFB, CFD, CFI, properdin, CR1, CR1L, CR2, CD46, vitronectin, C3, C5, C6, C7, C8A, C8B, C8G and C9). The study group were 434 cases with infra-renal aortic diameter ≥30 mm and 378 disease-free controls from two UK centres, all with self-reported European ancestry. There was no evidence for significant association with presence or size of aneurysm following correction for multiple testing. This study suggests that variation in the genes of the alternative pathway is not an important cause of abdominal aortic aneurysm development. PMID:21352499

  9. Genetic Investigation of Complement Pathway Genes in Type 2 Diabetic Retinopathy: An Inflammatory Perspective

    PubMed Central

    Yang, Ming Ming; Wang, Jun; Ren, Hong; Sun, Yun Duan; Fan, Jiao Jie; Teng, Yan; Li, Yan Bo

    2016-01-01

    Diabetic retinopathy (DR) has complex multifactorial pathogenesis. This study aimed to investigate the association of complement pathway genes with susceptibility to DR. Eight haplotype-tagging SNPs of SERPING1 and C5 were genotyped in 570 subjects with type 2 diabetes: 295 DR patients (138 nonproliferative DR [NPDR] and 157 proliferative DR [PDR]) and 275 diabetic controls. Among the six C5 SNPs, a marginal association was first detected between rs17611 and total DR patients (P = 0.009, OR = 0.53 for recessive model). In stratification analysis, a significant decrease in the frequencies of G allele and GG homozygosity for rs17611 was observed in PDR patients compared with diabetic controls (Pcorr = 0.032, OR = 0.65 and Pcorr = 0.016, OR = 0.37, resp.); it was linked with a disease progression. A haplotype AA defined by the major alleles of rs17611 and rs1548782 was significantly predisposed to PDR with increased risk of 1.54 (Pcorr = 0.023). Regarding other variants in C5 and SERPING1, none of the tagging SNPs had a significant association with DR and its subgroups (all P > 0.05). Our study revealed an association between DR and C5 polymorphisms with clinical significance, whereas SERPING1 is not a major genetic component of DR. Our data suggest a link of complement pathway with DR pathogenesis. PMID:26989329

  10. Inhibition of the classical pathway of complement by meningococcal capsular polysaccharides

    PubMed Central

    Agarwal, Sarika; Vasudhev, Shreekant; DeOliveira, Rosane; Ram, Sanjay

    2014-01-01

    Almost all invasive Neisseria meningitidis isolates express capsular polysaccharide. Antibody (Ab) is required for complement-dependent killing of meningococci. While alternative pathway evasion has received considerable attention, little is known about classical pathway (CP) inhibition by meningococci and forms the basis of this study. We engineered capsulated and unencapsulated isogenic mutant strains of groups A, B, C, W and Y meningococci to express similar amounts of the same factor H-binding protein (fHbp; a key component of group B meningococcal vaccines) molecule. Despite similar anti-fHbp mAb binding, significantly less C4b was deposited on all five encapsulated mutants compared to their unencapsulated counterparts (P<0.01), when purified C1 and C4 were used to deposit C4b. Reduced C4b deposition was the result of capsule-mediated inhibition of C1q engagement by Ab. C4b deposition correlated linearly with C1q engagement by anti-fHbp. While B, C, W and Y capsules limited CP-mediated killing by anti-fHbp, the unencapsulated group A mutant paradoxically was more resistant than its encapsulated counterpart. Strains varied considerably in their susceptibility to anti-fHbp and complement despite similar Ab binding, which may have implications for the activity of fHbp-based vaccines. Capsule also limited C4b deposition by anti-porin A mAbs. Capsule expression decreased binding of an anti-LOS IgM mAb (~1.2 to 2-fold reduction in fluorescence). Akin to observations with IgG, capsule also decreased IgM-mediated C4b deposition when IgM binding to the mutant strain pairs was normalized. In conclusion, we show that capsular polysaccharide, a critical meningococcal virulence factor, inhibits the CP of complement. PMID:25015832

  11. Bioactive Lysophospholipids Generated by Hepatic Lipase Degradation of Lipoproteins Lead to Complement Activation via the Classical Pathway

    PubMed Central

    Ma, Wanchao; Paik, David C.; Barile, Gaetano R.

    2014-01-01

    Purpose. We determined bioactivity of lysophospholipids generated by degradation of the low-density (LDL), very low-density (VLDL), and high-density (HDL) lipoproteins with hepatic lipase (HL), cholesterol esterase (CE), and lipoprotein-associated phospholipase A2 (Lp-PLA2). Methods. The LDL, VLDL, and HDL were treated with HL, CE, and Lp-PLA2 after immobilization on plates, and complement activation studies were performed with diluted human serum. Complement component 3 (C3) fixation, a marker for complement activation, was determined with a monoclonal anti-human C3d antibody. Enzymatic properties of HL and CE were assayed with triglyceride and phosphatidylcholine substrates for triglyceride hydrolase and phospholipase A activities. The ARPE-19 cells were used for viability studies. Results. The HL degradation of human lipoproteins LDL, VLDL, or HDL results in the formation of modified lipoproteins that can activate the complement pathway. Complement activation is dose- and time-dependent upon HL and occurs via the classical pathway. Enzymatic studies suggest that the phospholipase A1 activity of HL generates complement-activating lysophospholipids. C-reactive protein (CRP), known to simultaneously interact with complement C1 and complement factor H (CFH), further enhances HL-induced complement activation. The lysophospholipids, 1-Palmitoyl-sn-glycero-3-phosphocholine and 1-Oleoyl-sn-glycero-3-phosphocholine, can be directly cytotoxic to ARPE-19 cells. Conclusions. The HL degradation of lipoproteins, known to accumulate in the outer retina and in drusen, can lead to the formation of bioactive lysophospholipids that can trigger complement activation and induce RPE cellular dysfunction. Given the known risk associations for age-related macular degeneration (AMD) with HL, CRP, and CFH, this study elucidates a possible damage pathway for age-related macular degeneration (AMD) in genetically predisposed individuals, that HL activity may lead to accumulation of lysophospholipids to initiate complement activation, with CFH dysregulation exacerbating the effects of this process. PMID:25205869

  12. Differences in Cryptococcus neoformans capsular polysaccharide structure influence assembly of alternative complement pathway C3 convertase on fungal surfaces.

    PubMed

    Washburn, R G; Bryant-Varela, B J; Julian, N C; Bennett, J E

    1991-01-01

    Binding of complement component C3 and Factor B to Cryptococcus neoformans serotypes A through D via the alternative complement pathway was measured in a system containing fresh nonimmune human serum. Serotypes B and C (C. neoformans var. gattii) bound approximately half as many molecules of both complement components as serotypes A and D (C. neoformans var. neoformans). In contrast, removal of xylosyl and glucuronyl side chains from the mannan main chain of capsular polysaccharide by the Smith degradation procedure resulted in binding of similar quantities of C3 to each of the four serotypes. We conclude that the relatively high degree of side chain substitution of capsular polysaccharide from C. neoformans variety gattii contributes to inefficient surface assembly of the alternative pathway C3 convertase. Inefficient binding of alternative pathway complement components to serotypes B and C may contribute to the relative difficulty in successfully treating infections caused by these organisms. PMID:2062324

  13. Interspecies Complementation of the LuxR Family Pathway-Specific Regulator Involved in Macrolide Biosynthesis.

    PubMed

    Mo, SangJoon; Yoon, Yeo Joon

    2016-01-28

    PikD is a widely known pathway-specific regulator for controlling pikromycin production in Streptomyces venezuelae ATCC 15439, which is a representative of the large ATP-binding regulator of the LuxR family (LAL) in Streptomyces sp. RapH and FkbN also belong to the LAL family of transcriptional regulators, which show greatest homology with the ATP-binding motif and helix-turn-helix DNA-binding motif of PikD. Overexpression of pikD and heterologous expression of rapH and fkbN led to enhanced production of pikromycin by approximately 1.8-, 1.6-, and 1.6-fold in S. venezuelae, respectively. Cross-complementation of rapH and fkbN in the pikD deletion mutant (?pikD) restored pikromycin and derived macrolactone production. Overall, these results show that heterologous expression of rapH and fkbN leads to the overproduction of pikromycin and its congeners from the pikromycin biosynthetic pathway in S. venezuelae, and they have the same functionality as the pathwayspecific transcriptional activator for the pikromycin biosynthetic pathway in the ?pikD strain. These results also show extensive "cross-communication" between pathway-specific regulators of streptomycetes and suggest revision of the current paradigm for pathwayspecific versus global regulation of secondary metabolism in Streptomyces species. PMID:26608164

  14. Role of the terminal complement pathway in experimental membranous nephropathy in the rabbit.

    PubMed Central

    Groggel, G C; Adler, S; Rennke, H G; Couser, W G; Salant, D J

    1983-01-01

    Our recent observations of a complement-mediated, cell-independent mechanism of altered glomerular permeability in rat membranous nephropathy suggested a possible role for the terminal complement pathway in the mediation of proteinuria in certain forms of glomerular disease. To directly determine whether the membranolytic terminal complement components (C5b-C9) are involved in glomerular injury, we studied the development of proteinuria in normal and C6-deficient (C6D) rabbits, in both of which a membranous nephropathy-like lesion develops early in the course of immunization with cationized bovine serum albumin (cBSA) (pI 8.9-9.2). C6 hemolytic activity of C6D was 0.01% that of control rabbits. After 1 wk of daily intravenous injections of cBSA, proteinuria developed in 71% of controls (median 154, range 1-3,010 mg/24 h, n = 24), whereas none of C6D were proteinuric (median 6, range 2-12 mg/24 h, n = 12, P less than 0.01). After 1 wk of cBSA, both groups had qualitatively identical glomerular deposits of BSA, rabbit IgG, and C3 on immunofluorescence microscopy, predominantly subepithelial electron-dense deposits on electron microscopy, and minimal glomerular inflammatory cell infiltration of glomeruli. Glomeruli were isolated from individual animals after 1 wk of cBSA and deposits of rabbit IgG antibody were quantitated by a standardized in vitro assay using anti-rabbit IgG-125I. Rabbit IgG deposits were found to be similar in control (29.8 +/- 13.2, range 12.7-48.6 micrograms anti-IgG/2,000 glomeruli, n = 6) and C6D rabbits (32.6 +/- 13.8, range 16.8-48.8 micrograms anti-IgG/2,000 glomeruli, n = 5, P greater than 0.05). After 2 wk, coincident with a prominent influx of mononuclear cells and neutrophils, proteinuria developed in C6D rabbits. These results document, for the first time, a requirement for a terminal complement component in the development of immunologic glomerular injury. Since the only known action of C6 is in the assembly of the membrane attack complex, these observations suggest that the membranolytic properties of complement may contribute to glomerular damage. Images PMID:6227634

  15. A component of the medicinal herb ephedra blocks activation in the classical and alternative pathways of complement.

    PubMed

    Ling, M; Piddlesden, S J; Morgan, B P

    1995-12-01

    Extracts of the herb Ephedra sinica have long been used in traditional Chinese medicine for the treatment of, among other conditions, acute nephritis. In preliminary studies it was shown that extracts of ephedra caused inhibition of complement in vitro. We thus set out to isolate the active component(s) of this herb, to examine the complement-inhibiting capacity in sera from different species, and to characterize the mechanism(s) by which it inhibits complement. Aqueous extraction of the herb followed by fractionation using thin layer chromatography (TLC) demonstrated that complement-inhibiting activity resided within a single band, hereafter termed the complement-inhibiting component (CIC), which represents an as yet uncharacterized polyanionic carbohydrate molecule. TLC-purified CIC inhibited the classical complement pathway in all species tested (human, pig, guinea pig, rat and rabbit). Using erythrocyte intermediates and sera specifically depleted of individual components it was apparent that CIC inhibited C2. This finding was confirmed using purified human C2, CIC causing a dose-dependent loss of C2 haemolytic activity. At much higher doses, CIC also showed some inhibiting effect in the terminal pathway, and this was shown to be due to inhibition of C9. In the alternative pathway CIC also showed inhibitory activity, although its site of action in this pathway remains unresolved. In Chinese medicine the herb is taken orally, though no studies of complement levels in patients taking the herb have been reported. Preliminary data indicate that oral administration in rats causes a partial inhibition of serum complement activity. Given the current enthusiasm for complement inhibition as a therapy for inflammatory diseases, this non-toxic, naturally occurring agent might be of therapeutic value. PMID:8536376

  16. All-trans-Retinal Sensitizes Human RPE Cells to Alternative Complement PathwayInduced Cell Death

    PubMed Central

    Berchuck, Jacob E.; Yang, Ping; Toimil, Brett A.; Ma, Zhe; Baciu, Peter; Jaffe, Glenn J.

    2013-01-01

    Purpose. Retinal pigment epithelial (RPE) cell death occurs early in the pathogenesis of age-related macular degeneration (AMD) and Stargardt's disease. Emerging evidence suggests that all-trans-retinal (atRal) and alternative complement pathway (AP) activation contribute to RPE cell death in both of these retinal disorders. The aim of this study was to investigate the combined effect of atRal and AP activation on RPE cell viability. Methods. RPE cells were treated with atRal and then incubated with a complement-fixing antibody followed by stimulation with C1q-depleted serum to activate AP. Cell viability was assessed by tetrazolium salt and lactate dehydrogenase release assays. Changes in cell surface CD46 and CD59 expression were assessed by flow cytometry. Cells were pretreated with the antioxidant resveratrol, and C1q-depleted serum was incubated with an anti-C5 antibody prior to initiating AP attack to determine the protective effects of antioxidant therapy and complement inhibition, respectively. Results. Both atRal and AP activation independently caused RPE cell death. When AP attack was initiated following atRal treatment, a synergistic increase in cell death was observed. Following 24-hour atRal treatment, CD46 and CD59 expression decreased, corresponding temporally to increased susceptibility to AP attack. Resveratrol and the anti-C5 antibody both protected against AP-induced cell death following atRal exposure and were most effective when used in combination. Conclusions. atRal sensitizes RPE cells to AP attack, which may be mediated in part by atRal-induced downregulation of CD46 and CD59. Despite increased susceptibility to AP attack following exposure to atRal, resveratrol and anti-C5 antibody effectively prevent AP-mediated cell death. PMID:23518773

  17. Early Components of the Complement Classical Activation Pathway in Human Systemic Autoimmune Diseases

    PubMed Central

    Lintner, Katherine E.; Wu, Yee Ling; Yang, Yan; Spencer, Charles H.; Hauptmann, Georges; Hebert, Lee A.; Atkinson, John P.; Yu, C. Yung

    2016-01-01

    The complement system consists of effector proteins, regulators, and receptors that participate in host defense against pathogens. Activation of the complement system, via the classical pathway (CP), has long been recognized in immune complex-mediated tissue injury, most notably systemic lupus erythematosus (SLE). Paradoxically, a complete deficiency of an early component of the CP, as evidenced by homozygous genetic deficiencies reported in human, are strongly associated with the risk of developing SLE or a lupus-like disease. Similarly, isotype deficiency attributable to a gene copy-number (GCN) variation and/or the presence of autoantibodies directed against a CP component or a regulatory protein that result in an acquired deficiency are relatively common in SLE patients. Applying accurate assay methodologies with rigorous data validations, low GCNs of total C4, and heterozygous and homozygous deficiencies of C4A have been shown as medium to large effect size risk factors, while high copy numbers of total C4 or C4A as prevalent protective factors, of European and East-Asian SLE. Here, we summarize the current knowledge related to genetic deficiency and insufficiency, and acquired protein deficiencies for C1q, C1r, C1s, C4A/C4B, and C2 in disease pathogenesis and prognosis of SLE, and, briefly, for other systemic autoimmune diseases. As the complement system is increasingly found to be associated with autoimmune diseases and immune-mediated diseases, it has become an attractive therapeutic target. We highlight the recent developments and offer a balanced perspective concerning future investigations and therapeutic applications with a focus on early components of the CP in human systemic autoimmune diseases. PMID:26913032

  18. Peptide Inhibitor of Complement C1 (PIC1) Rapidly Inhibits Complement Activation after Intravascular Injection in Rats

    PubMed Central

    Sharp, Julia A.; Hair, Pamela S.; Pallera, Haree K.; Kumar, Parvathi S.; Mauriello, Clifford T.; Nyalwidhe, Julius O.; Phelps, Cody A.; Park, Dalnam; Thielens, Nicole M.; Pascal, Stephen M.; Chen, Waldon; Duffy, Diane M.; Lattanzio, Frank A.; Cunnion, Kenji M.; Krishna, Neel K.

    2015-01-01

    The complement system has been increasingly recognized to play a pivotal role in a variety of inflammatory and autoimmune diseases. Consequently, therapeutic modulators of the classical, lectin and alternative pathways of the complement system are currently in pre-clinical and clinical development. Our laboratory has identified a peptide that specifically inhibits the classical and lectin pathways of complement and is referred to as Peptide Inhibitor of Complement C1 (PIC1). In this study, we determined that the lead PIC1 variant demonstrates a salt-dependent binding to C1q, the initiator molecule of the classical pathway. Additionally, this peptide bound to the lectin pathway initiator molecule MBL as well as the ficolins H, M and L, suggesting a common mechanism of PIC1 inhibitory activity occurs via binding to the collagen-like tails of these collectin molecules. We further analyzed the effect of arginine and glutamic acid residue substitution on the complement inhibitory activity of our lead derivative in a hemolytic assay and found that the original sequence demonstrated superior inhibitory activity. To improve upon the solubility of the lead derivative, a pegylated, water soluble variant was developed, structurally characterized and demonstrated to inhibit complement activation in mouse plasma, as well as rat, non-human primate and human serum in vitro. After intravenous injection in rats, the pegylated derivative inhibited complement activation in the blood by 90% after 30 seconds, demonstrating extremely rapid function. Additionally, no adverse toxicological effects were observed in limited testing. Together these results show that PIC1 rapidly inhibits classical complement activation in vitro and in vivo and is functional for a variety of animal species, suggesting its utility in animal models of classical complement-mediated diseases. PMID:26196285

  19. Complement regulators in human disease: lessons from modern genetics.

    PubMed

    K Liszewski, M; Atkinson, J P

    2015-03-01

    First identified in human serum in the late 19th century as a 'complement' to antibodies in mediating bacterial lysis, the complement system emerged more than a billion years ago probably as the first humoral immune system. The contemporary complement system consists of nearly 60 proteins in three activation pathways (classical, alternative and lectin) and a terminal cytolytic pathway common to all. Modern molecular biology and genetics have not only led to further elucidation of the structure of complement system components, but have also revealed function-altering rare variants and common polymorphisms, particularly in regulators of the alternative pathway, that predispose to human disease by creating 'hyperinflammatory complement phenotypes'. To treat these 'complementopathies', a monoclonal antibody against the initiator of the membrane attack complex, C5, has received approval for use. Additional therapeutic reagents are on the horizon. PMID:25495259

  20. Sodium arsenite-induced alteration in hepatocyte function of rat with special emphasis on superoxide dismutase expression pathway and its prevention by mushroom lectin.

    PubMed

    Bera, Asit K; Rana, Tanmoy; Bhattacharya, Debasis; Das, Subhashree; Pan, Diganta; Das, Subrata K

    2011-10-01

    This study was accomplished to exemplify the possible protective role of ascorbic acid and mushroom lectin against arsenic-induced cytotoxicity and impairment of superoxide dismutase (SOD) production pathway in hepatocytes of rat. Hepatocytes were isolated from rat and treated with sodium arsenite (AS), arsenic plus ascorbic acid (AS + AA) and arsenic plus mushroom lectin (AS + ML). A placebo control was also included. Arsenic treatment resulted in the depletion of cell proliferation, phagocytic activity (nitro blue tetrazolium index) and superoxide dismutase (SOD) activity, relative mRNA expression of superoxide dismutase 2 (SOD(2)) and enhanced production of nitric oxide (NO). Ascorbic acid, a standard antioxidant, could normalize cellular perturbation and SOD production pathway relating to gene expression, whereas partially purified Pleurotus florida lectin (PFL), an edible mushroom containing protein complex, maintained cellular activity and prevented stress by normalizing phagocytic (NBT index) and SOD activities vis--vis relative gene expression. It could further defend NO production of hepatocytes. Mushroom lectin strongly prevented sodium arsenite-induced damage of SOD production pathway in hepatocytes, and its effect was also comparable to a standard antioxidant, i.e. ascorbic acid. PMID:21554548

  1. Pseudomonas aeruginosa Uses Dihydrolipoamide Dehydrogenase (Lpd) to Bind to the Human Terminal Pathway Regulators Vitronectin and Clusterin to Inhibit Terminal Pathway Complement Attack

    PubMed Central

    Hallström, Teresia; Uhde, Melanie; Singh, Birendra; Skerka, Christine; Riesbeck, Kristian; Zipfel, Peter F.

    2015-01-01

    The opportunistic human pathogen Pseudomonas aeruginosa controls host innate immune and complement attack. Here we identify Dihydrolipoamide dehydrogenase (Lpd), a 57 kDa moonlighting protein, as the first P. aeruginosa protein that binds the two human terminal pathway inhibitors vitronectin and clusterin. Both human regulators when bound to the bacterium inhibited effector function of the terminal complement, blocked C5b-9 deposition and protected the bacterium from complement damage. P. aeruginosa when challenged with complement active human serum depleted from vitronectin was severely damaged and bacterial survival was reduced by over 50%. Similarly, when in human serum clusterin was blocked by a mAb, bacterial survival was reduced by 44%. Thus, demonstrating that Pseudomonas benefits from attachment of each human regulator and controls complement attack. The Lpd binding site in vitronectin was localized to the C-terminal region, i.e. to residues 354–363. Thus, Lpd of P. aeruginosa is a surface exposed moonlighting protein that binds two human terminal pathway inhibitors, vitronectin and clusterin and each human inhibitor when attached protected the bacterial pathogen from the action of the terminal complement pathway. Our results showed insights into the important function of Lpd as a complement regulator binding protein that might play an important role in virulence of P. aeruginosa. PMID:26368530

  2. MytiLec, a Mussel R-Type Lectin, Interacts with Surface Glycan Gb3 on Burkitt's Lymphoma Cells to Trigger Apoptosis through Multiple Pathways.

    PubMed

    Hasan, Imtiaj; Sugawara, Shigeki; Fujii, Yuki; Koide, Yasuhiro; Terada, Daiki; Iimura, Naoya; Fujiwara, Toshiyuki; Takahashi, Keisuke G; Kojima, Nobuhiko; Rajia, Sultana; Kawsar, Sarkar M A; Kanaly, Robert A; Uchiyama, Hideho; Hosono, Masahiro; Ogawa, Yukiko; Fujita, Hideaki; Hamako, Jiharu; Matsui, Taei; Ozeki, Yasuhiro

    2015-01-01

    MytiLec; a novel lectin isolated from the Mediterranean mussel (Mytilus galloprovincialis); shows strong binding affinity to globotriose (Gb3: Galα1-4Galβ1-4Glc). MytiLec revealed β-trefoil folding as also found in the ricin B-subunit type (R-type) lectin family, although the amino acid sequences were quite different. Classification of R-type lectin family members therefore needs to be based on conformation as well as on primary structure. MytiLec specifically killed Burkitt's lymphoma Ramos cells, which express Gb3. Fluorescein-labeling assay revealed that MytiLec was incorporated inside the cells. MytiLec treatment of Ramos cells resulted in activation of both classical MAPK/ extracellular signal-regulated kinase and extracellular signal-regulated kinase (MEK-ERK) and stress-activated (p38 kinase and JNK) Mitogen-activated protein kinases (MAPK) pathways. In the cells, MytiLec treatment triggered expression of tumor necrosis factor (TNF)-α (a ligand of death receptor-dependent apoptosis) and activation of mitochondria-controlling caspase-9 (initiator caspase) and caspase-3 (activator caspase). Experiments using the specific MEK inhibitor U0126 showed that MytiLec-induced phosphorylation of the MEK-ERK pathway up-regulated expression of the cyclin-dependent kinase inhibitor p21, leading to cell cycle arrest and TNF-α production. Activation of caspase-3 by MytiLec appeared to be regulated by multiple different pathways. Our findings, taken together, indicate that the novel R-type lectin MytiLec initiates programmed cell death of Burkitt's lymphoma cells through multiple pathways (MAPK cascade, death receptor signaling; caspase activation) based on interaction of the lectin with Gb3-containing glycosphingolipid-enriched microdomains on the cell surface. PMID:26694420

  3. MytiLec, a Mussel R-Type Lectin, Interacts with Surface Glycan Gb3 on Burkitt’s Lymphoma Cells to Trigger Apoptosis through Multiple Pathways

    PubMed Central

    Hasan, Imtiaj; Sugawara, Shigeki; Fujii, Yuki; Koide, Yasuhiro; Terada, Daiki; Iimura, Naoya; Fujiwara, Toshiyuki; Takahashi, Keisuke G.; Kojima, Nobuhiko; Rajia, Sultana; Kawsar, Sarkar M. A.; Kanaly, Robert A.; Uchiyama, Hideho; Hosono, Masahiro; Ogawa, Yukiko; Fujita, Hideaki; Hamako, Jiharu; Matsui, Taei; Ozeki, Yasuhiro

    2015-01-01

    MytiLec; a novel lectin isolated from the Mediterranean mussel (Mytilus galloprovincialis); shows strong binding affinity to globotriose (Gb3: Galα1-4Galβ1-4Glc). MytiLec revealed β-trefoil folding as also found in the ricin B-subunit type (R-type) lectin family, although the amino acid sequences were quite different. Classification of R-type lectin family members therefore needs to be based on conformation as well as on primary structure. MytiLec specifically killed Burkitt's lymphoma Ramos cells, which express Gb3. Fluorescein-labeling assay revealed that MytiLec was incorporated inside the cells. MytiLec treatment of Ramos cells resulted in activation of both classical MAPK/ extracellular signal-regulated kinase and extracellular signal-regulated kinase (MEK-ERK) and stress-activated (p38 kinase and JNK) Mitogen-activated protein kinases (MAPK) pathways. In the cells, MytiLec treatment triggered expression of tumor necrosis factor (TNF)-α (a ligand of death receptor-dependent apoptosis) and activation of mitochondria-controlling caspase-9 (initiator caspase) and caspase-3 (activator caspase). Experiments using the specific MEK inhibitor U0126 showed that MytiLec-induced phosphorylation of the MEK-ERK pathway up-regulated expression of the cyclin-dependent kinase inhibitor p21, leading to cell cycle arrest and TNF-α production. Activation of caspase-3 by MytiLec appeared to be regulated by multiple different pathways. Our findings, taken together, indicate that the novel R-type lectin MytiLec initiates programmed cell death of Burkitt’s lymphoma cells through multiple pathways (MAPK cascade, death receptor signaling; caspase activation) based on interaction of the lectin with Gb3-containing glycosphingolipid-enriched microdomains on the cell surface. PMID:26694420

  4. Complementing the inflammasome.

    PubMed

    Triantafilou, Martha; Hughes, Timothy R; Morgan, Bryan Paul; Triantafilou, Kathy

    2016-02-01

    The innate immune system is an ancient surveillance system able to sense microbial invaders as well as aberrations in normal cell function. No longer viewed as a static and non-specific part of immunity, the innate immune system employs a plethora of specialized pattern recognition sensors to monitor and achieve homeostasis; these include the Toll-like receptors, the retinoic acid-inducible gene-like receptors, the nucleotide-binding oligomerization domain receptors (NLRs), the C-type lectins and the complement system. In order to increase specificity and diversity, innate immunity uses homotypic and heterotypic associations among these different components. Multi-molecular assemblies are formed both on the cell surface and in the cytosol to respond to pathogen and danger signals. Diverse, but tailored, responses to a changing environment are orchestrated depending on the the nature of the challenge and the repertoire of interacting receptors and components available in the sensing cell. It is now emerging that innate immunity operates a system of 'checks and balances' where interaction among the sensors is key in maintaining normal cell function. Complement sits at the heart of this alarm system and it is becoming apparent that it is capable of interacting with all the other pathways to effect a tailored immune response. In this review, we will focus on complement interactions with NLRs, the so-called 'inflammasomes', describing the molecular mechanisms that have been revealed so far and discussing the circumstantial evidence that exists for these interactions in disease states. PMID:26572245

  5. A Novel Factor I Activity in Nipah Virus Inhibits Human Complement Pathways through Cleavage of C3b

    PubMed Central

    Johnson, John B.; Borisevich, Viktoriya; Rockx, Barry

    2014-01-01

    ABSTRACT Complement is an innate immune system that most animal viruses must face during natural infections. Given that replication and dissemination of the highly pathogenic Nipah virus (NiV) include exposure to environments rich in complement factors, we tested the in vitro sensitivity of NiV to complement-mediated neutralization. Here we show that NiV was completely resistant to in vitro neutralization by normal human serum (NHS). Treatment of purified NiV with NHS activated complement pathways, but there was very little C3 deposition on virus particles. In in vitro reconstitution experiments, NiV particles provided time- and dose-dependent factor I-like protease activity capable of cleaving C3b into inactive C3b (iC3b). NiV-dependent inactivation of C3b only occurred with the cofactors factor H and soluble CR1 but not with CD46. Purified NiV particles did not support C4b cleavage. Electron microscopy of purified NiV particles showed immunogold labeling with anti-factor I antibodies. Our results suggest a novel mechanism by which NiV evades the human complement system through a unique factor I-like activity. IMPORTANCE Viruses have evolved mechanisms to limit complement-mediated neutralization, some of which involve hijacking cellular proteins involved in control of inappropriate complement activation. Here we report a previously unknown mechanism whereby NiV provides a novel protease activity capable of in vitro cleavage and inactivation of C3b, a key component of the complement cascade. These data help to explain how an enveloped virus such as NiV can infect and disseminate through body fluids that are rich in complement activity. Disruption of the ability of NiV to recruit complement inhibitors could form the basis for the development of effective therapies and safer vaccines to combat these highly pathogenic emerging viruses. PMID:25355897

  6. Natural IgG antibodies in normal rabbit serum are involved in killing of the ompP2 mutant of Haemophilus parasuis SC096 strain via the classical complement pathway.

    PubMed

    Zhou, S M; Xu, C G; Zhang, B; Feng, S X; Zhang, L Y; Zou, Y; Liao, M

    2013-04-01

    Serum resistance in Haemophilus parasuis strain SC096 has been shown to be dependent on expression of the outer membrane protein P2 (OmpP2) and loss of the ompP2 gene results in significantly greater sensitivity to both porcine and rabbit sera. However, the mechanism of complement activation by the serum sensitive ?ompP2 strain is unknown. In this study, the classical complement pathway is demonstrated to be the main pathway for killing the H. parasuis ?ompP2 strain, and not the mannan-binding lectin (MBL) or alternative pathway. In addition, absorption of antibodies against ?ompP2 strain or depletion of IgGs from serum inhibited serum killing activity, which could be restored by addition of heat-inactivated serum or purified IgGs. Western blot analysis indicated that the OmpP2 mutant could bind significantly more IgGs than the wild type strain SC096 when incubated with serum. Finally, IgGs in normal rabbit serum targeted to the OMPs, but not lipooligosaccharide (LOS) in the OmpP2 mutant strain were found to be involved in bacterial killing indicating that the bactericidal epitope(s) is in the outer membrane proteins. PMID:23103218

  7. Complement Defects in Patients with Chronic Rhinosinusitis

    PubMed Central

    Gaunsbaek, Maria Q.; Lange, Bibi; Kjeldsen, Anette D.; Svane-Knudsen, Viggo; Skjoedt, Karsten; Henriksen, Maiken L.; Nielsen, Christian; Palarasah, Yaseelan; Hansen, Soren

    2012-01-01

    The complement system is an important part of our immune system, and complement defects lead generally to increased susceptibility to infections and autoimmune diseases. We have studied the role of complement activity in relation with chronic rhinosinusitis (CRS), and more specifically studied whether complement defects collectively predispose individuals for CRS or affect CRS severity. The participants comprised 87 CRS patients randomly selected from the general population, and a control group of 150 healthy blood donors. The CRS patients were diagnosed according to the European Position Paper on Rhinosinusitis and nasal Polyps criteria, and severity was evaluated by the Sino-nasal Outcome Test-22. Serum samples were analysed by ELISA for activity of the respective pathways of complement, and subsequently for serum levels of relevant components. We found that the frequency of complement defects was significantly higher among CRS patients than among healthy control subjects. A majority of Mannan-binding lectin deficient CRS patients was observed. The presence of complement defects had no influence on the severity of subjective symptoms. Our studies show that defects in the complement system collectively may play an immunological role related to the development of CRS. However, an association between severity of symptoms and presence of complement defects could not be demonstrated. PMID:23144819

  8. Characterization of a Factor H Mutation That Perturbs the Alternative Pathway of Complement in a Family with Membranoproliferative GN

    PubMed Central

    Wong, Edwin K.S.; Anderson, Holly E.; Herbert, Andrew P.; Challis, Rachel C.; Brown, Paul; Reis, Geisilaine S.; Tellez, James O.; Strain, Lisa; Fluck, Nicholas; Humphrey, Ann; Macleod, Alison; Richards, Anna; Ahlert, Daniel; Santibanez-Koref, Mauro; Barlow, Paul N.; Marchbank, Kevin J.; Harris, Claire L.; Goodship, Timothy H.J.

    2014-01-01

    Complement C3 activation is a characteristic finding in membranoproliferative GN (MPGN). This activation can be caused by immune complex deposition or an acquired or inherited defect in complement regulation. Deficiency of complement factor H has long been associated with MPGN. More recently, heterozygous genetic variants have been reported in sporadic cases of MPGN, although their functional significance has not been assessed. We describe a family with MPGN and acquired partial lipodystrophy. Although C3 nephritic factor was shown in family members with acquired partial lipodystrophy, it did not segregate with the renal phenotype. Genetic analysis revealed a novel heterozygous mutation in complement factor H (R83S) in addition to known risk polymorphisms carried by individuals with MPGN. Patients with MPGN had normal levels of factor H, and structural analysis of the mutant revealed only subtle alterations. However, functional analysis revealed profoundly reduced C3b binding, cofactor activity, and decay accelerating activity leading to loss of regulation of the alternative pathway. In summary, this family showed a confluence of common and rare functionally significant genetic risk factors causing disease. Data from our analysis of these factors highlight the role of the alternative pathway of complement in MPGN. PMID:24722444

  9. Down-Regulation of Complement Receptors on the Surface of Host Monocyte Even as In Vitro Complement Pathway Blocking Interferes in Dengue Infection

    PubMed Central

    Marinho, Cintia Ferreira; Azeredo, Elzinandes Leal; Torrentes-Carvalho, Amanda; Marins-Dos-Santos, Alessandro; Kubelka, Claire Fernandes; de Souza, Luiz Jos; Cunha, Rivaldo Venncio; de-Oliveira-Pinto, Luzia Maria

    2014-01-01

    In dengue virus (DENV) infection, complement system (CS) activation appears to have protective and pathogenic effects. In severe dengue fever (DF), the levels of DENV non-structural-1 protein and of the products of complement activation, including C3a, C5a and SC5b-9, are higher before vascular leakage occurs, supporting the hypothesis that complement activation contributes to unfavourable outcomes. The clinical manifestations of DF range from asymptomatic to severe and even fatal. Here, we aimed to characterise CS by their receptors or activation product, in vivo in DF patients and in vitro by DENV-2 stimulation on monocytes. In comparison with healthy controls, DF patients showed lower expression of CR3 (CD11b), CR4 (CD11c) and, CD59 on monocytes. The DF patients who were high producers of SC5b-9 were also those that showed more pronounced bleeding or vascular leakage. Those findings encouraged us to investigate the role of CS in vitro, using monocytes isolated from healthy subjects. Prior blocking with CR3 alone (CD11b) or CR3 (CD11b/CD18) reduced viral infection, as quantified by the levels of intracellular viral antigen expression and soluble DENV non-structural viral protein. However, we found that CR3 alone (CD11b) or CR3 (CD11b/CD18) blocking did not influence major histocompatibility complex presentation neither active caspase-1 on monocytes, thus probably ruling out inflammasome-related mechanisms. Although it did impair the secretion of tumour necrosis factor alpha and interferon alpha. Our data provide strategies of blocking CR3 (CD11b) pathways could have implications for the treatment of viral infection by antiviral-related mechanisms. PMID:25061945

  10. The alternative complement pathway aids in vascular regression during the early stages of a murine model of proliferative retinopathy

    PubMed Central

    Kim, Clifford; Smith, Kaylee E.; Castillejos, Alexandra; Diaz-Aguilar, Daniel; Saint-Geniez, Magali; Connor, Kip M.

    2016-01-01

    Proliferative retinopathic diseases often progress in 2 phases: initial regression of retinal vasculature (phase 1) followed by subsequent neovascularization (NV) (phase 2). The immune system has been shown to aid in vascular pruning in such retinopathies; however, little is known about the role of the alternative complement pathway in the initial vascular regression phase. Using a mouse model of oxygen-induced retinopathy (OIR), we observed that alternative complement pathwaydeficient mice (Fb?/?) exhibited a mild decrease in vascular loss at postnatal day (P)8 compared with age- and strain-matched controls (P = 0.035). Laser capture microdissection was used to isolate the retinal blood vessels. Expression of the complement inhibitors Cd55 and Cd59 was significantly decreased in blood vessels isolated from hyperoxic retinas compared with those from normoxic control mice. Vegf expression was measured at P8 and found to be significantly lower in OIR mice than in normoxic control mice (P = 0.0048). Further examination of specific Vegf isoform expression revealed a significant decrease in Vegf120 (P = 0.00032) and Vegf188 (P = 0.0092). In conjunction with the major modulating effects of Vegf during early retinal vascular development, our data suggest a modest involvement of the alternative complement pathway in targeting vessels for regression in the initial vaso-obliteration stage of OIR.Kim, C., Smith, K. E., Castillejos, A., Diaz-Aguilar, D., Saint-Geniez, M., Connor, K. M. The alternative complement pathway aids in vascular regression during the early stages of a murine model of proliferative retinopathy. PMID:26631482

  11. The indoleamine 2,3-dioxygenase pathway controls complement-dependent enhancement of chemo-radiation therapy against murine glioblastoma

    PubMed Central

    2014-01-01

    Background Indoleamine 2,3-dioxygenase (IDO) is an enzyme with immune-suppressive properties that is commonly exploited by tumors to evade immune destruction. Anti-tumor T cell responses can be initiated in solid tumors, but are immediately suppressed by compensatory upregulation of immunological checkpoints, including IDO. In addition to these known effects on the adaptive immune system, we previously showed widespread, T cell-dependent complement deposition during allogeneic fetal rejection upon maternal treatment with IDO-blockade. We hypothesized that IDO protects glioblastoma from the full effects of chemo-radiation therapy by preventing vascular activation and complement-dependent tumor destruction. Methods To test this hypothesis, we utilized a syngeneic orthotopic glioblastoma model in which GL261 glioblastoma tumor cells were stereotactically implanted into the right frontal lobes of syngeneic mice. These mice were treated with IDO-blocking drugs in combination with chemotherapy and radiation therapy. Results Pharmacologic inhibition of IDO synergized with chemo-radiation therapy to prolong survival in mice bearing intracranial glioblastoma tumors. We now show that pharmacologic or genetic inhibition of IDO allowed chemo-radiation to trigger widespread complement deposition at sites of tumor growth. Chemotherapy treatment alone resulted in collections of perivascular leukocytes within tumors, but no complement deposition. Adding IDO-blockade led to upregulation of VCAM-1 on vascular endothelium within the tumor microenvironment, and further adding radiation in the presence of IDO-blockade led to widespread deposition of complement. Mice genetically deficient in complement component C3 lost all of the synergistic effects of IDO-blockade on chemo-radiation-induced survival. Conclusions Together these findings identify a novel mechanistic link between IDO and complement, and implicate complement as a major downstream effector mechanism for the beneficial effect of IDO-blockade after chemo-radiation therapy. We speculate that this represents a fundamental pathway by which the tumor regulates intratumoral vascular activation and protects itself from immune-mediated tumor destruction. PMID:25054064

  12. The alternative complement pathway control protein H binds to immune complexes and serves their detection

    SciTech Connect

    Nydegger, U.E.; Corvetta, A.; Spaeth, P.J.; Spycher, M.

    1983-01-01

    During solubilization of immune complexes C3b becomes fixed to the immunoglobulin part and serves as a receptor for the alternative complement pathway control protein H. The H-C3b immune complex interaction can be made detectable using 4% polyethyleneglycol to separate free from bound /sup 125/I-H. Tetanus toxoid (Te)/anti-Te complexes kept soluble with fresh serum and containing 125 IU of specific antibody bound 18% of /sup 125/I-H; when fresh serum was chelated with 10 mM EDTA, /sup 125/I-H binding was only 5%. On sucrose density gradients, the H-binding material sedimented in the range of 12 to 30 S. In 36 serum samples from rheumatoid arthritis (RA) patients and in 12 serum samples from patients with systemic lupus erythematosus (SLE), /sup 125/I-H binding was significantly elevated to 9.5 +/- 4.7% (mean +/- 1 SD) and 13.3 +/- 5.6%, respectively, while /sup 125/I-H binding by 36 normal human sera was 4 +/- 2%. RA samples (17/36, 47%) and SLE samples (9/12, 75%) had H-binding values increased by more than 2 SD above the normal mean. The serum samples were also assessed for conglutinin- and C1q-binding activities; a significant correlation between H and C1q binding was observed (P less than 0.001); there was no correlation between H and conglutinin binding. Although binding to immune complexes through its interaction with C3b, H clearly detects a population of complexes other than conglutinin, thus expanding the possibilities of further characterizing pathological complexes.

  13. Musca domestica larva lectin induces apoptosis in BEL-7402 cells through a Ca(2+)/JNK-mediated mitochondrial pathway.

    PubMed

    Wang, Chun-Ling; Xia, Yan; Nie, Jian-Zeng; Zhou, Minghui; Zhang, Rong-Ping; Niu, Li-Li; Hou, Li-Hua; Cao, Xiao-Hong

    2013-06-01

    Although Musca domestica larvae lectin (MLL) is able to inhibit cancer cell proliferation and to induce cancer cell apoptosis, the molecular mechanism(s) responsible for these processes remain elusive. In the current study, the signaling network underlying the MLL-induced apoptosis of human hepatoma BEL-7402 cell was investigated. Our data found out that MLL causes a sustained increase of the intracellular Ca(2+) and this process was prevented by the intracellular calcium chelator, BAPTA-AM, suggesting the involvement of intracellular Ca(2+) in MLL-induced cell apoptosis. MLL also causes the production of reactive oxygen species and elevates the phosphorylation status of JNK, processes associated with the increased cytoplasmic Ca(2+). The mitochondrial permeability transition pore (MPTP) opening study showed that MLL treatment of BEL-7402 cells results in the opening of MPTP and a reduction of mitochondrial transmembrane potential. In such condition, cytochrome-c was detected to be released from mitochondria to cytoplasm through the MPTP. This eventually activates caspase-3 and thus results in apoptosis of the tested BEL-7402 cells. According to a comprehensive review of all the evidence, it is concluded that MLL induces apoptosis of BEL-7402 cells through a Ca(2+)/JNK-mediated MPTP pathway. PMID:23247835

  14. Molecules Great and Small: The Complement System.

    PubMed

    Mathern, Douglas R; Heeger, Peter S

    2015-09-01

    The complement cascade, traditionally considered an effector arm of innate immunity required for host defense against pathogens, is now recognized as a crucial pathogenic mediator of various kidney diseases. Complement components produced by the liver and circulating in the plasma undergo activation through the classical and/or mannose-binding lectin pathways to mediate anti-HLA antibody-initiated kidney transplant rejection and autoantibody-initiated GN, the latter including membranous glomerulopathy, antiglomerular basement membrane disease, and lupus nephritis. Inherited and/or acquired abnormalities of complement regulators, which requisitely limit restraint on alternative pathway complement activation, contribute to the pathogenesis of the C3 nephropathies and atypical hemolytic uremic syndrome. Increasing evidence links complement produced by endothelial cells and/or tubular cells to the pathogenesis of kidney ischemia-reperfusion injury and progressive kidney fibrosis. Data emerging since the mid-2000s additionally show that immune cells, including T cells and antigen-presenting cells, produce alternative pathway complement components during cognate interactions. The subsequent local complement activation yields production of the anaphylatoxins C3a and C5a, which bind to their respective receptors (C3aR and C5aR) on both partners to augment effector T-cell proliferation and survival, while simultaneously inhibiting regulatory T-cell induction and function. This immune cell-derived complement enhances pathogenic alloreactive T-cell immunity that results in transplant rejection and likely contributes to the pathogenesis of other T cell-mediated kidney diseases. C5a/C5aR ligations on neutrophils have additionally been shown to contribute to vascular inflammation in models of ANCA-mediated renal vasculitis. New translational immunology efforts along with the development of pharmacologic agents that block human complement components and receptors now permit testing of the intriguing concept that targeting complement in patients with an assortment of kidney diseases has the potential to abrogate disease progression and improve patient health. PMID:25568220

  15. Alternative and classical complement pathway activity in sera from colostrum-fed and colostrum-deprived neonatal pigs.

    PubMed Central

    Renshaw, H W; Gilmore, R J

    1980-01-01

    Haemolytic assays were used to compare alternative and classical complement (C) pathway activities in sera obtained from neonatal pigs reared on porcine colostrum, bovine colostrum or an immunoglobulin-free synthetic diet. Dramatic increases in immunoglobulin concentrations were noted in the colostrum-fed animals during the first day of life, but there was not a concurrent, marked increase in either classical or alternative C pathway activity. Whether fed on homologous or heterologous colostrum, neonatal pigs had a similar gradual increase in alternative and classical C pathway activity in the post-natal period. If direct passive absorption of C components occurs in newborn pigs, it has only a minor influence on functional levels of alternative and classical C pathway activity in their sera. In pigs fed homologous and heterologous colostrum there was, respectively, an 83% and 80% increase in classical pathway activity, but only a 13% and 12% increase in alternative pathway activity during the first 3 days of life. Pigs fed the immunoglobulin-free synthetic diet had a 37% increase in classical C and a 24% increase in alternative C pathway activity. Part of the increase in classical C pathway activity in the post-natal period may be caused by a stimulating factor in colostrum. Most if not all of the increase in alternative C pathway activity and some of the increase in classical C pathway activity is most likely caused by normal humoral homeostatic mechanisms in the neonatal pig. PMID:7429550

  16. Molecular mechanism of anticancer effect of Sclerotium rolfsii lectin in HT29 cells involves differential expression of genes associated with multiple signaling pathways: A microarray analysis.

    PubMed

    Barkeer, Srikanth; Guha, Nilanjan; Hothpet, Vishwanathreddy; Saligrama Adavigowda, Deepak; Hegde, Prajna; Padmanaban, Arunkumar; Yu, Lu-Gang; Swamy, Bale M; Inamdar, Shashikala R

    2015-12-01

    Sclerotium rolfsii lectin (SRL) is a lectin isolated from fungus S. rolfsii and has high binding specificity toward the oncofetal Thomsen-Friedenreich carbohydrate antigen (Gal?1-3GalNAc-?-O-Ser/Thr, T or TF), which is expressed in more than 90% of human cancers. Our previous studies have shown that binding of SRL to human colon, breast and ovarian cancer cells induces cell apoptosis in vitro and suppresses tumor growth in vivo. This study investigated the SRL-mediated cell signaling in human colon cancer HT29 cells by mRNA and miRNA microarrays. It was found that SRL treatment results in altered expression of several hundred molecules including mitogen-activated protein kinase (MAPK) and c-JUN-associated, apoptosis-associated and cell cycle and DNA replication-associated signaling molecules. Pathway analysis using GeneSpring 12.6.1 revealed that SRL treatment induces changes of MAPK and c-JUN-associated signaling pathways as early as 2 h while changes of cell cycle, DNA replication and apoptosis pathways were significantly affected only after 24 h. A significant change of cell miRNA expression was also observed after 12 h treatment of the cells with SRL. These changes were further validated by quantitative real time polymerase chain reaction and immunoblotting. This study thus suggests that the presence of SRL affects multiple signaling pathways in cancer cells with early effects on cell proliferation pathways associated with MAPK and c-JUN, followed by miRNA-associated cell activity and apoptosis. This provides insight information into the molecular mechanism of the anticancer activity of this fungal lectin. PMID:26347523

  17. Mutations in Complement Factor H Impair Alternative Pathway Regulation on Mouse Glomerular Endothelial Cells in Vitro.

    PubMed

    Loeven, Markus A; Rops, Angelique L; Lehtinen, Markus J; van Kuppevelt, Toin H; Daha, Mohamed R; Smith, Richard J; Bakker, Marinka; Berden, Jo H; Rabelink, Ton J; Jokiranta, T Sakari; van der Vlag, Johan

    2016-03-01

    Complement factor H (FH) inhibits complement activation and interacts with glomerular endothelium via its complement control protein domains 19 and 20, which also recognize heparan sulfate (HS). Abnormalities in FH are associated with the renal diseases atypical hemolytic uremic syndrome and dense deposit disease and the ocular disease age-related macular degeneration. Although FH systemically controls complement activation, clinical phenotypes selectively manifest in kidneys and eyes, suggesting the presence of tissue-specific determinants of disease development. Recent results imply the importance of tissue-specifically expressed, sulfated glycosaminoglycans (GAGs), like HS, in determining FH binding to and activity on host tissues. Therefore, we investigated which GAGs mediate human FH and recombinant human FH complement control proteins domains 19 and 20 (FH19-20) binding to mouse glomerular endothelial cells (mGEnCs) in ELISA. Furthermore, we evaluated the functional defects of FH19-20 mutants during complement activation by measuring C3b deposition on mGEnCs using flow cytometry. FH and FH19-20 bound dose-dependently to mGEnCs and TNF-? treatment increased binding of both proteins, whereas heparinase digestion and competition with heparin/HS inhibited binding. Furthermore, 2-O-, and 6-O-, but not N-desulfation of heparin, significantly increased the inhibitory effect on FH19-20 binding to mGEnCs. Compared with wild type FH19-20, atypical hemolytic uremic syndrome-associated mutants were less able to compete with FH in normal human serum during complement activation on mGEnCs, confirming their potential glomerular pathogenicity. In conclusion, our study shows that FH and FH19-20 binding to glomerular endothelial cells is differentially mediated by HS but not other GAGs. Furthermore, we describe a novel, patient serum-independent competition assay for pathogenicity screening of FH19-20 mutants. PMID:26728463

  18. Activation of the Complement Classical Pathway (C1q Binding) by Mesophilic Aeromonas hydrophila Outer Membrane Protein

    PubMed Central

    Merino, Susana; Nogueras, Maria Mercedes; Aguilar, Alicia; Rubires, Xavier; Albert, Sebastian; Bened, Vicente Javier; Toms, Juan M.

    1998-01-01

    The mechanism of killing of Aeromonas hydrophila serum-sensitive strains in nonimmune serum by the complement classical pathway has been studied. The bacterial cell surface component that binds C1q more efficiently was identified as a major outer membrane protein of 39 kDa, presumably the porin II described by D. Jeanteur, N. Gletsu, F. Pattus, and J. T. Buckley (Mol. Microbiol. 6:33553363, 1992), of these microorganisms. We have demonstrated that the purified form of porin II binds C1q and activates the classical pathway in an antibody-independent manner, with the subsequent consumption of C4 and reduction of the serum total hemolytic activity. Activation of the classical pathway has been observed in human nonimmune serum and agammaglobulinemic serum (both depleted of factor D). Binding of C1q to other components of the bacterial outer membrane, in particular to rough lipopolysaccharide, could not be demonstrated. Activation of the classical pathway by this lipopolysaccharide was also much less efficient than activation by the outer membrane protein. The strains possessing O-antigen lipopolysaccharide bind less C1q than the serum-sensitive strains, because the outer membrane protein is less accessible, and are resistant to complement-mediated killing. Finally, a similar or identical outer membrane protein (presumably porin II) that binds C1q was shown to be present in strains from the most common mesophilic Aeromonas O serogroups. PMID:9673268

  19. Dissecting the complement pathway in hepatic injury and regeneration with a novel protective strategy

    PubMed Central

    Marshall, Keely M.; Zhong, Zhi; Atkinson, Carl

    2014-01-01

    Liver resection is commonly performed under ischemic conditions, resulting in two types of insult to the remnant liver: ischemia reperfusion injury (IRI) and loss of liver mass. Complement inhibition is recognized as a potential therapeutic modality for IRI, but early complement activation products are also essential for liver regeneration. We describe a novel site-targeted murine complement inhibitor, CR2-CD59, which specifically inhibits the terminal membrane attack complex (MAC), and we use this protein to investigate the complement-dependent balance between liver injury and regeneration in a clinical setting of pharmacological inhibition. CR2-CD59 did not impact in vivo generation of C3 and C5 activation products but was as effective as the C3 activation inhibitor CR2-Crry at ameliorating hepatic IRI, indicating that the MAC is the principle mediator of hepatic IRI. Furthermore, unlike C3 or C5 inhibition, CR2-CD59 was not only protective but significantly enhanced hepatocyte proliferation after partial hepatectomy, including when combined with ischemia and reperfusion. Remarkably, CR2-CD59 also enhanced regeneration after 90% hepatectomy and improved long-term survival from 0 to 70%. CR2-CD59 functioned by increasing hepatic TNF and IL-6 levels with associated STAT3 and Akt activation, and by preventing mitochondrial depolarization and allowing recovery of ATP stores. PMID:25113972

  20. C1- inactivator: its efficiency as a regulator of classical complement pathway activation by soluble IgG aggregates.

    PubMed Central

    Doekes, G; van Es, L A; Daha, M R

    1983-01-01

    The role of C1- inactivator (C1(-)-In) during activation of the classical complement pathway by soluble immune complexes was studied using purified human complement components C1, C4 and C1(-)-In, and stabilized soluble aggregates of normal human IgG as a model for soluble immune complexes. The C4-consuming ability that could be generated by incubation of precursor C1 with IgG aggregates was abolished completely by the presence of a large excess of C1(-)-In during the C1 activation step. Kinetic studies confirmed that this inhibition was due to a second-order reaction between C1- and C1(-)-In resulting in the irreversible inactivation of C1-. When aggregates of various sizes were enabled to induce C4 conversion in mixtures of C1, C4 and a variable concentration of C1(-)-In, the presence of C1(-)-In had two effects. Firstly, the efficiency of the aggregates in causing C4 consumption was reduced remarkably. At a C1(-)-In:C1 ratio of 8, which can be found in normal human serum, approximately eight to ten times as many aggregates were required for a given level of C4 consumption as when no C1(-)-In was present. Secondly, C1(-)-In diminished the maximum C4 consumption that could be achieved, especially with smaller aggregates. Thus, a complete or partial C1(-)-In deficiency probably facilitates complement activation by soluble immune complexes in two ways: it may enhance the efficiency of classical pathway activation by all C1-activating complexes, and it may enable small complexes, which normally cannot overcome the C1(-)-In barrier, to activate the classical pathway to the C4 level. PMID:6852866

  1. Inherited mitochondrial DNA variants can affect complement, inflammation and apoptosis pathways: insights into mitochondrial-nuclear interactions.

    PubMed

    Kenney, M Cristina; Chwa, Marilyn; Atilano, Shari R; Falatoonzadeh, Payam; Ramirez, Claudio; Malik, Deepika; Tarek, Mohamed; Cceres-del-Carpio, Javier; Nesburn, Anthony B; Boyer, David S; Kuppermann, Baruch D; Vawter, Marquis; Jazwinski, S Michal; Miceli, Michael; Wallace, Douglas C; Udar, Nitin

    2014-07-01

    Age-related macular degeneration (AMD) is the leading cause of vision loss in developed countries. While linked to genetic polymorphisms in the complement pathway, there are many individuals with high risk alleles that do not develop AMD, suggesting that other 'modifiers' may be involved. Mitochondrial (mt) haplogroups, defined by accumulations of specific mtDNA single nucleotide polymorphisms (SNPs) which represent population origins, may be one such modifier. J haplogroup has been associated with high risk for AMD while the H haplogroup is protective. It has been difficult to assign biological consequences for haplogroups so we created human ARPE-19 cybrids (cytoplasmic hybrids), which have identical nuclei but mitochondria of either J or H haplogroups, to investigate their effects upon bioenergetics and molecular pathways. J cybrids have altered bioenergetic profiles compared with H cybrids. Q-PCR analyses show significantly lower expression levels for seven respiratory complex genes encoded by mtDNA. J and H cybrids have significantly altered expression of eight nuclear genes of the alternative complement, inflammation and apoptosis pathways. Sequencing of the entire mtDNA was carried out for all the cybrids to identify haplogroup and non-haplogroup defining SNPs. mtDNA can mediate cellular bioenergetics and expression levels of nuclear genes related to complement, inflammation and apoptosis. Sequencing data suggest that observed effects are not due to rare mtDNA variants but rather the combination of SNPs representing the J versus H haplogroups. These findings represent a paradigm shift in our concepts of mt-nuclear interactions. PMID:24584571

  2. Inherited mitochondrial DNA variants can affect complement, inflammation and apoptosis pathways: insights into mitochondrial–nuclear interactions

    PubMed Central

    Cristina Kenney, M.; Chwa, Marilyn; Atilano, Shari R.; Falatoonzadeh, Payam; Ramirez, Claudio; Malik, Deepika; Tarek, Mohamed; Cáceres-del-Carpio, Javier; Nesburn, Anthony B.; Boyer, David S.; Kuppermann, Baruch D.; Vawter, Marquis; Michal Jazwinski, S.; Miceli, Michael; Wallace, Douglas C.; Udar, Nitin

    2014-01-01

    Age-related macular degeneration (AMD) is the leading cause of vision loss in developed countries. While linked to genetic polymorphisms in the complement pathway, there are many individuals with high risk alleles that do not develop AMD, suggesting that other ‘modifiers’ may be involved. Mitochondrial (mt) haplogroups, defined by accumulations of specific mtDNA single nucleotide polymorphisms (SNPs) which represent population origins, may be one such modifier. J haplogroup has been associated with high risk for AMD while the H haplogroup is protective. It has been difficult to assign biological consequences for haplogroups so we created human ARPE-19 cybrids (cytoplasmic hybrids), which have identical nuclei but mitochondria of either J or H haplogroups, to investigate their effects upon bioenergetics and molecular pathways. J cybrids have altered bioenergetic profiles compared with H cybrids. Q-PCR analyses show significantly lower expression levels for seven respiratory complex genes encoded by mtDNA. J and H cybrids have significantly altered expression of eight nuclear genes of the alternative complement, inflammation and apoptosis pathways. Sequencing of the entire mtDNA was carried out for all the cybrids to identify haplogroup and non-haplogroup defining SNPs. mtDNA can mediate cellular bioenergetics and expression levels of nuclear genes related to complement, inflammation and apoptosis. Sequencing data suggest that observed effects are not due to rare mtDNA variants but rather the combination of SNPs representing the J versus H haplogroups. These findings represent a paradigm shift in our concepts of mt–nuclear interactions. PMID:24584571

  3. Impact of Mannose-Binding Lectin Deficiency on Radiocontrast-Induced Renal Dysfunction

    PubMed Central

    Osthoff, Michael; Trendelenburg, Marten

    2013-01-01

    Contrast-induced nephropathy (CIN) is the third leading cause of acute renal failure in hospitalized patients. Endothelial dysfunction, renal medullary ischemia, and tubular toxicity are regarded as the most important factors in the pathogenesis of CIN. Mannose-binding lectin (MBL), a pattern recognition protein of the lectin pathway of complement, has been found to aggravate and mediate tissue damage during experimental renal ischemia/reperfusion (I/R) injury which was alleviated by inhibition with C1 inhibitor, a potent MBL, and lectin pathway inhibitor. In this paper, we highlight the potential role of MBL in the pathogenesis of human CIN. In experimental I/R models, MBL was previously found to induce tubular cell death independent of the complement system. In addition, after binding to vascular endothelial cells, MBL and its associated serine proteases were able to trigger a proinflammatory reaction and contribute to endothelial dysfunction. In humans, urinary MBL was increased after administration of contrast media and in individuals with CIN. Moreover, individuals with normal/high MBL levels were at increased risk to develop radiocontrast-induced renal dysfunction. Hence, MBL and the lectin pathway seem to be a promising target given that a licensed, powerful, human recombinant inhibitor exits to be added to the scarce armamentarium currently available for prophylaxis of CIN. PMID:24386641

  4. The lectin concanavalin-A signals MT1-MMP catalytic independent induction of COX-2 through an IKKgamma/NF-kappaB-dependent pathway.

    PubMed

    Sina, Asmaa; Proulx-Bonneau, Sbastien; Roy, Alain; Poliquin, Laurent; Cao, Jian; Annabi, Borhane

    2010-03-01

    The lectin from Canavalia ensiformis (Concanavalin-A, ConA), one of the most abundant lectins known, enables one to mimic biological lectin/carbohydrate interactions that regulate extracellular matrix protein recognition. As such, ConA is known to induce membrane type-1 matrix metalloproteinase (MT1-MMP) which expression is increased in brain cancer. Given that MT1-MMP correlated to high expression of cyclooxygenase (COX)-2 in gliomas with increasing histological grade, we specifically assessed the early proinflammatory cellular signaling processes triggered by ConA in the regulation of COX-2. We found that treatment with ConA or direct overexpression of a recombinant MT1-MMP resulted in the induction of COX-2 expression. This increase in COX-2 was correlated with a concomitant decrease in phosphorylated AKT suggestive of cell death induction, and was independent of MT1-MMP's catalytic function. ConA- and MT1-MMP-mediated intracellular signaling of COX-2 was also confirmed in wild-type and in Nuclear Factor-kappaB (NF-kappaB) p65(-/-) mutant mouse embryonic fibroblasts (MEF), but was abrogated in NF-kappaB1 (p50)(-/-) and in I kappaB kinase (IKK) gamma(-/-) mutant MEF cells. Collectively, our results highlight an IKK/NF-kappaB-dependent pathway linking MT1-MMP-mediated intracellular signaling to the induction of COX-2. That signaling pathway could account for the inflammatory balance responsible for the therapy resistance phenotype of glioblastoma cells, and prompts for the design of new therapeutic strategies that target cell surface carbohydrate structures and MT1-MMP-mediated signaling. Concise summary Concanavalin-A (ConA) mimics biological lectin/carbohydrate interactions that regulate the proinflammatory phenotype of cancer cells through yet undefined signaling. Here we highlight an IKK/NF-kappaB-dependent pathway linking MT1-MMP-mediated intracellular signaling to the induction of cyclooxygenase-2, and that could be responsible for the therapy resistance phenotype of glioblastoma cells. PMID:20195390

  5. Genetic Association of the Porcine C9 Complement Component with Hemolytic Complement Activity

    PubMed Central

    Khoa, D. V. A.; Wimmers, K.

    2015-01-01

    The complement system is a part of the natural immune regulation mechanism against invading pathogens. Complement activation from three different pathways (classical, lectin, and alternative) leads to the formation of C5-convertase, an enzyme for cleavage of C5 into C5a and C5b, followed by C6, C7, C8, and C9 in membrane attack complex. The C9 is the last complement component of the terminal lytic pathway, which plays an important role in lysis of the target cells depending on its self-polymerization to form transmembrane channels. To address the association of C9 with traits related to disease resistance, the complete porcine C9 cDNA was comparatively sequenced to detect single nucleotide polymorphisms (SNPs) in pigs of the breeds Hampshire (HS), Duroc (DU), Berlin miniature pig (BMP), German Landrace (LR), Pietrain (PIE), and Muong Khuong (Vietnamese potbelly pig). Genotyping was performed in 417 F2 animals of a resource population (DUMI: DU×BMP) that were vaccinated with Mycoplasma hyopneumoniae, Aujeszky diseases virus and porcine respiratory and reproductive syndrome virus at 6, 14 and 16 weeks of age, respectively. Two SNPs were detected within the third exon. One of them has an amino acid substitution. The European porcine breeds (LR and PIE) show higher allele frequency of these SNPs than Vietnamese porcine breed (MK). Association of the substitution SNP with hemolytic complement activity indicated statistically significant differences between genotypes in the classical pathway but not in the alternative pathway. The interactions between eight time points of measurement of complement activity before and after vaccinations and genotypes were significantly different. The difference in hemolytic complement activity in the both pathways depends on genotype, kind of vaccine, age and the interaction to the other complement components. These results promote the porcine C9 (pC9) as a candidate gene to improve general animal health in the future. PMID:26194222

  6. Genetic Association of the Porcine C9 Complement Component with Hemolytic Complement Activity.

    PubMed

    Khoa, D V A; Wimmers, K

    2015-09-01

    The complement system is a part of the natural immune regulation mechanism against invading pathogens. Complement activation from three different pathways (classical, lectin, and alternative) leads to the formation of C5-convertase, an enzyme for cleavage of C5 into C5a and C5b, followed by C6, C7, C8, and C9 in membrane attack complex. The C9 is the last complement component of the terminal lytic pathway, which plays an important role in lysis of the target cells depending on its self-polymerization to form transmembrane channels. To address the association of C9 with traits related to disease resistance, the complete porcine C9 cDNA was comparatively sequenced to detect single nucleotide polymorphisms (SNPs) in pigs of the breeds Hampshire (HS), Duroc (DU), Berlin miniature pig (BMP), German Landrace (LR), Pietrain (PIE), and Muong Khuong (Vietnamese potbelly pig). Genotyping was performed in 417 F2 animals of a resource population (DUMI: DUBMP) that were vaccinated with Mycoplasma hyopneumoniae, Aujeszky diseases virus and porcine respiratory and reproductive syndrome virus at 6, 14 and 16 weeks of age, respectively. Two SNPs were detected within the third exon. One of them has an amino acid substitution. The European porcine breeds (LR and PIE) show higher allele frequency of these SNPs than Vietnamese porcine breed (MK). Association of the substitution SNP with hemolytic complement activity indicated statistically significant differences between genotypes in the classical pathway but not in the alternative pathway. The interactions between eight time points of measurement of complement activity before and after vaccinations and genotypes were significantly different. The difference in hemolytic complement activity in the both pathways depends on genotype, kind of vaccine, age and the interaction to the other complement components. These results promote the porcine C9 (pC9) as a candidate gene to improve general animal health in the future. PMID:26194222

  7. Identification of human mannose binding lectin (MBL) recognition sites for novel inhibitory antibodies.

    PubMed

    Zhao, Hui; Wakamiya, Nobutaka; Suzuki, Yasuhiko; Hamonko, Matthew T; Stahl, Gregory L

    2002-02-01

    Mannose binding lectin (MBL) binding initiates activation of the lectin complement pathway. Recent studies from our laboratory have demonstrated that MBL-dependent complement activation mediates cellular injury following oxidative stress in vivo and in vitro. A panel of novel inhibitory monoclonal antibodies (MAbs) against MBL (e.g., MAb 3F8, 2A9, and hMBL1.2) has been developed that inhibit MBL binding and lectin pathway activation. Here, we further characterized the interactions of these MAbs and their Fab fragments to MBL. Whole MAbs or their Fab fragments bound to MBL with relatively high affinity. Fab fragments of 3F8 were functionally effective in inhibiting MBL-dependent complement activation, however, steric hindrance of MAb 2A9 was essential for inhibition of MBL-dependent complement activation. We identified the hinge region, and residues EDCVLLL within the carbohydrate recognition domain of MBL as the recognition sites for MAb 3F8 and 2A9, respectively. The interaction of MAbs (e.g., 3F8 and 2A9) to MBL was dependent on the conformation of their recognition sites. These findings demonstrate that MBL binding can be inhibited by at least two separate and independent mechanisms. PMID:11991814

  8. Soluble Collectin-12 (CL-12) Is a Pattern Recognition Molecule Initiating Complement Activation via the Alternative Pathway.

    PubMed

    Ma, Ying Jie; Hein, Estrid; Munthe-Fog, Lea; Skjoedt, Mikkel-Ole; Bayarri-Olmos, Rafael; Romani, Luigina; Garred, Peter

    2015-10-01

    Soluble defense collagens including the collectins play important roles in innate immunity. Recently, a new member of the collectin family named collectin-12 (CL-12 or CL-P1) has been identified. CL-12 is highly expressed in umbilical cord vascular endothelial cells as a transmembrane receptor and may recognize certain bacteria and fungi, leading to opsonophagocytosis. However, based on its structural and functional similarities with soluble collectins, we hypothesized the existence of a fluid-phase analog of CL-12 released from cells, which may function as a soluble pattern-recognition molecule. Using recombinant CL-12 full length or CL-12 extracellular domain, we determined the occurrence of soluble CL-12 shed from in vitro cultured cells. Western blot showed that soluble recombinant CL-12 migrated with a band corresponding to ? 120 kDa under reducing conditions, whereas under nonreducing conditions it presented multimeric assembly forms. Immunoprecipitation and Western blot analysis of human umbilical cord plasma enabled identification of a natural soluble form of CL-12 having an electrophoretic mobility pattern close to that of shed soluble recombinant CL-12. Soluble CL-12 could recognize Aspergillus fumigatus partially through the carbohydrate-recognition domain in a Ca(2+)-independent manner. This led to activation of the alternative pathway of complement exclusively via association with properdin on A. fumigatus as validated by detection of C3b deposition and formation of the terminal complement complex. These results demonstrate the existence of CL-12 in a soluble form and indicate a novel mechanism by which the alternative pathway of complement may be triggered directly by a soluble pattern-recognition molecule. PMID:26290605

  9. Protein load impairs factor H binding promoting complement-dependent dysfunction of proximal tubular cells.

    PubMed

    Buelli, Simona; Abbate, Mauro; Morigi, Marina; Moioli, Daniela; Zanchi, Cristina; Noris, Marina; Zoja, Carla; Pusey, Charles D; Zipfel, Peter F; Remuzzi, Giuseppe

    2009-05-01

    Intrarenal complement activation plays an important role in the progression of chronic kidney disease. A key target of the activated complement cascade is the proximal tubule, a site where abnormally filtered plasma proteins and complement factors combine to promote injury. This study determined whether protein overloading of human proximal tubular cells (HK-2) in culture enhances complement activation by impairing complement regulation. Addition of albumin or transferrin to the cells incubated with diluted human serum as a source of complement caused increased apical C3 deposition. Soluble complement receptor-1 (an inhibitor of all 3 activation pathways) blocked complement deposition while the classical and lectin pathway inhibitor, magnesium chloride-EGTA, was, ineffective. Media containing albumin as well as complement had additive proinflammatory effects as shown by increased fractalkine and transforming growth factor-beta mRNA expression. This paralleled active C3 and C5b-9 generations, effects not shared by transferrin. Factor H, one of the main natural inhibitors of the alternative pathway, binds to heparan sulfate proteoglycans. Both the density of heparan sulfate and factor H binding were reduced with protein loading, thereby enhancing the albumin- and serum-dependent complement activation potential. Thus, protein overload reduces the ability of the tubule cell to bind factor H and counteract complement activation, effects instrumental to renal disease progression. PMID:19242507

  10. Ouabain rescues rat nephrogenesis during intrauterine growth restriction by regulating the complement and coagulation cascades and calcium signaling pathway.

    PubMed

    Chen, L; Yue, J; Han, X; Li, J; Hu, Y

    2016-02-01

    Intrauterine growth restriction (IUGR) is associated with a reduction in the numbers of nephrons in neonates, which increases the risk of hypertension. Our previous study showed that ouabain protects the development of the embryonic kidney during IUGR. To explore this molecular mechanism, IUGR rats were induced by protein and calorie restriction throughout pregnancy, and ouabain was delivered using a mini osmotic pump. RNA sequencing technology was used to identify the differentially expressed genes (DEGs) of the embryonic kidneys. DEGs were submitted to the Database for Annotation and Visualization and Integrated Discovery, and gene ontology enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were conducted. Maternal malnutrition significantly reduced fetal weight, but ouabain treatment had no significant effect on body weight. A total of 322 (177 upregulated and 145 downregulated) DEGs were detected between control and the IUGR group. Meanwhile, 318 DEGs were found to be differentially expressed (180 increased and 138 decreased) between the IUGR group and the ouabain-treated group. KEGG pathway analysis indicated that maternal undernutrition mainly disrupts the complement and coagulation cascades and the calcium signaling pathway, which could be protected by ouabain treatment. Taken together, these two biological pathways may play an important role in nephrogenesis, indicating potential novel therapeutic targets against the unfavorable effects of IUGR. PMID:26442628

  11. Complement System Part I – Molecular Mechanisms of Activation and Regulation

    PubMed Central

    Merle, Nicolas S.; Church, Sarah Elizabeth; Fremeaux-Bacchi, Veronique; Roumenina, Lubka T.

    2015-01-01

    Complement is a complex innate immune surveillance system, playing a key role in defense against pathogens and in host homeostasis. The complement system is initiated by conformational changes in recognition molecular complexes upon sensing danger signals. The subsequent cascade of enzymatic reactions is tightly regulated to assure that complement is activated only at specific locations requiring defense against pathogens, thus avoiding host tissue damage. Here, we discuss the recent advances describing the molecular and structural basis of activation and regulation of the complement pathways and their implication on physiology and pathology. This article will review the mechanisms of activation of alternative, classical, and lectin pathways, the formation of C3 and C5 convertases, the action of anaphylatoxins, and the membrane-attack-complex. We will also discuss the importance of structure–function relationships using the example of atypical hemolytic uremic syndrome. Lastly, we will discuss the development and benefits of therapies using complement inhibitors. PMID:26082779

  12. Complement and membrane-bound complement regulatory proteins as biomarkers and therapeutic targets for autoimmune inflammatory disorders, RA and SLE.

    PubMed

    Das, Nibhriti

    2015-11-01

    Complement system is a major effecter system of the innate immunity that bridges with adaptive immunity. The system consists of about 40 humoral and cell surface proteins that include zymogens, receptors and regulators. The zymogens get activated in a cascade fashion by antigen-antibody complex, antigen alone or by polymannans, respectively, by the classical, alternative and mannose binding lectin (MBL) pathways. The ongoing research on complement regulators and complement receptors suggest key role of these proteins in the initiation, regulation and effecter mechanisms of the innate and adaptive immunity. Although, the complement system provides the first line of defence against the invading pathogens, its aberrant uncontrolled activation causes extensive self tissue injury. A large number of humoral and cell surface complement regulatory protein keep the system well-regulated in healthy individuals. Complement profiling had brought important information on the pathophysiology of several infectious and chronic inflammatory disorders. In view of the diversity of the clinical disorders involving abnormal complement activity or regulation, which include both acute and chronic diseases that affect a wide range of organs, diverse yet specifically tailored therapeutic approaches may be needed to shift complement back into balance. This brief review discusses on the complement system, its functions and its importance as biomarkers and therapeutic targets for autoimmune diseases with focus on SLE and RA. PMID:26669012

  13. Candidate Pathway-Based GWAS Identifies Novel Associations of Genomic Variants in the Complement System Associated with Coronary Artery Disease

    PubMed Central

    Xu, Chengqi; Yang, Qin; Xiong, Hongbo; Wang, Longfei; Cai, Jianping; Wang, Fan; Li, Sisi; Chen, Jing; Wang, Chuchu; Wang, Dan; Xiong, Xin; Wang, Pengyun; Zhao, Yuanyuan; Wang, Xiaojing; Huang, Yufeng; Chen, Shanshan; Yin, Dan; Li, Xiuchun; Liu, Ying; Liu, Jinqiu; Wang, Jingjing; Li, Hui; Ke, Tie; Ren, Xiang; Wu, Yanxia; Wu, Gang; Wan, Jing; Zhang, Rongfeng; Wu, Tangchun; Wang, Junhan; Xia, Yunlong; Yang, Yanzong; Cheng, Xiang; Liao, Yuhua; Chen, Qiuyun; Zhou, Yanhong; He, Qing; Tu, Xin; Wang, Qing K.

    2014-01-01

    Background Genomic variants identified by genome-wide association studies (GWAS) explain <20% of heritability of coronary artery disease (CAD), thus many risk variants remain missing for CAD. Identification of new variants may unravel new biological pathways and genetic mechanisms for CAD. To identify new variants associated with CAD, we developed a candidate pathway-based GWAS by integrating expression quantitative loci (eQTL) analysis and mining of GWAS data with variants in a candidate pathway. Methods and Results Mining of GWAS data was performed to analyze variants in 32 complement system genes for positive association with CAD. Functional variants in genes showing positive association were then identified by searching existing expression quantitative loci databases and validated by RT-PCR. A follow-up case control design was then used to determine whether the functional variants are associated with CAD in two independent GeneID Chinese populations. Candidate pathway-based GWAS identified positive association between variants in C3AR1 and C6 and CAD. Two functional variants, rs7842 in C3AR1 and rs4400166 in C6, were found to be associated with expression levels of C3AR1 and C6, respectively. Significant association was identified between rs7842 and CAD (P=3.99×10−6, OR=1.47) and between rs4400166 and CAD (P=9.30×10−3, OR=1.24) in the validation cohort. The significant findings were confirmed in the replication cohort (P=1.53×10−5, OR=1.37 for rs7842; P=8.41×10−3, OR=1.21 for rs4400166. Conclusions Integration of GWAS with biological pathways and eQTL is effective in identifying new risk variants for CAD. Functional variants increasing C3AR1 and C6 expression were shown to confer significant risk of CAD for the first time. PMID:25249547

  14. Classical complement activation and acquired immune response pathways are not essential for retinal degeneration in the rd1 mouse

    PubMed Central

    Rohrer, Brbel; Demos, Christina; Frigg, Rico; Grimm, Christian

    2007-01-01

    Misregulation of the innate immune response and other immune-related processes have been suggested to play a critical role in the pathogenesis of a number of different neurodegenerative diseases, including age related macular degeneration. In an animal model for photoreceptor degeneration, several genes of the innate and acquired immune system were found to be differentially regulated in the retina during the degenerative process. In addition to this differential regulation of individual genes, we found that in the rd1 retina a significantly higher number of genes involved in immune-related responses were expressed at any given time during the degenerative period. The peak of immune-related gene expression was at postnatal day 14, coinciding with the peak of photoreceptor apoptosis in the rd1 mouse. We directly tested the potential involvement of acquired and innate immune responses in initiation and progression of photoreceptor degeneration by analyzing double mutant animals. Retinal morphology and photoreceptor apoptosis of rd1 mice on a SCID genetic background (no mature T- and B-cells) or in combination with a RAG-1 (no functional B- and T-cells) or a C1q? (no functional classical complement activation pathway) knockout was followed during the degenerative process using light microscopy or TUNEL staining, respectively. Although complement factor C1q? was highly up-regulated in the rd1 retina concomitantly with the degenerative process, lack of this protein did not protect the rd1 retina. Similarly, retinal degeneration and photoreceptor apoptosis appeared to proceed normally in the rd1 mouse lacking functional B- and T-cells. Our results suggest that both, the classical complement system of innate immunity and a functional acquired immune response are not essential for the degenerative process in the rd1 mouse retina. PMID:17069800

  15. Classical complement activation and acquired immune response pathways are not essential for retinal degeneration in the rd1 mouse.

    PubMed

    Rohrer, Brbel; Demos, Christina; Frigg, Rico; Grimm, Christian

    2007-01-01

    Misregulation of the innate immune response and other immune-related processes have been suggested to play a critical role in the pathogenesis of a number of different neurodegenerative diseases, including age related macular degeneration. In an animal model for photoreceptor degeneration, several genes of the innate and acquired immune system were found to be differentially regulated in the retina during the degenerative process. In addition to this differential regulation of individual genes, we found that in the rd1 retina a significantly higher number of genes involved in immune-related responses were expressed at any given time during the degenerative period. The peak of immune-related gene expression was at postnatal day 14, coinciding with the peak of photoreceptor apoptosis in the rd1 mouse. We directly tested the potential involvement of acquired and innate immune responses in initiation and progression of photoreceptor degeneration by analyzing double mutant animals. Retinal morphology and photoreceptor apoptosis of rd1 mice on a SCID genetic background (no mature T- and B-cells) or in combination with a RAG1 (no functional B- and T-cells) or a C1qalpha (no functional classical complement activation pathway) knockout was followed during the degenerative process using light microscopy or TUNEL staining, respectively. Although complement factor C1qalpha was highly up-regulated in the rd1 retina concomitantly with the degenerative process, lack of this protein did not protect the rd1 retina. Similarly, retinal degeneration and photoreceptor apoptosis appeared to proceed normally in the rd1 mouse lacking functional B- and T-cells. Our results suggest that both, the classical complement system of innate immunity and a functional acquired immune response are not essential for the degenerative process in the rd1 mouse retina. PMID:17069800

  16. Visual Pathway Study Using in vivo DTI Tractography to Complement Classical Anatomy

    PubMed Central

    Wu, Wentao; Rigolo, Laura; O’Donnell, Lauren J.; Norton, Isaiah; Shriver, Sargent; Golby, Alexandra J.

    2011-01-01

    Background Knowledge of the individual course of the optic radiations (OR) is important to avoid post-operative visual deficits. Cadaveric studies of the visual pathways are limited because it has not been possible to accurately separate the OR from neighboring tracts and results may not apply to individual patients. Diffusion tensor imaging (DTI) studies may be able to demonstrate the relationships between the OR and neighboring fibers in vivo in individual subjects. Objective To use DTI tractography to study the OR and Meyer’s loop (ML) anatomy in vivo. Methods Ten healthy subjects underwent magnetic resonance imaging with diffusion imaging at 3T. Using a fiducial-based DTI tractography tool (Slicer 3.3), seeds were placed near the lateral geniculate nucleus (LGN) to reconstruct individual visual pathways and neighboring tracts. Projections of the optic radiations onto 3D brain models were shown individually in order to quantify relationships to key landmarks. Results Two patterns of visual pathways were found. The OR ran more commonly deep in the whole superior and middle temporal gyri and superior temporal sulcus. The OR was closely surrounded in all cases by an inferior longitudinal fascicle and a parieto/occipito/temporo-pontine fascicle. The mean left and right distances between the tip of the OR and temporal pole were 39.8± 3.8mm and 40.6±5.7 mm, respectively. Conclusion DTI tractography provides a practical complementary method to study the OR and ML anatomy in vivo, and with reference to individual 3D brain anatomy. PMID:21808220

  17. Viral bimolecular fluorescence complementation: a novel tool to study intracellular vesicular trafficking pathways.

    PubMed

    Dirk, Brennan S; Jacob, Rajesh Abraham; Johnson, Aaron L; Pawlak, Emily N; Cavanagh, P Craig; Van Nynatten, Logan; Haeryfar, S M Mansour; Dikeakos, Jimmy D

    2015-01-01

    The Human Immunodeficiency Virus type 1 (HIV-1) accessory protein Nef interacts with a multitude of cellular proteins, manipulating the host membrane trafficking machinery to evade immune surveillance. Nef interactions have been analyzed using various in vitro assays, co-immunoprecipitation studies, and more recently mass spectrometry. However, these methods do not evaluate Nef interactions in the context of viral infection nor do they define the sub-cellular location of these interactions. In this report, we describe a novel bimolecular fluorescence complementation (BiFC) lentiviral expression tool, termed viral BiFC, to study Nef interactions with host cellular proteins in the context of viral infection. Using the F2A cleavage site from the foot and mouth disease virus we generated a viral BiFC expression vector capable of concurrent expression of Nef and host cellular proteins; PACS-1, MHC-I and SNX18. Our studies confirmed the interaction between Nef and PACS-1, a host membrane trafficking protein involved in Nef-mediated immune evasion, and demonstrated co-localization of this complex with LAMP-1 positive endolysosomal vesicles. Furthermore, we utilized viral BiFC to localize the Nef/MHC-I interaction to an AP-1 positive endosomal compartment. Finally, viral BiFC was observed between Nef and the membrane trafficking regulator SNX18. This novel demonstration of an association between Nef and SNX18 was localized to AP-1 positive vesicles. In summary, viral BiFC is a unique tool designed to analyze the interaction between Nef and host cellular proteins by mapping the sub-cellular locations of their interactions during viral infection. PMID:25915798

  18. Shiga Toxin Promotes Podocyte Injury in Experimental Hemolytic Uremic Syndrome via Activation of the Alternative Pathway of Complement

    PubMed Central

    Locatelli, Monica; Buelli, Simona; Pezzotta, Anna; Corna, Daniela; Perico, Luca; Tomasoni, Susanna; Rottoli, Daniela; Rizzo, Paola; Conti, Debora; Thurman, Joshua M.; Remuzzi, Giuseppe; Zoja, Carlamaria

    2014-01-01

    Shiga toxin (Stx)producing Escherichia coli is the offending agent of postdiarrhea-associated hemolytic uremic syndrome (HUS), a disorder of glomerular ischemic damage and widespread microvascular thrombosis. We previously documented that Stx induces glomerular complement activation, generating C3a responsible for microvascular thrombosis in experimental HUS. Here, we show that the presence of C3 deposits on podocytes is associated with podocyte damage and loss in HUS mice generated by the coinjection of Stx2 and LPS. Because podocyte adhesion to the glomerular basement membrane is mediated by integrins, the relevance of integrin-linked kinase (ILK) signals in podocyte dysfunction was evaluated. Podocyte expression of ILK increased after the injection of Stx2/LPS and preceded the upregulation of Snail and downregulation of nephrin and ?-actinin-4. Factor B deficiency or pretreatment with an inhibitory antibody to factor B protected mice against Stx2/LPS-induced podocyte dysregulation. Similarly, pretreatment with a C3a receptor antagonist limited podocyte loss and changes in ILK, Snail, and ?-actinin-4 expression. In cultured podocytes, treatment with C3a reduced ?-actinin-4 expression and promoted ILK-dependent nuclear expression of Snail and cell motility. These results suggest that Stx-induced activation of the alternative pathway of complement and generation of C3a promotes ILK signaling, leading to podocyte dysfunction and loss in Stx-HUS. PMID:24578132

  19. Shiga toxin promotes podocyte injury in experimental hemolytic uremic syndrome via activation of the alternative pathway of complement.

    PubMed

    Locatelli, Monica; Buelli, Simona; Pezzotta, Anna; Corna, Daniela; Perico, Luca; Tomasoni, Susanna; Rottoli, Daniela; Rizzo, Paola; Conti, Debora; Thurman, Joshua M; Remuzzi, Giuseppe; Zoja, Carlamaria; Morigi, Marina

    2014-08-01

    Shiga toxin (Stx)-producing Escherichia coli is the offending agent of postdiarrhea-associated hemolytic uremic syndrome (HUS), a disorder of glomerular ischemic damage and widespread microvascular thrombosis. We previously documented that Stx induces glomerular complement activation, generating C3a responsible for microvascular thrombosis in experimental HUS. Here, we show that the presence of C3 deposits on podocytes is associated with podocyte damage and loss in HUS mice generated by the coinjection of Stx2 and LPS. Because podocyte adhesion to the glomerular basement membrane is mediated by integrins, the relevance of integrin-linked kinase (ILK) signals in podocyte dysfunction was evaluated. Podocyte expression of ILK increased after the injection of Stx2/LPS and preceded the upregulation of Snail and downregulation of nephrin and ?-actinin-4. Factor B deficiency or pretreatment with an inhibitory antibody to factor B protected mice against Stx2/LPS-induced podocyte dysregulation. Similarly, pretreatment with a C3a receptor antagonist limited podocyte loss and changes in ILK, Snail, and ?-actinin-4 expression. In cultured podocytes, treatment with C3a reduced ?-actinin-4 expression and promoted ILK-dependent nuclear expression of Snail and cell motility. These results suggest that Stx-induced activation of the alternative pathway of complement and generation of C3a promotes ILK signaling, leading to podocyte dysfunction and loss in Stx-HUS. PMID:24578132

  20. Anti-Mouse Properdin TSR 5/6 Monoclonal Antibodies Block Complement Alternative Pathway-dependent Pathogenesis

    PubMed Central

    Bertram, Paula; Akk, Antonina M.; Zhou, Hui-fang; Mitchell, Lynne M.; Pham, Christine T.N.

    2015-01-01

    The complement alternative pathway (AP) is a major contributor to a broad and growing spectrum of diseases that includes age-related macular degeneration, atypical hemolytic uremic syndrome, and preeclampsia. As a result, there is much interest in the therapeutic disruption of AP activity. Properdin, the only positive regulator of the AP, is a particularly promising AP target. Several issues need to be clarified before the potential for properdin-directed therapy can be realized. In this report we use a portion of the mouse properdin protein, expressed in a bacterial system, to raise rabbit polyclonal and hamster monoclonal antibodies that block properdin-dependent pathogenesis. These antibodies, when employed with AP-dependent mouse disease models, can help evaluate the feasibility of properdin-directed therapy. PMID:25723276

  1. Role of the complement anaphylatoxin C5a-receptor pathway in atopic dermatitis in mice

    PubMed Central

    DANG, LIN; HE, LEI; WANG, YAN; XIONG, JIKUI; BAI, BINGXUE; LI, YUZHEN

    2015-01-01

    Atopic dermatitis (AD) is a chronic inflammatory skin disease with a genetic background. The C5a-receptor (C5aR) pathway has been reported to be involved in AD; however, the precise pathogenesis remains to be elucidated. In the present study, the contribution of the C5aR pathway to AD in mice was investigated. A BALB/c mouse model of AD was induced by application of 2,4-dinitrochlorobenzene (DNCB) onto hairless dorsal skin. Following DNCB application for 2 weeks, C5aR expression in skin tissue was assessed by reverse transcription quantitative polymerase chain reaction. C5aR expression in skin tissue was significantly increased in mice with AD. In an additional experiment, C5aR antagonist (C5aRA) intracutaneously injected in combination with DNCB treatment. The skin-fold thickness, number of total infiltrating leukocytes and mast cells infiltrating in skin tissue were measured. Interleukin-4 (IL-4) and interferon-γ (IFN-γ) levels in skin tissue and IL-4, IFN-γ, histamine and immunoglobulin E (IgE) levels in serum were measured using ELISA. The skin-fold thickness, numbers of total infiltrating leukocytes and mast cells in skin tissue, as well as levels of IL-4, IFN-γ, histamine and IgE were significantly increased in mice with AD. However, simultaneous treatment with C5aRA significantly attenuated increases in skin fold thickness and the numbers of total infiltrating leukocytes and mast cells in skin tissue. Treatment with C5aRA also decreased IL-4 and IFN-γ levels in skin tissue, as well as the levels of IL-4, IFN-γ, histamine and IgE in the serum. In conclusion, C5aRA inhibited AD in mice, possibly through suppression of the C5aR-mediated cascade action of mast cells. PMID:25650554

  2. ConBr, a lectin from Canavalia brasiliensis seeds, modulates signaling pathways and increases BDNF expression probably via a glycosylated target.

    PubMed

    Rieger, Dbora K; Cunha, Rodrigo M S; Lopes, Mark William; Costa, Ana Paula; Budni, Josiani; Rodrigues, Ana Lcia S; Walz, Roger; Teixeira, Edson H; Nascimento, Kyria S; Cavada, Benildo S; Leal, Rodrigo B

    2014-12-01

    In the central nervous system, many receptors, ion channels and neurotransmitter transporters are glycoproteins, where the glycan chains are modulator elements. Lectins are proteins, which recognize and bind carbohydrate complexes. We have previously shown that ConBr, a lectin purified from Canavalia brasiliensis seeds, produced antidepressant-like effect and blocked hippocampal neurotoxicity induced by quinolinic acid and glutamate. Noteworthy, all these effects occurred in a dependence of its carbohydrate recognition domain. Therefore, the present study was undertaken in order to elucidate intracellular signaling pathways regulated by ConBr that may be potentially associated with the antidepressant and neuroprotective effects previously reported to be dependent on carbohydrate interaction. ConBr (10?g/site) was injected into the ventricle (i.c.v.) of mice, and the hippocampi were removed 0.5, 1, 3, 6, 8, 12, 18, and 24?h after treatment. Our results showed that in the period of 0.5-3?h, ConBr induced activation of the protein kinases Akt, ERK1, and PKA. Furthermore, the phosphorylation of CREB-Ser133 was stimulated by ConBr (1-6?h), while brain-derived neurotrophic factor (BDNF) mRNA was increased at 12?h and BDNF protein at 18-24?h. Our data suggest that an early activation of protein kinases may trigger CREB-dependent BDNF transcription, resulting in a subsequent increase of BDNF protein in response to ConBr. Later, increment of Akt phosphorylation was observed 24?h after ConBr administration, possibly due to BDNF/TrkB-dependent activation of Akt. Our findings indicate that ConBr is a multifunctional molecule capable to activate signaling pathways involved in neuroplasticity and neuroprotection. PMID:25319623

  3. Mannose-Binding Lectin Inhibits Monocyte Proliferation through Transforming Growth Factor-β1 and p38 Signaling Pathways

    PubMed Central

    Wang, Yan; Chen, A-De; Lei, Yan-Mei; Shan, Gui-Qiu; Zhang, Li-Yun; Lu, Xiao; Chen, Zheng-Liang

    2013-01-01

    Mannose-binding lectin (MBL), a plasma C-type lectin, plays an important role in innate immunity. However, the interaction, and the consequences of it, between MBL and the immune system remain ill defined. We have investigated the contributing mechanisms and effects of MBL on the proliferation of human monocytes. At lower concentrations (≤4 μg/ml) MBL was shown to partially enhance monocyte proliferation. By contrast, at higher concentrations (8–20 μg/ml) of MBL, cell proliferation was markedly attenuated. MBL-induced growth inhibition was associated with G0/G1 arrest, down-regulation of cyclin D1/D3, cyclin-dependent kinase (Cdk) 2/Cdk4 and up-regulation of the Cdk inhibitory protein Cip1/p21. Additionally, MBL induced apoptosis, and did so through caspase-3 activation and poly ADP-ribose polymerase (PARP) cleavage. Moreover, transforming growth factor (TGF)-β1 levels increased in the supernatants of MBL-stimulated monocyte cultures. We also found that MBL-dependent inhibition of monocyte proliferation could be reversed by the TGF-β receptor antagonist SB-431542, or by anti-TGF-β1 antibody, or by the mitogen-activated protein kinase (MAPK) inhibitors specific for p38 (SB203580), but not ERK (U0126) or JNK (SP600125). Thus, at high concentrations, MBL can affect the immune system by inhibiting monocyte proliferation, which suggests that MBL may exhibit anti-inflammatory effects. PMID:24039775

  4. Rapid Assays for Lectin Toxicity and Binding Changes that Reflect Altered Glycosylation in Mammalian Cells

    PubMed Central

    Stanley, Pamela; Sundaram, Subha

    2014-01-01

    Glycosylation engineering is used to generate glycoproteins, glycolipids or proteoglycans with a more defined complement of glycans on their glycoconjugates. For example, a mammalian cell glycosylation mutant lacking a specific glycosyltransferase generates glycoproteins, and/or glycolipids, and/or proteoglycans, with truncated glycans missing the sugar transferred by that glycosyltransferase, and also missing those sugars that would be added subsequently. In some cases, an alternative glycosyltransferase may then use the truncated glycans as acceptors, thereby generating a new or different glycan subset in the mutant cell. Another type of glycosylation mutant arises from gain-of-function mutations that, for example, activate a silent glycosyltransferase gene. In this case, glycoconjugates will have glycans with additional sugar(s) that are more elaborate than the glycans of wild type cells. Mutations in other genes that affect glycosylation, such as nucleotide sugar synthases or transporters, will alter the glycan complement in more general ways that usually affect several types of glycoconjugates. There are now many strategies for generating a precise mutation in a glycosylation gene in a mammalian cell. Large-volume cultures of mammalian cells may also give rise to spontaneous mutants in glycosylation pathways. This article will focus on how to rapidly characterize mammalian cells with an altered glycosylation activity. The key reagents for the protocols described are plant lectins that bind mammalian glycans with varying avidities, depending on the specific structure of those glycans. Cells with altered glycosylation generally become resistant or hypersensitive to lectin toxicity, and have reduced or increased lectin or antibody binding. Here we describe rapid assays to compare the cytotoxicity of lectins in a lectin resistance test, and the binding of lectins or antibodies by flow cytometry in a glycan-binding assay. Based on these tests, glycosylation changes expressed by a cell can be revealed, and glycosylation mutants classified into phenotypic groups that may reflect a loss-of-function or gain-of-function mutation in a specific gene involved in glycan synthesis. PMID:24903886

  5. Activation of Ras and mitogen-activated protein kinase pathway by terminal complement complexes is G protein dependent.

    PubMed

    Niculescu, F; Rus, H; van Biesen, T; Shin, M L

    1997-05-01

    Assembly of terminal complement complexes (TCC) C5b-7, C5b-8, and C5b-9 on target cells during acute and chronic inflammation induces hydrolysis of plasma membrane phospholipids and heterotrimeric G protein activation. TCC also stimulate a variety of cellular activities, which include cytokine synthesis, proto-oncogene activation, and mitotic signaling. Now we report that sublytic TCC induced Ras, Raf-1, and extracellular signal-regulated kinase (ERK) 1 activation in JY25 B cell line. When cells were exposed to C5b-9, GTP-bound Ras in anti-C5b-9 immunoprecipitates was increased 3.2-fold at 2 min, while GTP-bound Ras in anti-Ras immunoprecipitates was increased 2-fold at 10 min. Both C5b-9 and C5b-7, but not C5b6, increased Raf-1 kinase activity maximum 3.3-fold at 2 min and 2.8-fold at 5 min, respectively. ERK1 activity was 2-fold increased by C5b-9 at 2 min and by C5b-7 at 10 min, over the C5b6 level. The role of mitogen-activated protein kinase (MAPK) pathway on TCC-inducible mitotic signaling was evaluated by assessing DNA synthesis and activator protein 1 (AP-1) DNA-binding activity. The MAPK/ERK-specific inhibitor PD 098,059 abolished the C5b-9-induced DNA synthesis. Involvement of G protein in the activation of MAPK pathway by TCC was indicated by inhibition of Raf-1 and ERK1 kinase activity, as well as the DNA synthesis by pretreatment of cells with pertussis toxin. Overexpression of beta-adrenergic receptor kinase 1 carboxyl-terminal peptide in JY25 cells also inhibited Raf-1 and ERK1 activity, indicating a direct involvement of G betagamma subunits in the signal transduction generated through activation of MAPK pathway by TCC assembly in the plasma membrane. PMID:9127005

  6. Outer Membrane Protein P5 Is Required for Resistance of Nontypeable Haemophilus influenzae to Both the Classical and Alternative Complement Pathways

    PubMed Central

    Rosadini, Charles V.; Ram, Sanjay

    2014-01-01

    The complement system is an important first line of defense against the human pathogen Haemophilus influenzae. To survive and propagate in vivo, H. influenzae has evolved mechanisms for subverting this host defense, most of which have been shown to involve outer surface structures, including lipooligosaccharide glycans and outer surface proteins. Bacterial defense against complement acts at multiple steps in the pathway by mechanisms that are not fully understood. Here we identify outer membrane protein P5 as an essential factor in serum resistance of both H. influenzae strain Rd and nontypeable H. influenzae (NTHi) clinical isolate NT127. P5 was essential for resistance of Rd and NT127 to complement in pooled human serum. Further investigation determined that P5 expression decreased cell surface binding of IgM, a potent activator of the classical pathway of complement, to both Rd and NT127. Additionally, P5 expression was required for NT127 to bind factor H (fH), an important inhibitor of alternative pathway (AP) activation. Collectively, the results obtained in this work highlight the ability of H. influenzae to utilize a single protein to perform multiple protective functions for evading host immunity. PMID:24478079

  7. SALSA: A Regulator of the Early Steps of Complement Activation on Mucosal Surfaces

    PubMed Central

    Reichhardt, Martin Parnov; Meri, Seppo

    2016-01-01

    Complement is present mainly in blood. However, following mechanical damage or inflammation, serous exudates enter the mucosal surfaces. Here, the complement proteins interact with other endogenous molecules to keep microbes from entering the parenteral tissues. One of the mucosal proteins known to interact with the early complement components of both the classical and the lectin pathway is the salivary scavenger and agglutinin (SALSA). SALSA is also known as deleted in malignant brain tumors 1 and gp340. It is found both attached to the epithelium and secreted into the surrounding fluids of most mucosal surfaces. SALSA has been shown to bind directly to C1q, mannose-binding lectin, and the ficolins. Through these interactions SALSA regulates activation of the complement system. In addition, SALSA interacts with surfactant proteins A and D, secretory IgA, and lactoferrin. Ulcerative colitis and Crohn’s disease are examples of diseases, where complement activation in mucosal tissues may occur. This review describes the latest advances in our understanding of how the early complement components interact with the SALSA molecule. Furthermore, we discuss how these interactions may affect disease propagation on mucosal surfaces in immunological and inflammatory diseases. PMID:27014265

  8. Complement system in zebrafish.

    PubMed

    Zhang, Shicui; Cui, Pengfei

    2014-09-01

    Zebrafish is recently emerging as a model species for the study of immunology and human diseases. Complement system is the humoral backbone of the innate immune defense, and our knowledge as such in zebrafish has dramatically increased in the recent years. This review summarizes the current research progress of zebrafish complement system. The global searching for complement components in genome database, together with published data, has unveiled the existence of all the orthologues of mammalian complement components identified thus far, including the complement regulatory proteins and complement receptors, in zebrafish. Interestingly, zebrafish complement components also display some distinctive features, such as prominent levels of extrahepatic expression and isotypic diversity of the complement components. Future studies should focus on the following issues that would be of special importance for understanding the physiological role of complement components in zebrafish: conclusive identification of complement genes, especially those with isotypic diversity; analysis and elucidation of function and mechanism of complement components; modulation of innate and adaptive immune response by complement system; and unconventional roles of complement-triggered pathways. PMID:24462834

  9. The Contribution of Mannose Binding Lectin to Reperfusion Injury after Ischemic Stroke

    PubMed Central

    Morrison, Helena; Frye, Jennifer; Davis-Gorman, Grace; Funk, Janet; McDonagh, Paul; Stahl, Gregory; Ritter, Leslie

    2012-01-01

    After complement system (CS) activation, the sequential production of complement products increases cell injury and death through opsonophagocytosis, cytolysis, adaptive, and inflammatory cell responses. These responses potentiate cerebral ischemia-reperfusion (IR) injury after ischemic stroke and reperfusion. Activation of the CS via mannose binding lectin (MBL)-initiated lectin pathway is known to increase tissue damage in response to IR in muscle, myocardium and intestine tissue. In contrast, the contribution of this pathway to cerebral IR injury, a neutrophil-mediated event, is less clear. Therefore, we investigated the potential protective role of MBL deficiency in neutrophil-mediated cerebral injury after IR. Using an intraluminal filament method, neutrophil activation and cerebral injury were compared between MBL-deficient and wild type C57Bl/6 mice subjected to 60 minutes of MCA ischemia and reperfusion. Systemic neutrophil activation was not decreased in MBL-deficient animals after IR. In MBL-deficient animals, cerebral injury was significantly decreased only in the striatum (p < 0.05). Despite MBL deficiency, C3 depositions were evident in the injured hemisphere during reperfusion. These results indicate that while MBL deficiency results in a modest protection of a sub-cortical brain region during IR, redundant complement pathway activation may overwhelm further beneficial effects of MBL deficiency during reperfusion. PMID:21208161

  10. Mannose-binding lectin polymorphisms and rheumatoid arthritis: A short review and meta-analysis.

    PubMed

    Epp Boschmann, Stefanie; Goeldner, Isabela; Tuon, Felipe Francisco; Schiel, Wagner; Aoyama, Fernanda; de Messias-Reason, Iara J

    2016-01-01

    Mannose-binding lectin (MBL) is a pattern recognition receptor of the lectin pathway of complement system. MBL binds to carbohydrates on microorganism's surfaces leading to complement activation, opsonization and phagocytosis. Polymorphisms in the MBL gene (MBL2) are associated with variations on MBL serum levels and with the susceptibility to various infectious and autoimmune diseases. The involvement of the lectin pathway in rheumatoid arthritis (RA) has been demonstrated by several studies and although MBL has been considered to have a dual role in the pathogenesis of the disease, the association between MBL and RA remains inconclusive. In an attempt to clarify this relationship, we developed this short review summarizing accumulated evidences in regard to MBL and RA and a meta-analysis to evaluate the influence of MBL2 polymorphisms on the susceptibility to RA. Among a total of 217 articles that were identified following a predefined search strategy on PubMed, Scopus, Scielo, EMBASE and Cochrane databases, only 13 met all inclusion criteria and were included in the meta-analysis. Data assessment was conducted by three independent investigators and presented in odds ratio (OR) and 95% confidence intervals (CIs) using forest plot charts. Both heterogeneity and publication bias were analyzed. The results of the meta-analysis evidenced that MBL2 low producing OO and XX genotypes do not confer higher risk to RA, even when data were analyzed according to cohort's ethnicity. Further studies are needed in order to clarify the importance of other genes of the lectin pathway in the pathogenesis of RA. PMID:26608926

  11. Functional assessment of mouse complement pathway activities and quantification of C3b/C3c/iC3b in an experimental model of mouse renal ischaemia/reperfusion injury.

    PubMed

    Kotimaa, Juha P; van Werkhoven, Maaike B; O'Flynn, Joseph; Klar-Mohamad, Ngaisah; van Groningen, Jan; Schilders, Geurt; Rutjes, Helma; Daha, Mohamed R; Seelen, Marc A; van Kooten, Cees

    2015-04-01

    The complement system is an essential component of our innate immunity, both for the protection against infections and for proper handling of dying cells. However, the complement system can also contribute to tissue injury and inflammatory responses. In view of novel therapeutic possibilities, there is an increasing interest in measurement of the complement system activation in the systemic compartment, both in the clinical setting as well as in experimental models. Here we describe in parallel a sensitive and specific sandwich ELISA detecting mouse C3 activation fragments C3b/C3c/iC3b, as well as functional complement ELISAs detecting specific activities of the three complement pathways at the level of C3 and at the level of C9 activation. In a murine model of renal ischaemia/reperfusion injury (IRI) we found transient complement activation as shown by generation of C3b/C3c/iC3b fragments at 24 h following reperfusion, which returned to base-line at 3 and 7 days post reperfusion. When the pathway specific complement activities were measured at the level of C3 activation, we found no significant reduction in any of the pathways. However, the functional complement activity of all three pathways was significantly reduced when measured at the level of C9, with the strongest reduction being observed in the alternative pathway. For all three pathways there was a strong correlation between the amount of C3 fragments and the reduction in functional complement activity. Moreover, at 24 h both C3 fragments and the functional complement activities showed a correlation with the rise in serum creatinine. Together our results show that determination of the systemic pathway specific complement activity is feasible in experimental mouse models and that they are useful in understanding complement activation and inhibition in vivo. PMID:25733354

  12. Sundanese Complementation

    ERIC Educational Resources Information Center

    Kurniawan, Eri

    2013-01-01

    The focus of this thesis is the description and analysis of clausal complementation in Sundanese, an Austronesian language spoken in Indonesia. The thesis examined a range of clausal complement types in Sundanese, which consists of (i) "yen/(wi)rehna" "that" complements, (ii) "pikeun" "for" complements,

  13. Sundanese Complementation

    ERIC Educational Resources Information Center

    Kurniawan, Eri

    2013-01-01

    The focus of this thesis is the description and analysis of clausal complementation in Sundanese, an Austronesian language spoken in Indonesia. The thesis examined a range of clausal complement types in Sundanese, which consists of (i) "yen/(wi)rehna" "that" complements, (ii) "pikeun" "for" complements,…

  14. Splenic RNA and MicroRNA Mimics Promote Complement Factor B Production and Alternative Pathway Activation via Innate Immune Signaling.

    PubMed

    Zou, Lin; Feng, Yan; Xu, Ganqiong; Jian, Wenling; Chao, Wei

    2016-03-15

    Complement factor B (cfB) is an essential component of the alternative pathway (AP) and plays an important role in the pathogenesis of polymicrobial sepsis. However, the mechanism leading to cfB production and AP activation during sepsis remains poorly understood. In this study, we found that plasma cell-free RNA was significantly increased following cecal ligation and puncture (CLP), an animal model of polymicrobial sepsis, and was closely associated with sepsis severity. Quantitative RT-PCR and microRNA (miRNA) array analysis revealed an increase in bacterial RNA and multiple host miRNAs (miR-145, miR-146a, miR-122, miR-210) in the blood following CLP. Treatment with tissue RNA or synthetic miRNA mimics (miR-145, miR-146a, miR-122, miR-34a) induced a marked increase in cfB production in cardiomyocytes or macrophages. The newly synthesized cfB released into medium was biologically active because it participated in AP activation initiated by cobra venom factor. Genetic deletion of TLR7 or MyD88, but not TLR3, and inhibition of the MAPKs (JNK and p38) or NF-κB abolished miR-146a-induced cfB production. In vivo, CLP led to a significant increase in splenic cfB expression that correlated with the plasma RNA or miRNA levels. Peritoneal injection of RNA or miR-146a led to an increase in cfB expression in the peritoneal space that was attenuated in MyD88-knockout or TLR7-knockout mice, respectively. These findings demonstrate that host cellular RNA and specific miRNAs are released into the circulation during polymicrobial sepsis and may function as extracellular mediators capable of promoting cfB production and AP activation through specific TLR7 and MyD88 signaling. PMID:26889043

  15. Human Mannose-Binding Lectin Inhibitor Prevents Myocardial Injury and Arterial Thrombogenesis in a Novel Animal Model

    PubMed Central

    Pavlov, Vasile I.; Tan, Ying S.; McClure, Erin E.; La Bonte, Laura R.; Zou, Chenhui; Gorsuch, William B.; Stahl, Gregory L.

    2016-01-01

    Myocardial infarction and coagulation disorders are leading causes of disability and death in the world. An important role of the lectin complement pathway in myocardial infarction and coagulation has been demonstrated in mice genetically deficient in lectin complement pathway proteins. However, these studies are limited to comparisons between wild-type and deficient mice and lack the ability to examine reversal/inhibition of injury after disease establishment. We developed a novel mouse that expresses functional human mannose-binding lectin (MBL) 2 under the control of Mbl1 promoter. Serum MBL2 concentrations averaged approximately 3 ?g/mL in MBL2+/+Mbl1?/?Mbl2?/? [MBL2 knock in (KI)] mice. Serum MBL2 level in MBL2 KI mice significantly increased after 7 (8 ?g/mL) or 14 (9 ?g/mL) days of hyperglycemia compared to normoglycemic mice (P<0.001). Monoclonal antibody 3F8 inhibited C3 deposition on mannan-coated plates in MBL2 KI, but not wild-type, mice. Myocardial ischemia/reperfusion in MBL2 KI mice revealed that 3F8 preserved cardiac function and decreased infarct size and fibrin deposition in a time-dependent manner. Furthermore, 3F8 prevented ferric chlorideinduced occlusive arterial thrombogenesis invivo. MBL2 KI mice represent a novel animal model that can be used to study the lectin complement pathway in acute and chronic models of human disease. Furthermore, these novel mice demonstrate the therapeutic window for MBL2 inhibition for effective treatment of disease and its complications. PMID:25482922

  16. Infections of People with Complement Deficiencies and Patients Who Have Undergone Splenectomy

    PubMed Central

    Ram, Sanjay; Lewis, Lisa A.; Rice, Peter A.

    2010-01-01

    Summary: The complement system comprises several fluid-phase and membrane-associated proteins. Under physiological conditions, activation of the fluid-phase components of complement is maintained under tight control and complement activation occurs primarily on surfaces recognized as “nonself” in an attempt to minimize damage to bystander host cells. Membrane complement components act to limit complement activation on host cells or to facilitate uptake of antigens or microbes “tagged” with complement fragments. While this review focuses on the role of complement in infectious diseases, work over the past couple of decades has defined several important functions of complement distinct from that of combating infections. Activation of complement in the fluid phase can occur through the classical, lectin, or alternative pathway. Deficiencies of components of the classical pathway lead to the development of autoimmune disorders and predispose individuals to recurrent respiratory infections and infections caused by encapsulated organisms, including Streptococcus pneumoniae, Neisseria meningitidis, and Haemophilus influenzae. While no individual with complete mannan-binding lectin (MBL) deficiency has been identified, low MBL levels have been linked to predisposition to, or severity of, several diseases. It appears that MBL may play an important role in children, who have a relatively immature adaptive immune response. C3 is the point at which all complement pathways converge, and complete deficiency of C3 invariably leads to severe infections, including those caused by meningococci and pneumococci. Deficiencies of the alternative and terminal complement pathways result in an almost exclusive predisposition to invasive meningococcal disease. The spleen plays an important role in antigen processing and the production of antibodies. Splenic macrophages are critical in clearing opsonized encapsulated bacteria (such as pneumococci, meningococci, and Escherichia coli) and intraerythrocytic parasites such as those causing malaria and babesiosis, which explains the fulminant nature of these infections in persons with anatomic or functional asplenia. Paramount to the management of patients with complement deficiencies and asplenia is educating patients about their predisposition to infection and the importance of preventive immunizations and seeking prompt medical attention. PMID:20930072

  17. Complement and Viral Pathogenesis

    PubMed Central

    Stoermer, Kristina A.; Morrison, Thomas E.

    2011-01-01

    The complement system functions as an immune surveillance system that rapidly responds to infection. Activation of the complement system by specific recognition pathways triggers a protease cascade, generating cleavage products that function to eliminate pathogens, regulate inflammatory responses, and shape adaptive immune responses. However, when dysregulated, these powerful functions can become destructive and the complement system has been implicated as a pathogenic effector in numerous diseases, including infectious diseases. This review highlights recent discoveries that have identified critical roles for the complement system in the pathogenesis of viral infection. PMID:21292294

  18. Mapping of Functional Domains in Herpesvirus Saimiri Complement Control Protein Homolog: Complement Control Protein Domain 2 Is the Smallest Structural Unit Displaying Cofactor and Decay-Accelerating Activities?

    PubMed Central

    Singh, Akhilesh K.; Yadav, Viveka Nand; Pyaram, Kalyani; Mullick, Jayati; Sahu, Arvind

    2009-01-01

    Herpesvirus saimiri encodes a functional homolog of human regulator-of-complement-activation proteins named CCPH that inactivates complement by accelerating the decay of C3 convertases and by serving as a cofactor in factor I-mediated inactivation of their subunits C3b and C4b. Here, we map the functional domains of CCPH. We demonstrate that short consensus repeat 2 (SCR2) is the minimum domain essential for classical/lectin pathway C3 convertase decay-accelerating activity as well as for factor I cofactor activity for C3b and C4b. Thus, CCPH is the first example wherein a single SCR domain has been shown to display complement regulatory functions. PMID:19640995

  19. Laboratory tests for disorders of complement and complement regulatory proteins.

    PubMed

    Shih, Angela R; Murali, Mandakolathur R

    2015-12-01

    The complement pathway is a cascade of proteases that is involved in immune surveillance and innate immunity, as well as adaptive immunity. Dysfunction of the complement cascade may be mediated by aberrations in the pathways of activation, complement regulatory proteins, or complement deficiencies, and has been linked to a number of hematologic disorders, including paroxysmal noctural hemoglobinuria (PNH), hereditary angioedema (HAE), and atypical hemolytic-uremic syndrome (aHUS). Here, current laboratory tests for disorders of the complement pathway are reviewed, and their utility and limitations in hematologic disorders and systemic diseases are discussed. Current therapeutic advances targeting the complement pathway in treatment of complement-mediated hematologic disorders are also reviewed. Am. J. Hematol. 90:1180-1186, 2015. 2015 Wiley Periodicals, Inc. PMID:26437749

  20. Complement mediators: key regulators of airway tissue remodeling in asthma.

    PubMed

    Khan, Mohammad Afzal; Assiri, Abdullah Mohammed; Broering, Dieter Clemens

    2015-01-01

    The complement mediators are the major effectors of the immune balance, which operates at the interface between the innate and adaptive immunity, and is vital for many immunoregulatory functions. Activation of the complement cascade through the classical, alternative or lectin pathways thus generating opsonins like C3b and C5b, anaphylatoxins C3a and C5a, chemotaxin, and inflammatory mediators, which leads to cellular death. Complement mediators that accelerate the airway remodeling are not well defined; however, an uncontrolled Th2-driven adaptive immune response has been linked to the major pathophysiologic features of asthma, including bronchoconstriction, airway hyperresponsiveness, and airway inflammation. The mechanisms leading to complement mediated airway tissue remodeling, and the effect of therapy on preventing and/or reversing it are not clearly understood. This review highlights complement-mediated inflammation, and the mechanism through it triggers the airway tissue injury and remodeling in the airway epithelium that could serve as potential targets for developing a new drug to rescue the asthma patients. PMID:26289385

  1. Use of peanut lectin and rat mammary stem cell lines to identify a cellular differentiation pathway for the alveolar cell in the rat mammary gland.

    PubMed

    Rudland, P S

    1992-10-01

    The presence of the carbohydrate receptor for PNL has been used to identify the previously described morphological types of epithelial cell produced as the stem cell line rat mammary 25 (Rama 25) differentiates to casein secretory alveolar-like cells in vitro. Thus when cultures of the epithelial stem cell line Rama 25 are treated with neuraminidase, fluorescently-conjugated PNL fails to stain cuboidal cells, stains weakly grey cells, and stains strongly the surface of dark cells. When superconfluent cultures of Rama 25 are treated with dimethyl sulfoxide or retinoic acid and prolactin, estradiol, hydrocortisone, and insulin to induce differentiation to alveolar cells, PNL stains strongly the untreated surfaces of droplet cells and casein-secreting vacuolated cells. PNL-staining of the derivative cell lines with truncated cellular pathways, and quantitative binding of [125I]-labeled PNL to the cultured cells are consistent with this cellular staining pattern. The presence of the carbohydrate receptor for peanut lectin (PNL) has also been used to identify specific epithelial cell types in different mammary structures of the developing rat mammary gland, as they differentiate to casein secretory alveolar cells in vivo. Thus when different structures of the developing rat mammary gland are treated with neuraminidase, peroxidase-conjugated PNL fails to stain histochemically the majority of epithelial cells in ducts, stains the cytoplasm of the majority of epithelial cells in terminal end-buds (TEBs), and stains strongly the luminal surfaces of the majority of epithelial cells in alveolar buds (ABs). PNL also stains the untreated luminal surfaces of alveolar cells, whether or not the cells can be stained with a monoclonal antibody to rat beta-casein. Stimulation of mammary differentiation by an analogue of ethyl retinoate or by perphenazine causes cells in end-buds to bind PNL without the necessity for their desialylation similar to that seen in casein secretory alveoli of lactating rats. In conclusion the different interconverting cell types of Rama 25 which form a pathway to casein-secretory cells in vitro are thus equated with recognisable epithelial cell types in vivo. These results suggest that casein-secretory cells in vivo are generated by similar successive interconversions between the major epithelial cell types present in the different mammary structures in the order: ducts, TEBs, ABs, alveoli, and secretory alveoli. PMID:1522129

  2. Commercially Available Complement Component-Depleted Sera Are Unexpectedly Codepleted of Ficolin-2

    PubMed Central

    Brady, Allison M.; Geno, K. Aaron; Dalecki, Alex G.; Cheng, Xiaogang

    2014-01-01

    The ficolins are a family of innate pattern recognition molecules that are known to bind acetylated compounds and activate complement through the association of mannose binding lectin (MBL)/ficolin-associated serine proteases (MASPs). Their importance has more recently become appreciated, as they have been shown to play a role in a variety of disease processes from infection to autoimmunity. While studying ficolin-2-mediated complement deposition on Streptococcus pneumoniae, we found that sera depleted of C1q or other complement components were also codepleted of ficolin-2 but not ficolin-1, ficolin-3, or MBL. MBL present in C1q-depleted sera was able to mediate complement deposition on Saccharomyces cerevisiae, suggesting the presence of MASPs. We found that complement was activated on pneumococci in C1q-depleted serum only after opsonization with exogenous recombinant ficolin-2 (rFicolin-2). Also, no complement deposition was observed in C1q-depleted serum when pneumococci were opsonized with rFicolin-2 mutated at its lysine-57 residue, where MASPs are known to associate. Thus, these depleted sera are a unique tool to study ficolin-2-mediated complement pathways; however, one should be aware that ficolin-2 is absent from complement component-depleted sera. PMID:25030054

  3. The role of complement in the pathogenesis of renal ischemia-reperfusion injury and fibrosis

    PubMed Central

    2014-01-01

    The complement system is a major component of innate immunity and has been commonly identified as a central element in host defense, clearance of immune complexes, and tissue homeostasis. After ischemia-reperfusion injury (IRI), the complement system is activated by endogenous ligands that trigger proteolytic cleavage of complement components via the classical, lectin and/or alternative pathway. The result is the formation of terminal complement components C3a, C5a, and the membrane attack complex (C5b-9 or MAC), all of which play pivotal roles in the amplification of the inflammatory response, chemotaxis, neutrophil/monocyte recruitment and activation, and direct tubular cell injury. However, recent evidence suggests that complement activity transcends innate host defense and there is increasing data suggesting complement as a regulator in processes such as allo-immunity, stem cell differentiation, tissue repair, and progression to fibrosis. In this review, we discuss recent advances addressing the role of complement as a regulator of IRI and renal fibrosis after organ donation for transplantation. We will also briefly discuss currently approved therapies that target complement activity in kidney ischemia-reperfusion and transplantation. PMID:25383094

  4. Regulatory Components of the Alternative Complement Pathway in Endothelial Cell Cytoplasm, Factor H and Factor I, Are Not Packaged in Weibel-Palade Bodies

    PubMed Central

    Turner, Nancy A.; Sartain, Sarah E.; Hui, Shiu-Ki; Moake, Joel L.

    2015-01-01

    It was recently reported that factor H, a regulatory component of the alternative complement pathway, is stored with von Willebrand factor (VWF) in the Weibel-Palade bodies of endothelial cells. If this were to be the case, it would have therapeutic importance for patients with the atypical hemolytic-uremic syndrome that can be caused either by a heterozygous defect in the factor H gene or by the presence of an autoantibody against factor H. The in vivo Weibel-Palade body secretagogue, des-amino-D-arginine vasopressin (DDAVP), would be expected to increase transiently the circulating factor H levels, in addition to increasing the circulating levels of VWF. We describe experiments demonstrating that factor H is released from endothelial cell cytoplasm without a secondary storage site. These experiments showed that factor H is not stored with VWF in endothelial cell Weibel-Palade bodies, and is not secreted in response in vitro in response to the Weibel-Palade body secretagogue, histamine. Furthermore, the in vivo Weibel-Palade body secretagogue, DDAVP does not increase the circulating factor H levels concomitantly with DDAVP-induced increased VWF. Factor I, a regulatory component of the alternative complement pathway that is functionally related to factor H, is also located in endothelial cell cytoplasm, and is also not present in endothelial cell Weibel-Palade bodies. Our data demonstrate that the factor H and factor I regulatory proteins of the alternative complement pathway are not stored in Weibel-Palade bodies. DDAVP induces the secretion into human plasma of VWF - but not factor H. PMID:25803806

  5. Complement in hemolytic anemia.

    PubMed

    Brodsky, Robert A

    2015-12-01

    Complement is increasingly being recognized as an important driver of human disease, including many hemolytic anemias. Paroxysmal nocturnal hemoglobinuria (PNH) cells are susceptible to hemolysis because of a loss of the complement regulatory proteins CD59 and CD55. Patients with atypical hemolytic uremic syndrome (aHUS) develop a thrombotic microangiopathy (TMA) that in most cases is attributable to mutations that lead to activation of the alternative pathway of complement. For optimal therapy, it is critical, but often difficult, to distinguish aHUS from other TMAs, such as thrombotic thrombocytopenic purpura; however, novel bioassays are being developed. In cold agglutinin disease (CAD), immunoglobulin M autoantibodies fix complement on the surface of red cells, resulting in extravascular hemolysis by the reticuloendothelial system. Drugs that inhibit complement activation are increasingly being used to treat these diseases. This article discusses the pathophysiology, diagnosis, and therapy for PNH, aHUS, and CAD. PMID:26637747

  6. Complement in hemolytic anemia.

    PubMed

    Brodsky, Robert A

    2015-11-26

    Complement is increasingly being recognized as an important driver of human disease, including many hemolytic anemias. Paroxysmal nocturnal hemoglobinuria (PNH) cells are susceptible to hemolysis because of a loss of the complement regulatory proteins CD59 and CD55. Patients with atypical hemolytic uremic syndrome (aHUS) develop a thrombotic microangiopathy (TMA) that in most cases is attributable to mutations that lead to activation of the alternative pathway of complement. For optimal therapy, it is critical, but often difficult, to distinguish aHUS from other TMAs, such as thrombotic thrombocytopenic purpura; however, novel bioassays are being developed. In cold agglutinin disease (CAD), immunoglobulin M autoantibodies fix complement on the surface of red cells, resulting in extravascular hemolysis by the reticuloendothelial system. Drugs that inhibit complement activation are increasingly being used to treat these diseases. This article discusses the pathophysiology, diagnosis, and therapy for PNH, aHUS, and CAD. PMID:26582375

  7. Early graft failure of GalTKO pig organs in baboons is reduced by expression of a human complement pathway-regulatory protein.

    PubMed

    Azimzadeh, Agnes M; Kelishadi, Sean S; Ezzelarab, Mohamed B; Singh, Avneesh K; Stoddard, Tiffany; Iwase, Hayato; Zhang, Tianshu; Burdorf, Lars; Sievert, Evelyn; Avon, Chris; Cheng, Xiangfei; Ayares, David; Horvath, Keith A; Corcoran, Philip C; Mohiuddin, Muhammad M; Barth, Rolf N; Cooper, David K C; Pierson, Richard N

    2015-01-01

    We describe the incidence of early graft failure (EGF, defined as loss of function from any cause within 3 days after transplant) in a large cohort of GalTKO pig organs transplanted into baboons in three centers, and the effect of additional expression of a human complement pathway-regulatory protein, CD46 or CD55 (GalTKO.hCPRP). Baboon recipients of life-supporting GalTKO kidney (n = 7) or heterotopic heart (n = 14) grafts received either no immunosuppression (n = 4), or one of several partial or full immunosuppressive regimens (n = 17). Fourteen additional baboons received a GalTKO.hCPRP kidney (n = 5) or heart (n = 9) and similar treatment regimens. Immunologic, pathologic, and coagulation parameters were measured at frequent intervals. EGF of GalTKO organs occurred in 9/21 baboons (43%). hCPRP expression reduced the GalTKO EGF incidence to 7% (1/14; P < 0.01 vs. GalTKO alone). At 30 mins, complement deposits were more intense in organs in which EGF developed (P < 0.005). The intensity of peri-transplant platelet activation (as ?-thromboglobulin release) correlated with EGF, as did the cumulative coagulation score (P < 0.01). We conclude that (i) the transgenic expression of a hCPRP on the vascular endothelium of a GalTKO pig reduces the incidence of EGF and reduces complement deposition, (ii) complement deposition and platelet activation correlate with early GalTKO organ failure, and (iii) the expression of a hCPRP reduces EGF but does not prevent systemic coagulation activation. Additional strategies will be required to control coagulation activation. PMID:26174749

  8. Modulation of glycan detection on specific glycoproteins by lectin multimerization

    PubMed Central

    Cao, Zheng; Partyka, Katie; McDonald, Mitchell; Brouhard, Elizabeth; Hincapie, Marina; Brand, Randall E.; Hancock, William S.; Haab, Brian B.

    2013-01-01

    Improved methods for studying glycans could spur significant advances in the understanding and application of glycobiology. The use of affinity reagents such as lectins and glycan-binding antibodies is a valuable complement to methods involving mass spectrometry and chromatography. Many lectins, however, are not useful as analytic tools due to low affinity in vitro. As an approach to increasing lectin avidity to targeted glycans, we tested the use of lectin multimerization. Several biotinylated lectins were linked together through streptavidin interactions. The binding of certain lectins for purified glycoproteins and glycoproteins captured directly out of biological solutions was increased using multimerization, resulting in the detection of lower concentrations of glycoprotein than possible using monomeric detection. The analysis of glycoproteins in plasma samples showed that the level of binding enhancement through multimerization was not equivalent across patient samples. Wheat germ agglutinin (WGA) reactive glycans on fibronectin and thrombospondin-5 were preferentially bound by multimers in pancreatic cancer patient samples relative to control samples, suggesting a cancer-associated change in glycan density that could be detected only through lectin multimerization. This strategy could lead to the more sensitive and informative detection of glycans in biological samples and a broader spectrum of lectins that are useful as analytical reagents. PMID:23286506

  9. Mannan binding lectin attenuates double-stranded RNA-mediated TLR3 activation and innate immunity.

    PubMed

    Liu, Hongzhi; Zhou, Jia; Ma, Di; Lu, Xiao; Ming, Siqi; Shan, Guiqiu; Zhang, Xiaoyong; Hou, Jinlin; Chen, Zhengliang; Zuo, Daming

    2014-03-18

    Mannan binding lectin (MBL) functions as a pattern recognition molecule (PRM) which is able to initiate complement activation. Here, we characterize a previously unrecognized attribute of MBL as a double-stranded RNA (dsRNA) binding protein capable of modifying Toll like receptor 3 (TLR3) activation. MBL interacts with poly(I:C) and suppresses poly(I:C)-induced activation of TLR3 pathways and subsequent cytokine production. In addition, MBL binds to TLR3 directly. Surprisingly, disrupting the interaction between MBL and complement receptor 1 (CR1) or restraining the traffic of MBL to phagosome reversed the MBL limited TLR3 activation. We demonstrate the importance of MBL guided ligands intracellular localization, emphasizing the significance of understanding the dynamics of TLR agonists complexed with MBL or other PRMs inside the cell in immune defense. PMID:24530528

  10. Sushi domain-containing protein 4 (SUSD4) inhibits complement by disrupting the formation of the classical C3 convertase.

    PubMed

    Holmquist, Emelie; Okroj, Marcin; Nodin, Bjrn; Jirstrm, Karin; Blom, Anna M

    2013-06-01

    Recently discovered Sushi domain-containing protein 4 (SUSD4) contains several Sushi or complement control protein domains; therefore, we hypothesized that it may act as complement inhibitor. Two isoforms of human SUSD4, fused to the Fc part of human IgG, were recombinantly expressed in Chinese hamster ovary (CHO) cells. The secreted soluble isoform of SUSD4 (SUSD4b) inhibited the classical and lectin complement pathways by 50% at a concentration of 0.5 ?M. This effect was due to the fact that 1 ?M SUSD4b inhibited the formation of the classical C3 convertase by 90%. The membrane-bound isoform (SUSD4a) inhibited the classical and alternative complement pathways when expressed on the surface of CHO cells but not when expressed as a soluble, truncated protein. In all functional studies, we used known complement inhibitors as positive controls, while Coxsackie adenovirus receptor, which has no effect on complement, expressed with Fc tag, was a negative control. We also studied the mRNA expression of both isoforms of SUSD4 in a panel of human tissues using quantitative PCR and primarily found SUSD4a in esophagus and brain, while SUSD4b was highly expressed in esophagus, ovary, and heart. Overall, our results show that SUSD4 is a novel complement inhibitor with restricted expression. PMID:23482636

  11. Enzyme treatment of Trypanosoma danilewskyi (Laveran and Mesnil) increases its susceptibility to lysis by the alternative complement pathway of goldfish, Carassius auratus (L.).

    PubMed

    Plouffe, D A; Belosevic, M

    2004-05-01

    This study examined whether in vitro-cultured Trypanosoma danilewskyi were susceptible to lysis in the presence or absence of anti-parasite antibodies and complement. Cultured trypanosomes were resistant to lysis by either immune or non-immune goldfish serum. However, trypanosomes treated with the proteolytic enzyme trypsin, which destroys surface proteins of the parasites, became susceptible to lysis when exposed to either immune or non-immune goldfish serum. The lysis by goldfish serum was dependent on the presence of heat-labile factors and occurred at 4 and 20 degrees C. The lysis was also dependent on the presence of Mg(2+) ions but not Ca(2+) ions. Furthermore, treatment of the parasites with different sialidases did not enhance their susceptibility to lysis by goldfish serum. Trypsinized parasites regained resistance to lysis after at least 6-h cultivation in the absence of trypsin and the restoration of full resistance was observed after 24-h cultivation. The resistance to lysis was abrogated when the protein synthesis inhibitor, puromycin, was added to the cultures. These results suggest that trypsinized trypanosomes were susceptible to lysis by goldfish complement (alternative pathway) and that protective surface proteins of the parasite were required for the resistance of normal trypanosomes to lysis. PMID:15139906

  12. Transcriptome and analysis on the complement and coagulation cascades pathway of large yellow croaker (Larimichthys crocea) to ciliate ectoparasite Cryptocaryon irritans infection.

    PubMed

    Yin, Fei; Gao, Quanxin; Tang, Baojun; Sun, Peng; Han, Kunhuang; Huang, Weiqing

    2016-03-01

    Large yellow croaker (Larimichthys crocea) is one of the most valuable marine fish in southern China. Given to the rapid development of aquaculture industry, the L. crocea was subjected to ciliate ectoparasite Cryptocaryon irritans. It therefore is indispensable and urgent to understand the mechanism of L. crocea host defense against C. irritans infection. In the present study, the extensively analysis at the transcriptome level for Cryptocaryoniasis in L. crocea was carried out. These results showed that 15,826,911, 16,462,921, and 15,625,433 paired-end clean reads were obtained from three cDNA libraries (A: 0 theronts/fish, B: 12,000 theronts/fish, and C: 24,000 theronts/fish) of the L. crocea immune-related tissues by Illumina paired-end sequencing technology. Totally, 30,509 unique transcript fragments (unigenes) were assembled, with an average length of 1715 bp. In B/A, C/A, and C/B pairwise comparison, 972, 900, and 1126 genes showed differential expression respectively. Differently expressed immune-related genes (DEIGs) were scrutinized, in B/A pairwise comparison, 48 genes showed differential expression, including 26 up-regulated genes and 22 down-regulated genes in B; in C/A pairwise comparison, there were 39 DEIGs, including 7 up-regulated genes and 32 down-regulated genes in C; in C/B pairwise comparison, 40 genes showed differential expression, including 11 up-regulated genes and 29 down-regulated genes in C. There were 16 DEIGs enriched KEGG pathways, in which the complement and coagulation cascades pathway was the top most DEIGs enriched pathway (B:A=42; C:A=28; C:B=42). The coagulation and fibrinolytic system was in a highly active state after infected by C. irritans with non-lethal concentration; the alternative complement pathway may play an important role in the early stages of C. irritans infection. These results demonstrated that low-concentration infection can significantly induce the immunological response in fishes, however, when fishes were in fatal conditions, the immunity was suppressed. PMID:26804649

  13. Lectins of marine hydrobionts.

    PubMed

    Chernikov, O V; Molchanova, V I; Chikalovets, I V; Kondrashina, A S; Li, W; Lukyanov, P A

    2013-07-01

    Data from the literature and results of our research on lectins isolated from some kinds of marine hydrobionts such as clams, ascidians, sea worms, sponges, and algae are presented in this review. Results of comparative analysis of the basic physicochemical properties and biological activity of lectins isolated from various sources are discussed. PMID:24010839

  14. Role of complement in glomerular diseases.

    PubMed

    Mao, Song; Zhang, Jianhua

    2016-06-01

    The complement system, composed of nearly 30 proteins, is a key regulator of immunity. The complement system is critical for protecting hosts from invading pathogens. Dysregulation of this system is associated with susceptibility to infection and various autoimmune diseases. Furthermore, complement activation due to the defective regulation of the alternative pathway will induce glomerular diseases. Anti-complement therapy has been applied in various glomerular diseases. Signaling pathways might be very important in the pathogenesis of glomerular diseases. This review will give a relatively complete signaling pathway flowchart for complement and a comprehensive understanding of the underlying role of complement in glomerular diseases. PMID:26400225

  15. CD45-mediated signaling pathway is involved in Rhizoctonia bataticola lectin (RBL)-induced proliferation and Th1/Th2 cytokine secretion in human PBMC

    SciTech Connect

    Pujari, Radha; Eligar, Sachin M.; Kumar, Natesh; Nagre, Nagaraja N.; Inamdar, Shashikala R.; Swamy, Bale M.; Shastry, Padma

    2012-03-23

    Highlights: Black-Right-Pointing-Pointer RBL, a potent mitogenic and complex N-glycan specific lectin binds to CD45 on PBMC. Black-Right-Pointing-Pointer RBL triggers CD45-mediated signaling involved in activation of p38MAPK and STAT-5. Black-Right-Pointing-Pointer Inhibition of CD45 PTPase signaling blocks RBL-induced ZAP70 phosphorylation. Black-Right-Pointing-Pointer RBL-CD45 mediated signaling is crucial for RBL-induced immunodulatory activities. -- Abstract: We earlier reported the mitogenic and immunostimulatory activities of Rhizoctonia bataticola lectin (RBL), purified from phytopathogenic fungus R. bataticola in human PBMC. The lectin demonstrates specificity towards glycoproteins containing complex N-glycans. Since CD45-protein tyrosine phosphatase that abundantly expresses N-glycans is important in T-cell signaling, the study aimed to investigate the involvement of CD45 in the immunomodulatory activities of RBL. Flowcytometry and confocal microscopy studies revealed that RBL exhibited binding to PBMC and colocalized with CD45. The binding was comparable in cells expressing different CD45 isoforms-RA, -RB and -RO. CD45 blocking antibody reduced the binding and proliferation of PBMC induced by RBL. CD45-PTPase inhibitor dephostatin inhibited RBL-induced proliferation, expression of CD25 and pZAP-70. RBL-induced secretion of Th1/Th2 cytokines were significantly inhibited in presence of dephostatin. Also, dephostatin blocked phosphorylation of p38MAPK and STAT-5 that was crucial for the biological functions of RBL. The study demonstrates the involvement of CD45-mediated signaling in RBL-induced PBMC proliferation and Th1/Th2 cytokine secretion through activation of p38MAPK and STAT-5.

  16. Mechanisms of complement activation, C4d deposition, and their contribution to the pathogenesis of antibody mediated rejection

    PubMed Central

    Murata, Kazunori; Baldwin, William M

    2009-01-01

    Complement split products have emerged as useful markers of antibody mediated rejection in solid organ transplants. One split product, C4d, is now widely accepted as a marker for antibody mediated rejection in renal and cardiac allografts. This review summarizes the rationale for the use of C4d as a marker of antibody mediated rejection, along with the clinical evidence supporting its use in the clinical diagnosis of antibody mediated rejection. Antibody-independent mechanisms by which C4d can be activated by the classical and lectin pathways of complement activation are also identified. Finally, mechanisms by which complement activation stimulates effector cells (neutrophils, monocytes, macrophages, platelets, and B and T lymphocytes) as well as target cells (endothelial cells) are discussed in relation to antibody mediated allograft rejection. PMID:19362461

  17. Complement-mediated injury and protection of endothelium: Lessons from atypical haemolytic uraemic syndrome

    PubMed Central

    Kerr, Heather; Richards, Anna

    2014-01-01

    The complement system provides a vital defence against invading pathogens. As an intrinsic system it is always on, in a state of constant, low level activation. This activation is principally mediated through the deposition of C3b on to pathogenic surfaces and host tissues. C3b is generated by spontaneous tick over and formal activation of the alternative pathway, and by activation of the classical and lectin pathways. If the deposited C3b is not appropriately regulated, there is progression to terminal pathway complement activation via the C5 convertases, generating the potent anaphylotoxin C5a and the membrane attack complex C5b-9. Unsurprisingly, these highly active components have the potential to cause injury to bystander host tissue, including the vascular endothelium. As such, complement activation on endothelium is normally tightly controlled by a large number of fluid-phase and membrane bound inhibitors, in an attempt to ensure that propagation of complement activation is appropriately restricted to invading pathogens and altered self, e.g. apoptotic and necrotic cells. The kidney is increasingly recognised as a site at particular risk from complement-mediated endothelial injury. Both genetic and acquired defects which impact on complement regulation predispose to this susceptibility. The thrombotic microangiopathy, haemolytic uraemic syndrome (HUS), will be used to illustrate the mechanisms by which the endothelial cell injury occurs. Finally, the underlying rationale for current and future potential therapeutic interventions in HUS and also the opportunities for enhancing endothelial defence to prevent relapsing disease through increased complement cytoprotective strategies will be summarised. PMID:21855165

  18. Complement Factor C4d Is a Common Denominator in Thrombotic Microangiopathy.

    PubMed

    Chua, Jamie S; Baelde, Hans J; Zandbergen, Malu; Wilhelmus, Suzanne; van Es, Leendert A; de Fijter, Johan W; Bruijn, Jan A; Bajema, Ingeborg M; Cohen, Danielle

    2015-09-01

    Complement activation has a major role in thrombotic microangiopathy (TMA), a disorder that can occur in a variety of clinical conditions. Promising results of recent trials with terminal complement-inhibiting drugs call for biomarkers identifying patients who might benefit from this treatment. The primary aim of this study was to determine the prevalence and localization of complement factor C4d in kidneys of patients with TMA. The secondary aims were to determine which complement pathways lead to C4d deposition and to determine whether complement activation results in deposition of the terminal complement complex. We examined 42 renal sections with histologically confirmed TMA obtained from a heterogeneous patient group. Deposits of C4d, mannose-binding lectin, C1q, IgM, and C5b-9 were scored in the glomeruli, peritubular capillaries, and arterioles. Notably, C4d deposits were present in 88.1% of TMA cases, and the various clinical conditions had distinct staining patterns within the various compartments of the renal vasculature. Classical pathway activation was observed in 90.5% of TMA cases. C5b-9 deposits were present in 78.6% of TMA cases and in 39.6% of controls (n=53), but the staining pattern differed between cases and controls. In conclusion, C4d is a common finding in TMA, regardless of the underlying clinical condition. Moreover, C5b-9 was present in >75% of the TMA samples, suggesting that terminal complement inhibitors may have a beneficial effect in these patients. C4d and C5b-9 should be investigated as possible diagnostic biomarkers in the clinical work-up of patients suspected of having complement-mediated TMA. PMID:25573909

  19. Surface-bound capsular polysaccharide of type Ia group B Streptococcus mediates C1 binding and activation of the classic complement pathway

    SciTech Connect

    Levy, N.J.; Kasper, D.L.

    1986-06-01

    The role of surface-bound type Ia group B Streptococcus (GBS) capsular polysaccharide in anti-body-independent binding of C1 and activation of the classic component pathway was investigated. In a radiolabeled bacterial-polymorphonuclear leukocyte (PMN) association assay, a measure of bacterial opsonization, preincubation of /sup 3/H-type Ia GBS with purified F(ab')/sub 2/ to the organism blocked the association of the bacteria with PMN', and the inhibitory effect was dose dependent. The specificity of F(ab')/sub 2/ blocking was shown after adsorption of F(ab')/sub 2/ with type Ia polysaccharide-sensitized erythrocytes. Polysaccharide-adsorbed F(ab')/sub 2/ had a 70% decrease in ability to block the association of bacteria with PMN. Neuraminidase digestion removed 80% of the terminal sialic acid residues from the native polysaccharide. These neuraminidase-digested organisms had a 72% decrease in binding and transfer of purified C1 compared with non-enzyme-treated organisms. Type Ia capsular polysaccharide bound to sheep erythrocytes promoted classic complement pathway-mediated hemolysis of the cells. The role of C1 inhibitor (INH) in modulation of C1 activation by the organisms was investigated. The possibility existed that the C1 INH could be bound by the bacteria, allowing C1 activation to occur in the fluid phase. The inhibitor was purified from human serum, and its activity was measured before and after incubation with type Ia GBS. The organisms had no effect on C1 INH activity. Thus surface-bound capsular polysacchardie of type Ia GBS mediates C1 binding and classic pathway activation, and this does not involve the C1 INH.

  20. Identification of Complin, a novel complement inhibitor that targets complement proteins factor B and C2.

    PubMed

    Kadam, Archana P; Sahu, Arvind

    2010-06-15

    Complement factor B (fB) is a key constituent of the alternative pathway (AP). Its central role in causing inflammation and tissue injury through activation of the AP urges the need for its therapeutic targeting. In the current study, we have screened phage-displayed random peptide libraries against fB and identified a novel cyclic hendecapeptide that inhibits activation of fB and the AP. Structure-activity studies revealed that: 1) the cysteine-constrained structure of the peptide is essential for its activity; 2) Ile5, Arg6, Leu7, and Tyr8 contribute significantly to its inhibitory activity; and 3) retro-inverso modification of the peptide results in loss of its activity. Binding studies performed using surface plasmon resonance suggested that the peptide has two binding sites on fB, which are located on the Ba and Bb fragments. Studies on the mechanism of inhibition revealed that the peptide does not block the interaction of fB with the activated form of C3, thereby suggesting that the peptide inhibits fB activation primarily by inhibiting its cleavage by factor D. The peptide showed a weak effect on preformed C3 and C5 convertases. Like inhibition of fB cleavage, the peptide also inhibited C2 cleavage by activated C1s and activation of the classical as well as lectin pathways. Based on its inhibitory activities, we named the peptide Complin. PMID:20483772

  1. Inhibition of the Alternative Pathway of Nonhuman Infant Complement by Porin B2 Contributes to Virulence of Neisseria meningitidis in the Infant Rat Model

    PubMed Central

    Vu, David M.; Granoff, Dan M.; Ram, Sanjay

    2014-01-01

    Neisseria meningitidis utilizes capsular polysaccharide, lipooligosaccharide (LOS) sialic acid, factor H binding protein (fHbp), and neisserial surface protein A (NspA) to regulate the alternative pathway (AP) of complement. Using meningococcal mutants that lacked all four of the above-mentioned molecules (quadruple mutants), we recently identified a role for PorB2 in attenuating the human AP; inhibition was mediated by human fH, a key downregulatory protein of the AP. Previous studies showed that fH downregulation of the AP via fHbp or NspA is specific for human fH. Here, we report that PorB2-expressing quadruple mutants also regulate the AP of baby rabbit and infant rat complement. Blocking a human fH binding region on PorB2 of the quadruple mutant of strain 4243 with a chimeric protein that comprised human fH domains 6 and 7 fused to murine IgG Fc enhanced AP-mediated baby rabbit C3 deposition, which provided evidence for an fH-dependent mechanism of nonhuman AP regulation by PorB2. Using isogenic mutants of strain H44/76 that differed only in their PorB molecules, we confirmed a role for PorB2 in resistance to killing by infant rat serum. The PorB2-expressing strain also caused higher levels of bacteremia in infant rats than its isogenic PorB3-expressing counterpart, thus providing a molecular basis for increased survival of PorB2 isolates in this model. These studies link PorB2 expression with infection of infant rats, which could inform the choice of meningococcal strains for use in animal models, and reveals, for the first time, that PorB2-expressing strains of N. meningitidis regulate the AP of baby rabbits and rats. PMID:24686052

  2. Lectins with potential for anti-cancer therapy.

    PubMed

    Yau, Tammy; Dan, Xiuli; Ng, Charlene Cheuk Wing; Ng, Tzi Bun

    2015-01-01

    This article reviews lectins of animal and plant origin that induce apoptosis and autophagy of cancer cells and hence possess the potential of being developed into anticancer drugs. Apoptosis-inducing lectins encompass galectins, C-type lectins, annexins, Haliotis discus discus lectin, Polygonatum odoratum lectin, mistletoe lectin, and concanavalin A, fucose-binding Dicentrarchus labrax lectin, and Strongylocentrotus purpuratus lectin, Polygonatum odoratum lectin, and mistletoe lectin, Polygonatum odoratum lectin, autophagy inducing lectins include annexins and Polygonatum odoratum lectin. PMID:25730388

  3. M. leprae components induce nerve damage by complement activation: identification of lipoarabinomannan as the dominant complement activator.

    PubMed

    Bahia El Idrissi, Nawal; Das, Pranab K; Fluiter, Kees; Rosa, Patricia S; Vreijling, Jeroen; Troost, Dirk; Morgan, B Paul; Baas, Frank; Ramaglia, Valeria

    2015-05-01

    Peripheral nerve damage is the hallmark of leprosy pathology but its etiology is unclear. We previously identified the membrane attack complex (MAC) of the complement system as a key determinant of post-traumatic nerve damage and demonstrated that its inhibition is neuroprotective. Here, we determined the contribution of the MAC to nerve damage caused by Mycobacterium leprae and its components in mouse. Furthermore, we studied the association between MAC and the key M. leprae component lipoarabinomannan (LAM) in nerve biopsies of leprosy patients. Intraneural injections of M. leprae sonicate induced MAC deposition and pathological changes in the mouse nerve, whereas MAC inhibition preserved myelin and axons. Complement activation occurred mainly via the lectin pathway and the principal activator was LAM. In leprosy nerves, the extent of LAM and MAC immunoreactivity was robust and significantly higher in multibacillary compared to paucibacillary donors (p = 0.01 and p = 0.001, respectively), with a highly significant association between LAM and MAC in the diseased samples (r = 0.9601, p = 0.0001). Further, MAC co-localized with LAM on axons, pointing to a role for this M. leprae antigen in complement activation and nerve damage in leprosy. Our findings demonstrate that MAC contributes to nerve damage in a model of M. leprae-induced nerve injury and its inhibition is neuroprotective. In addition, our data identified LAM as the key pathogen associated molecule that activates complement and causes nerve damage. Taken together our data imply an important role of complement in nerve damage in leprosy and may inform the development of novel therapeutics for patients. PMID:25772973

  4. Complement Activation by Merozoite Antigens of Plasmodium falciparum

    PubMed Central

    Korir, Jackson C.; Nyakoe, Nancy K.; Awinda, George; Waitumbi, John N.

    2014-01-01

    Background Complement (C) is a crucial part of the innate immune system and becomes over activated during malaria, resulting in depletion of C components, especially those for lectin pathway (LP), thereby compromising the host's innate defense. In this study, involvement of P. falciparum antigens in C activation was investigated. Methods A highly synchronous culture of the Dd2 clone of P. falciparum was established in a serum free medium. Supernatants harvested from rings, trophozoites and schizonts at various parasite densities were tested for ability to activate C by quantifying amount of C3b deposited on erythrocytes (E). Uninfected sham culture was used as control. Remnants of each C pathway were determined using Wieslab complement System Screenkit (Euro-diagnostica, Sweden). To identify MBL binding antigens of LP, culture supernatants were added to MBL sepharose columns and trapped antigens eluted with increasing concentrations of EDTA (10 mM, 50 mM and 100 mM) and then desalted before being tested for ability to activate C. The EDTA eluate with highest activity was run on a polyacrylamide gel and silver stained proteins analyzed by mass spectroscopy. Results Antigens released by P. falciparum growing in culture activated C leading to C3b deposition on E. Maximal activation at 7% parasitemia was associated with schizont stage (36.7%) compared to 22% for rings, 21% for trophozoites and 3% for sham culture. All the three pathways of C were activated, with highest activation being for the alternative pathway (only 6% of C activation potential remained), 65% for classiical and 43% for the LP. Seven MBL binding merozoite proteins were identified by mass spectrometry in the 50 mM EDTA eluate. Conclusions MBL binding merozoite adhesins with ability to activate C pathway were identified. The survival advantage for such pronounced C activation is unclear, but opsonisation could facilitate recognition and invasion of E. PMID:25144772

  5. Dissection of Functional Sites in Herpesvirus Saimiri Complement Control Protein Homolog

    PubMed Central

    Reza, Malik Johid; Kamble, Ashish; Ahmad, Muzammil; Krishnasastry, Musti V.

    2013-01-01

    Herpesvirus saimiri is known to encode a homolog of human complement regulators named complement control protein homolog (CCPH). We have previously reported that this virally encoded inhibitor effectively inactivates complement by supporting factor I-mediated inactivation of complement proteins C3b and C4b (termed cofactor activity), as well as by accelerating the irreversible decay of the classical/lectin and alternative pathway C3 convertases (termed decay-accelerating activity). To fine map its functional sites, in the present study, we have generated a homology model of CCPH and performed substitution mutagenesis of its conserved residues. Functional analyses of 24 substitution mutants of CCPH indicated that (i) amino acids R118 and F144 play a critical role in imparting C3b and C4b cofactor activities, (ii) amino acids R35, K142, and K191 are required for efficient decay of the C3 convertases, (iii) positively charged amino acids of the linker regions, which are dubbed to be critical for functioning in other complement regulators, are not crucial for its function, and (iv) S100K and G110D mutations substantially enhance its decay-accelerating activities without affecting the cofactor activities. Overall, our data point out that ionic interactions form a major component of the binding interface between CCPH and its interacting partners. PMID:23077301

  6. Binding of flavivirus nonstructural protein NS1 to C4b binding protein modulates complement activation.

    PubMed

    Avirutnan, Panisadee; Hauhart, Richard E; Somnuke, Pawit; Blom, Anna M; Diamond, Michael S; Atkinson, John P

    2011-07-01

    The complement system plays a pivotal protective role in the innate immune response to many pathogens including flaviviruses. Flavivirus nonstructural protein 1 (NS1) is a secreted nonstructural glycoprotein that accumulates in plasma to high levels and is displayed on the surface of infected cells but absent from viral particles. Previous work has defined an immune evasion role of flavivirus NS1 in limiting complement activation by forming a complex with C1s and C4 to promote cleavage of C4 to C4b. In this study, we demonstrate a second mechanism, also involving C4 and its active fragment C4b, by which NS1 antagonizes complement activation. Dengue, West Nile, or yellow fever virus NS1 directly associated with C4b binding protein (C4BP), a complement regulatory plasma protein that attenuates the classical and lectin pathways. Soluble NS1 recruited C4BP to inactivate C4b in solution and on the plasma membrane. Mapping studies revealed that the interaction sites of NS1 on C4BP partially overlap with the C4b binding sites. Together, these studies further define the immune evasion potential of NS1 in reducing the functional capacity of C4 in complement activation and control of flavivirus infection. PMID:21642539

  7. Structural and functional diversity of the lectin repertoire in teleost fish: Relevance to innate and adaptive immunity

    PubMed Central

    Vasta, Gerardo R.; Nita-Lazar, Mihai; Giomarelli, Barbara; Ahmed, Hafiz; Du, Shaojun; Cammarata, Matteo; Parrinello, Nicol; Bianchet, Mario A.; Amzel, L. Mario

    2012-01-01

    Proteincarbohydrate interactions mediated by lectins have been recognized as key components of innate immunity in vertebrates and invertebrates, not only for recognition of potential pathogens, but also for participating in downstream effector functions, such as their agglutination, immobilization, and complement-mediated opsonization and killing. More recently, lectins have been identified as critical regulators of mammalian adaptive immune responses. Fish are endowed with virtually all components of the mammalian adaptive immunity, and are equipped with a complex lectin repertoire. In this review, we discuss evidence suggesting that: (a) lectin repertoires in teleost fish are highly diversified, and include not only representatives of the lectin families described in mammals, but also members of lectin families described for the first time in fish species; (b) the tissue-specific expression and localization of the diverse lectin repertoires and their molecular partners is consistent with their distinct biological roles in innate and adaptive immunity; (c) although some lectins may bind endogenous ligands, others bind sugars on the surface of potential pathogens; (d) in addition to pathogen recognition and opsonization, some lectins display additional effector roles, such as complement activation and regulation of immune functions; (e) some lectins that recognize exogenous ligands mediate processes unrelated to immunity: they may act as anti-freeze proteins or prevent polyspermia during fertilization. PMID:21896283

  8. Lectins: production and practical applications

    PubMed Central

    2010-01-01

    Lectins are proteins found in a diversity of organisms. They possess the ability to agglutinate erythrocytes with known carbohydrate specificity since they have at least one non-catalytic domain that binds reversibly to specific monosaccharides or oligosaccharides. This articles aims to review the production and practical applications of lectins. Lectins are isolated from their natural sources by chromatographic procedures or produced by recombinant DNA technology. The yields of animal lectins are usually low compared with the yields of plant lectins such as legume lectins. Lectins manifest a diversity of activities including antitumor, immunomodulatory, antifungal, HIV-1 reverse transcriptase inhibitory, and anti-insect activities, which may find practical applications. A small number of lectins demonstrate antibacterial and anti-nematode activities. PMID:20890754

  9. A Serine Protease Isolated from the Bristles of the Amazonic Caterpillar, Premolis semirufa, Is a Potent Complement System Activator

    PubMed Central

    Villas Boas, Isadora Maria; Pidde-Queiroz, Giselle; Magnoli, Fabio Carlos; Gonalves-de-Andrade, Rute M.; van den Berg, Carmen W.; Tambourgi, Denise V.

    2015-01-01

    Background The caterpillar of the moth Premolis semirufa, commonly named pararama, is found in the Brazilian Amazon region. Accidental contact with the caterpillar bristles causes an intense itching sensation, followed by symptoms of an acute inflammation, which last for three to seven days after the first incident. After multiple accidents a chronic inflammatory reaction, called Pararamose, characterized by articular synovial membrane thickening with joint deformities common to chronic synovitis, frequently occurs. Although complement mediated inflammation may aid the host defense, inappropriate or excessive activation of the complement system and generation of anaphylatoxins can lead to inflammatory disorder and pathologies. The aim of the present study was to evaluate, in vitro, whether the Premolis semirufas bristles extract could interfere with the human complement system. Results The bristles extract was able to inhibit the haemolytic activity of the alternative pathway, as well as the activation of the lectin pathway, but had no effect on the classical pathway, and this inhibition seemed to be caused by activation and consumption of complement components. The extract induced the production of significant amounts of all three anaphylatoxins, C3a, C4a and C5a, promoted direct cleavage of C3, C4 and C5 and induced a significant generation of terminal complement complexes in normal human serum. By using molecular exclusion chromatography, a serine protease of 82 kDa, which activates complement, was isolated from P. semirufa bristles extract. The protease, named here as Ps82, reduced the haemolytic activity of the alternative and classical pathways and inhibited the lectin pathway. In addition, Ps82 induced the cleavage of C3, C4 and C5 and the generation of C3a and C4a in normal human serum and it was capable to cleave human purified C5 and generate C5a. The use of Phenanthroline, metalloprotease inhibitor, in the reactions did not significantly interfere with the activity of the Ps82, whereas the presence of PMSF, serine protease inhibitor, totally blocked the activity. Conclusion These data show that a serine protease present in the Premolis semirufas bristles extract has the ability to activate the complement system, which may contribute to the inflammatory process presented in humans after envenomation. PMID:25760458

  10. Lectins with anti-HIV activity: a review.

    PubMed

    Akkouh, Ouafae; Ng, Tzi Bun; Singh, Senjam Sunil; Yin, Cuiming; Dan, Xiuli; Chan, Yau Sang; Pan, Wenliang; Cheung, Randy Chi Fai

    2015-01-01

    Lectins including flowering plant lectins, algal lectins, cyanobacterial lectins, actinomycete lectin, worm lectins, and the nonpeptidic lectin mimics pradimicins and benanomicins, exhibit anti-HIV activity. The anti-HIV plant lectins include Artocarpus heterophyllus (jacalin) lectin, concanavalin A, Galanthus nivalis (snowdrop) agglutinin-related lectins, Musa acuminata (banana) lectin, Myrianthus holstii lectin, Narcissus pseudonarcissus lectin, and Urtica diocia agglutinin. The anti-HIV algal lectins comprise Boodlea coacta lectin, Griffithsin, Oscillatoria agardhii agglutinin. The anti-HIV cyanobacterial lectins are cyanovirin-N, scytovirin, Microcystis viridis lectin, and microvirin. Actinohivin is an anti-HIV actinomycete lectin. The anti-HIV worm lectins include Chaetopterus variopedatus polychaete marine worm lectin, Serpula vermicularis sea worm lectin, and C-type lectin Mermaid from nematode (Laxus oneistus). The anti-HIV nonpeptidic lectin mimics comprise pradimicins and benanomicins. Their anti-HIV mechanisms are discussed. PMID:25569520

  11. Model-Driven Redox Pathway Manipulation for Improved Isobutanol Production in Bacillus subtilis Complemented with Experimental Validation and Metabolic Profiling Analysis

    PubMed Central

    Qi, Haishan; Li, Shanshan; Zhao, Sumin; Huang, Di; Xia, Menglei; Wen, Jianping

    2014-01-01

    To rationally guide the improvement of isobutanol production, metabolic network and metabolic profiling analysis were performed to provide global and profound insights into cell metabolism of isobutanol-producing Bacillus subtilis. The metabolic flux distribution of strains with different isobutanol production capacity (BSUL03, BSUL04 and BSUL05) drops a hint of the importance of NADPH on isobutanol biosynthesis. Therefore, the redox pathways were redesigned in this study. To increase NADPH concentration, glucose-6-phosphate isomerase was inactivated (BSUL06) and glucose-6-phosphate dehydrogenase was overexpressed (BSUL07) successively. As expected, NADPH pool size in BSUL07 was 4.4-fold higher than that in parental strain BSUL05. However, cell growth, isobutanol yield and production were decreased by 46%, 22%, and 80%, respectively. Metabolic profiling analysis suggested that the severely imbalanced redox status might be the primary reason. To solve this problem, gene udhA of Escherichia coli encoding transhydrogenase was further overexpressed (BSUL08), which not only well balanced the cellular ratio of NAD(P)H/NAD(P)+, but also increased NADH and ATP concentration. In addition, a straightforward engineering approach for improving NADPH concentrations was employed in BSUL05 by overexpressing exogenous gene pntAB and obtained BSUL09. The performance for isobutanol production by BSUL09 was poorer than BSUL08 but better than other engineered strains. Furthermore, in fed-batch fermentation the isobutanol production and yield of BSUL08 increased by 11% and 19%, up to the value of 6.12 g/L and 0.37 C-mol isobutanol/C-mol glucose (63% of the theoretical value), respectively, compared with parental strain BSUL05. These results demonstrated that model-driven complemented with metabolic profiling analysis could serve as a useful approach in the strain improvement for higher bio-productivity in further application. PMID:24705866

  12. Lectins: getting familiar with translators of the sugar code.

    PubMed

    André, Sabine; Kaltner, Herbert; Manning, Joachim C; Murphy, Paul V; Gabius, Hans-Joachim

    2015-01-01

    The view on the significance of the presence of glycans in glycoconjugates is undergoing a paradigmatic change. Initially mostly considered to be rather inert and passive, the concept of the sugar code identifies glycans as highly versatile platform to store information. Their chemical properties endow carbohydrates to form oligomers with unsurpassed structural variability. Owing to their capacity to engage in hydrogen (and coordination) bonding and C-H/π-interactions these "code words" can be "read" (in Latin, legere) by specific receptors. A distinct class of carbohydrate-binding proteins are the lectins. More than a dozen protein folds have developed carbohydrate-binding capacity in vertebrates. Taking galectins as an example, distinct expression patterns are traced. The availability of labeled endogenous lectins facilitates monitoring of tissue reactivity, extending the scope of lectin histochemistry beyond that which traditionally involved plant lectins. Presentation of glycan and its cognate lectin can be orchestrated, making a glycan-based effector pathway in growth control of tumor and activated T cells possible. In order to unravel the structural basis of lectin specificity for particular glycoconjugates mimetics of branched glycans and programmable models of cell surfaces are being developed by strategic combination of lectin research with synthetic and supramolecular chemistry. PMID:25621423

  13. Role of complement in a murine model of peanut-induced anaphylaxis.

    PubMed

    Kodama, Toshihisa; Sekine, Hideharu; Takahashi, Minoru; Iwaki, Daisuke; Machida, Takeshi; Kanno, Kazuko; Ishida, Yumi; Endo, Yuichi; Fujita, Teizo

    2013-06-01

    Peanut allergy is severe and persisting from childhood to adulthood. However, there is no effective prophylaxis or treatment for peanut allergy. Little is known to about the molecular process in the pathogenesis of peanuts allergy, especially in innate immunity. Thus we investigated the role of complement activation in murine peanut anaphylaxis. Complement component C3 deposition on peanut extract (PE) was evaluated using sera from wild-type (WT), mannose-binding lectin associated serine protease (MASP)-1/3 deficient, MASP-2 deficient, and C4 deficient mice. Sera from interferon regulatory factor-4 (IRF-4) deficient mice, which lack serum immunoglobulin, were also used. In anaphylaxis study, mice were pretreated with propranolol and a long-acting form of IL-4, and injected with PE. Mice were then assessed for plasma C3a levels and hypothermia shock by ELISA and rectal temperature measurement, respectively. C3 deposition on PE was abolished in immunoglobulin- and C4-deficient sera. No difference in C3 deposition levels were observed among WT, MASP-1/3 deficient and MASP-2 deficient sera. IgM, IgG2b, IgG3, C1q, and ficolin-A deposits were detected on PE. In anaphylaxis study, MASP-1/3 deficient mice showed elevation of plasma C3a levels similar to WT mice. However, they were significantly reduced in C4- and MASP-2-deficient mice compared to WT mice. Consistently, PE-induced anaphylactic shock was prevented in C4 deficient mice and partially in MASP-2 deficient mice. In conclusion, PE activates complement via both the lectin and classical pathways in vivo, and the complement activation contributes to hypothermia shock in mice. PMID:23182714

  14. Lectin-dependent enhancement of Ebola virus infection via soluble and transmembrane C-type lectin receptors.

    PubMed

    Brudner, Matthew; Karpel, Marshall; Lear, Calli; Chen, Li; Yantosca, L Michael; Scully, Corinne; Sarraju, Ashish; Sokolovska, Anna; Zariffard, M Reza; Eisen, Damon P; Mungall, Bruce A; Kotton, Darrell N; Omari, Amel; Huang, I-Chueh; Farzan, Michael; Takahashi, Kazue; Stuart, Lynda; Stahl, Gregory L; Ezekowitz, Alan B; Spear, Gregory T; Olinger, Gene G; Schmidt, Emmett V; Michelow, Ian C

    2013-01-01

    Mannose-binding lectin (MBL) is a key soluble effector of the innate immune system that recognizes pathogen-specific surface glycans. Surprisingly, low-producing MBL genetic variants that may predispose children and immunocompromised individuals to infectious diseases are more common than would be expected in human populations. Since certain immune defense molecules, such as immunoglobulins, can be exploited by invasive pathogens, we hypothesized that MBL might also enhance infections in some circumstances. Consequently, the low and intermediate MBL levels commonly found in human populations might be the result of balancing selection. Using model infection systems with pseudotyped and authentic glycosylated viruses, we demonstrated that MBL indeed enhances infection of Ebola, Hendra, Nipah and West Nile viruses in low complement conditions. Mechanistic studies with Ebola virus (EBOV) glycoprotein pseudotyped lentiviruses confirmed that MBL binds to N-linked glycan epitopes on viral surfaces in a specific manner via the MBL carbohydrate recognition domain, which is necessary for enhanced infection. MBL mediates lipid-raft-dependent macropinocytosis of EBOV via a pathway that appears to require less actin or early endosomal processing compared with the filovirus canonical endocytic pathway. Using a validated RNA interference screen, we identified C1QBP (gC1qR) as a candidate surface receptor that mediates MBL-dependent enhancement of EBOV infection. We also identified dectin-2 (CLEC6A) as a potentially novel candidate attachment factor for EBOV. Our findings support the concept of an innate immune haplotype that represents critical interactions between MBL and complement component C4 genes and that may modify susceptibility or resistance to certain glycosylated pathogens. Therefore, higher levels of native or exogenous MBL could be deleterious in the setting of relative hypocomplementemia which can occur genetically or because of immunodepletion during active infections. Our findings confirm our hypothesis that the pressure of infectious diseases may have contributed in part to evolutionary selection of MBL mutant haplotypes. PMID:23573288

  15. Lectin-Dependent Enhancement of Ebola Virus Infection via Soluble and Transmembrane C-type Lectin Receptors

    PubMed Central

    Lear, Calli; Chen, Li; Yantosca, L. Michael; Scully, Corinne; Sarraju, Ashish; Sokolovska, Anna; Zariffard, M. Reza; Eisen, Damon P.; Mungall, Bruce A.; Kotton, Darrell N.; Omari, Amel; Huang, I-Chueh; Farzan, Michael; Takahashi, Kazue; Stuart, Lynda; Stahl, Gregory L.; Ezekowitz, Alan B.; Spear, Gregory T.; Olinger, Gene G.; Schmidt, Emmett V.; Michelow, Ian C.

    2013-01-01

    Mannose-binding lectin (MBL) is a key soluble effector of the innate immune system that recognizes pathogen-specific surface glycans. Surprisingly, low-producing MBL genetic variants that may predispose children and immunocompromised individuals to infectious diseases are more common than would be expected in human populations. Since certain immune defense molecules, such as immunoglobulins, can be exploited by invasive pathogens, we hypothesized that MBL might also enhance infections in some circumstances. Consequently, the low and intermediate MBL levels commonly found in human populations might be the result of balancing selection. Using model infection systems with pseudotyped and authentic glycosylated viruses, we demonstrated that MBL indeed enhances infection of Ebola, Hendra, Nipah and West Nile viruses in low complement conditions. Mechanistic studies with Ebola virus (EBOV) glycoprotein pseudotyped lentiviruses confirmed that MBL binds to N-linked glycan epitopes on viral surfaces in a specific manner via the MBL carbohydrate recognition domain, which is necessary for enhanced infection. MBL mediates lipid-raft-dependent macropinocytosis of EBOV via a pathway that appears to require less actin or early endosomal processing compared with the filovirus canonical endocytic pathway. Using a validated RNA interference screen, we identified C1QBP (gC1qR) as a candidate surface receptor that mediates MBL-dependent enhancement of EBOV infection. We also identified dectin-2 (CLEC6A) as a potentially novel candidate attachment factor for EBOV. Our findings support the concept of an innate immune haplotype that represents critical interactions between MBL and complement component C4 genes and that may modify susceptibility or resistance to certain glycosylated pathogens. Therefore, higher levels of native or exogenous MBL could be deleterious in the setting of relative hypocomplementemia which can occur genetically or because of immunodepletion during active infections. Our findings confirm our hypothesis that the pressure of infectious diseases may have contributed in part to evolutionary selection of MBL mutant haplotypes. PMID:23573288

  16. Complement, interferon and lupus.

    PubMed

    Elkon, Keith B; Santer, Deanna M

    2012-12-01

    The complement pathway was implicated in the immunopathogenesis of lupus and other autoimmune disorders decades ago. The apparent paradox that early complement component (C1q, C2 and C4) deficiencies predispose to lupus has been explained by the beneficial roles of these proteins in promoting the clearance of immune complexes (ICs) and apoptotic cells. Recent findings demonstrate that, in the absence of C1q, instead of ICs binding to monocytes, they preferentially engage plasmacytoid dendritic cells (pDC) so generating interferon (IFN) alpha, the cytokine with potent immune adjuvant properties. C1q opsonized apoptotic cells also exert an immunosuppressive effect through cytokine regulation and the stimulation of additional opsonins by macrophages. C1q was recently reported to impede neutrophil extracellular trap (NET) degradation. NETs are known to promote type I IFN production in SLE by providing a source of antigen for the formation of ICs as well as through direct pDC activation by cathelicidin (LL37). Together, these findings provide both direct and indirect links between two key pathways implicated in lupus pathogenesis: complement and IFN. PMID:22999705

  17. Microencapsulation of lectin anti-cancer agent and controlled release by alginate beads, biosafety approach.

    PubMed

    El-Aassar, M R; Hafez, Elsayed E; El-Deeb, Nehal M; Fouda, Moustafa M G

    2014-08-01

    Hepatocellular carcinoma (HCC) is considered as one of the most aggressive cancer worldwide. In Egypt, the prevalence of HCC is increasing during last years. Recently, drug-loaded microparticles were used to improve the efficiency of various medical treatments. This study is designed to evaluate the anticancer potentialities of lectins against HCC while hinting to its safety usage. The aim is also extended to encapsulate lectins in alginate microbeads for oral drug delivery purposes. The extracted lectins showed anti-proliferative effect against HCC with a percentage of 60.76% by using its nontoxic dose with an up-regulation of P53 gene expression. Concerning the handling of lectin alginate microbeads for oral drug delivery, the prepared lectin alginate beads were ?100?m in diameter. The efficiency of the microcapsules was checked by scanning electron microscopy, the SEM showed the change on the alginate beads surface revealing the successful lectin encapsulation. The release of lectins from the microbeads depended on a variety of factors as the microbeads forming carriers and the amount-encapsulated lectins. The Pisum sativum extracted lectins may be considered as a promising agent in controlling HCC and this solid dosage form could be suitable for oral administration complemented with/or without the standard HCC drugs. PMID:24857870

  18. Banana lectin: a brief review.

    PubMed

    Singh, Senjam Sunil; Devi, Sanjenbam Kunjeshwori; Ng, Tzi Bun

    2014-01-01

    Lectins are a group of proteins of non-immune origin that recognize and bind to carbohydrates without modifying them. Banana is the common name for both herbaceous plants of the genus Musa and for the fruit they produce. They are indeed a promising source for many medicinal applications. Banana lectins have the potential for inhibiting HIV-1 reverse transcriptase activity, suppressing cancer cell proliferation and stimulating macrophage activities. Nevertheless, compared to other plant lectins, there is relatively little information in the literature on banana lectins, particularly with respect to their structure and biological functions. Herein we focus our review on the structure, functions and exploitable properties of banana lectins. PMID:25407720

  19. Antagonism of the complement component C4 by flavivirus nonstructural protein NS1.

    PubMed

    Avirutnan, Panisadee; Fuchs, Anja; Hauhart, Richard E; Somnuke, Pawit; Youn, Soonjeon; Diamond, Michael S; Atkinson, John P

    2010-04-12

    The complement system plays an essential protective role in the initial defense against many microorganisms. Flavivirus NS1 is a secreted nonstructural glycoprotein that accumulates in blood, is displayed on the surface of infected cells, and has been hypothesized to have immune evasion functions. Herein, we demonstrate that dengue virus (DENV), West Nile virus (WNV), and yellow fever virus (YFV) NS1 attenuate classical and lectin pathway activation by directly interacting with C4. Binding of NS1 to C4 reduced C4b deposition and C3 convertase (C4b2a) activity. Although NS1 bound C4b, it lacked intrinsic cofactor activity to degrade C4b, and did not block C3 convertase formation or accelerate decay of the C3 and C5 convertases. Instead, NS1 enhanced C4 cleavage by recruiting and activating the complement-specific protease C1s. By binding C1s and C4 in a complex, NS1 promotes efficient degradation of C4 to C4b. Through this mechanism, NS1 protects DENV from complement-dependent neutralization in solution. These studies define a novel immune evasion mechanism for restricting complement control of microbial infection. PMID:20308361

  20. Complement and systemic lupus erythematosus

    PubMed Central

    2002-01-01

    Chapter summary Complement is implicated in the pathogenesis of systemic lupus erythematosus (SLE) in several ways and may act as both friend and foe. Homozygous deficiency of any of the proteins of the classical pathway is causally associated with susceptibility to the development of SLE, especially deficiency of the earliest proteins of the activation pathway. However, complement is also implicated in the effector inflammatory phase of the autoimmune response that characterizes the disease. Complement proteins are deposited in inflamed tissues and, in experimental models, inhibition of C5 ameliorates disease in a murine model. As a further twist to the associations between the complement system and SLE, autoantibodies to some complement proteins, especially to C1q, develop as part of the autoantibody response. The presence of anti-C1q autoantibodies is associated with severe illness, including glomerulonephritis. In this chapter the role of the complement system in SLE is reviewed and hypotheses are advanced to explain the complex relationships between complement and lupus. PMID:12110148

  1. Could plant lectins become promising anti-tumour drugs for causing autophagic cell death?

    PubMed

    Liu, Z; Luo, Y; Zhou, T-T; Zhang, W-Z

    2013-10-01

    Plant lectins, a group of highly diverse carbohydrate-binding proteins of non-immune origin, are ubiquitously distributed through a variety of plant species, and have recently drawn rising attention due to their remarkable ability to kill tumour cells using mechanisms implicated in autophagy. In this review, we provide a brief outline of structures of some representative plant lectins such as concanavalin A, Polygonatum cyrtonema lectin and mistletoe lectins. These can target autophagy by modulating BNIP-3, ROS-p38-p53, Ras-Raf and PI3KCI-Akt pathways, as well as Beclin-1, in many types of cancer cells. In addition, we further discuss how plant lectins are able to kill cancer cells by modulating autophagic death, for therapeutic purposes. Together, these findings provide a comprehensive perspective concerning plant lectins as promising new anti-tumour drugs, with respect to autophagic cell death in future cancer therapeutics. PMID:24033443

  2. Effect of lectins on the transport of food ingredients in Caco-2 cell cultures.

    PubMed

    Ohno, Y; Naganuma, T; Ogawa, T; Muramoto, K

    2004-01-01

    We investigated the effect of several lectins, such as soy bean lectin (SBA), concanavalin A (Con A), and wheat germ agglutinin (WGA), on the transport of some food ingredients (isoflavones, quercetin glycosides, carnosine/anserine) across Caco-2 cell monolayers. After incubation of food ingredients (0.03 approximately 2 mmol/L) in the presence or absence of lectins (1 approximately 180 microg/ml) on the apical side, aliquots were taken from the apical and basolateral solution, and were subjected to HPLC analysis. We also examined the effect of lectins on the permeability of the tight junction by measuring the transepithelial electrical resistance (TER) value of the Caco-2 cell monolayer. Isoflavones, which was not transported to the basolateral solution without lectins, could be transported in the presence of lectins, whereas their aglycones were detected at the same levels with or without the lectin treatment. The transport of quercetin glycosides also increased in the presence of lectins, however, that of peptides was not affected by the lectins. Con A and WGA, but SBA, decreased the TER value, indicating that Con A and WGA increased the transport via paracellular pathway, whereas SBA did via a different pathway. PMID:15630235

  3. Mannose-Binding Lectin Gene Polymorphism and Chronic Hepatitis B Infection in Children

    PubMed Central

    Erdemir, Gulin; Ozkan, Tanju B.; Ozgur, Taner; Budak, Ferah; Kilic, Sara S.; Onay, Huseyin

    2015-01-01

    Background/Aims: Mannose-binding lectin (MBL) is a member of innate immune system that activates complement system through lectin pathway. MBL deficiency is associated with susceptibility to infectious diseases. In this study, the relation between MBL gene polymorphism and chronic hepatitis B infection in children is evaluated. Patients and Methods: The study included 67 children with chronic hepatitis B and 99 healthy controls. The hepatitis B patients were divided into immuntolerant, chronic inactive, and treatment groups according to their laboratory findings. MBL gene codon 52, 54, and 57 polymorphisms were studied with polymerase chain reaction in all patients and controls. The associations of MBL gene polymorphism with clinical, laboratory, and histopathologic findings were evaluated. Results: Homozygous codon 54 polymorphism of MBL was found significantly higher in chronic hepatitis B patients than controls. Rate of the polymorphism was similar in all groups and, responsive and nonresponsive patients in the treatment group. Conclusions: The hepatitis B patients who are homozygous for codon 54 of MBL are prone to develop chronic infection. Longitudinal studies with larger groups are needed. PMID:25843194

  4. Mannose-binding lectin and the balance between immune protection and complication

    PubMed Central

    Takahashi, Kazue

    2012-01-01

    The innate immune system is evolutionarily ancient and biologically primitive. Historically, it was first identified as an element of the immune system that provides the first-line response to pathogens, and increasingly it is recognized for its central housekeeping role and its essential functions in tissue homeostasis, including coagulation and inflammation, among others. A pivotal link between the innate immune system and other functions is mannose-binding lectin (MBL), a pattern recognition molecule. Multiple studies have demonstrated that MBL deficiency increases susceptibility to infection, and the mechanisms associated with this susceptibility to infection include reduced opsonophagocytic killing and reduced activation of the lectin complement pathway. Results from our laboratory have demonstrated that MBL and MBL-associated serine protease (MASP)-1/3 together mediate coagulation factor-like activities, including thrombin-like activity. MBL and/or MASP-1/3-deficient hosts demonstrate in vivo evidence that MBL and MASP-1/3 are involved with hemostasis following injury. Staphylococcus aureus-infected MBL null mice developed disseminated intravascular coagulation, which was associated with elevated blood IL-6 levels (but not TNF-?) and systemic inflammatory responses. Infected MBL null mice also develop liver injury. These findings suggest that MBL deficiency may manifest as disseminated intravascular coagulation and organ failure with infection. Beginning from these observations, this review focuses on the interaction of innate immunity and other homeostatic systems, the derangement of which may lead to complications in infection and other inflammatory states. PMID:22114968

  5. Lysyl Hydroxylase 3 Modifies Lysine Residues to Facilitate Oligomerization of Mannan-Binding Lectin

    PubMed Central

    Risteli, Maija; Ruotsalainen, Heli; Bergmann, Ulrich; Venkatraman Girija, Umakhanth; Wallis, Russell; Myllyl, Raili

    2014-01-01

    Lysyl hydroxylase 3 (LH3) is a multifunctional protein with lysyl hydroxylase, galactosyltransferase and glucosyltransferase activities. The LH3 has been shown to modify the lysine residues both in collagens and also in some collagenous proteins. In this study we show for the first time that LH3 is essential for catalyzing formation of the glucosylgalactosylhydroxylysines of mannan-binding lectin (MBL), the first component of the lectin pathway of complement activation. Furthermore, loss of the terminal glucose units on the derivatized lysine residues in mouse embryonic fibroblasts lacking the LH3 protein leads to defective disulphide bonding and oligomerization of rat MBL-A, with a decrease in the proportion of the larger functional MBL oligomers. The oligomerization could be completely restored with the full length LH3 or the amino-terminal fragment of LH3 that possesses the glycosyltransferase activities. Our results confirm that LH3 is the only enzyme capable of glucosylating the galactosylhydroxylysine residues in proteins with a collagenous domain. In mice lacking the lysyl hydroxylase activity of LH3, but with untouched galactosyltransferase and glucosyltransferase activities, reduced circulating MBL-A levels were observed. Oligomerization was normal, however and residual lysyl hydroxylation was compensated in part by other lysyl hydroxylase isoenzymes. Our data suggest that LH3 is commonly involved in biosynthesis of collagenous proteins and the glucosylation of galactosylhydroxylysines residues by LH3 is crucial for the formation of the functional high-molecular weight MBL oligomers. PMID:25419660

  6. The Serine Protease Pic From Enteroaggregative Escherichia coli Mediates Immune Evasion by the Direct Cleavage of Complement Proteins.

    PubMed

    Abreu, Afonso G; Fraga, Tatiana R; Granados Martnez, Adriana P; Kondo, Marcia Y; Juliano, Maria A; Juliano, Luiz; Navarro-Garcia, Fernando; Isaac, Lourdes; Barbosa, Angela S; Elias, Waldir P

    2015-07-01

    Enteroaggregative and uropathogenic Escherichia coli, Shigella flexneri 2a, and the hybrid enteroaggregative/Shiga toxin-producing E. coli strain (O104:H4) are important pathogens responsible for intestinal and urinary tract infections, as well as sepsis and hemolytic uremic syndrome. They have in common the production of a serine protease called Pic. Several biological roles for Pic have been described, including protection of E. coli DH5? from complement-mediated killing. Hereby we showed that Pic significantly reduces complement activation by all 3 pathways. Pic cleaves purified C3/C3b and other proteins from the classic and lectin pathways, such as C4 and C2. Cleavage fragments of C3, C4, and C2 were also observed with HB101(pPic1) culture supernatants, and C3 cleavage sites were mapped by fluorescence resonance energy transfer peptides. Experiments using human serum as a source of complement proteins confirmed Pic proteolytic activity on these proteins. Furthermore, Pic works synergistically with the human complement regulators factor I and factor H, promoting inactivation of C3b. In the presence of both regulators, further degradation of C3 ?' chain was observed. Therefore, Pic may contribute to immune evasion of E. coli and S. flexneri, favoring invasiveness and increasing the severity of the disorders caused by these pathogens. PMID:25583166

  7. Lectins in human pathogenic fungi.

    PubMed

    Gallegos, Belém; Martínez, Ruth; Pérez, Laura; Del Socorro Pina, María; Perez, Eduardo; Hernández, Pedro

    2014-01-01

    Lectins are carbohydrate-binding proteins widely distributed in nature. They constitute a highly diverse group of proteins consisting of many different protein families that are, in general, structurally unrelated. In the last few years, mushroom and other fungal lectins have attracted wide attention due to their antitumour, antiproliferative and immunomodulatory activities. The present mini-review provides concise information about recent developments in understanding lectins from human pathogenic fungi. A bibliographic search was performed in the Science Direct and PubMed databases, using the following keywords "lectin", "fungi", "human" and "pathogenic". Lectins present in fungi have been classified; however, the role played by lectins derived from human pathogenic fungi in infectious processes remains uncertain; thus, this is a scientific field requiring more research. This manuscript is part of the series of works presented at the "V International Workshop: Molecular genetic approaches to the study of human pathogenic fungi" (Oaxaca, Mexico, 2012). PMID:24270074

  8. Critical roles of complement and antibodies in host defense mechanisms against Neisseria meningitidis as revealed by human complement genetic deficiencies.

    PubMed

    Hellerud, Bernt Christian; Aase, Audun; Herstad, Tove Karin; Naess, Lisbeth Meyer; Kristiansen, Lisa Høyem; Trøseid, Anne-Marie Siebke; Harboe, Morten; Lappegård, Knut Tore; Brandtzaeg, Petter; Høiby, E Arne; Mollnes, Tom Eirik

    2010-02-01

    Certain complement defects are associated with an increased propensity to contract Neisseria meningitidis infections. We performed detailed analyses of complement-mediated defense mechanisms against N. meningitidis 44/76 with whole blood and serum from two adult patients who were completely C2 or C5 deficient. The C5-deficient patient and the matched control were also deficient in mannose-binding lectin (MBL). The proliferation of meningococci incubated in freshly drawn whole blood was estimated by CFU and quantitative DNA real-time PCR. The serum bactericidal activity and opsonophagocytic activity by granulocytes were investigated, including heat-inactivated postvaccination sera, to examine the influence of antimeningococcal antibodies. The meningococci proliferated equally in C2- and C5-deficient blood, with a 2 log(10) increase of CFU and 4- to 5-log(10) increase in DNA copies. Proliferation was modestly decreased in reconstituted C2-deficient and control blood. After reconstitution of C5-deficient blood, all meningococci were killed, which is consistent with high antibody titers being present. The opsonophagocytic activity was strictly C2 dependent, appeared with normal serum, and increased with postvaccination serum. Serum bactericidal activity was strictly dependent on C2, C5, and high antibody titers. MBL did not influence any of the parameters observed. Complement-mediated defense against meningococci was thus dependent on the classical pathway. Some opsonophagocytic activity occurred despite low levels of antimeningococcal antibodies but was more efficient with immune sera. Serum bactericidal activity was dependent on C2, C5, and immune sera. MBL did not influence any of the parameters observed. PMID:19933829

  9. Complement fixation by rheumatoid factor.

    PubMed Central

    Tanimoto, K; Cooper, N R; Johnson, J S; Vaughan, J H

    1975-01-01

    The capacity for fixation and activation of hemolytic complement by polyclonal IgM rheumatoid factors (RF) isolated from sera of patients with rheumatoid arthritis and monoclonal IgM-RF isolated from the cryoprecipitates of patients with IgM-IgG mixed cryoglobulinemia was examined. RF mixed with aggregated, reduced, and alkylated human IgG (Agg-R/A-IgG) in the fluid phase failed to significantly reduce the level of total hemolytic complement, CH50, or of individual complement components, C1, C2, C3, and C5. However, sheep erythrocytes (SRC) coated with Agg-R/A-IgG or with reduced and alkylated rabbit IgG anti-SRC antibody were hemolyzed by complement in the presence of polyclonal IgM-RF. Human and guinea pig complement worked equally well. The degree of hemolysis was in direct proportion to the hemagglutination titer of the RF against the same coated cells. Monoclonal IgM-RF, normal human IgM, and purified Waldenstrm macroglobulins without antiglobulin activity were all inert. Hemolysis of coated SRC by RF and complement was inhibited by prior treatment of the complement source with chelating agents, hydrazine, cobra venom factor, specific antisera to C1q, CR, C5, C6, or C8, or by heating at 56 degrees C for 30 min. Purified radiolabeled C4, C3, and C8 included in the complement source were bound to hemolysed SRC in direct proportion to the degree of hemolysis. These data indicate that polyclonal IgM-RF fix and activate complement via the classic pathway. The system described for assessing complement fixation by isolated RF is readily adaptable to use with whole human serum. PMID:1078825

  10. The outer membrane protease PgtE of Salmonella enterica interferes with the alternative complement pathway by cleaving factors B and H

    PubMed Central

    Riva, Rauna; Korhonen, Timo K.; Meri, Seppo

    2015-01-01

    The virulence factor PgtE is an outer membrane protease (omptin) of the zoonotic pathogen Salmonella enterica that causes diseases ranging from gastroenteritis to severe enteric fever. It is surface exposed in bacteria that have a short-chain, i.e., rough LPS, as observed e.g., in bacteria residing inside macrophages or just emerging from them. We investigated whether PgtE cleaves the complement factors B (B) and H (H), key proteins controlling formation and inactivation of the complement protein C3b and thereby the activity of the complement system. S. enterica serovar Typhimurium or omptin-expressing recombinant E. coli bacteria were incubated with purified human complement proteins or recombinant H fragments. PgtE cleaved both B and H, whereas its close homolog Pla of Yersinia pestis cleaved only H. H was cleaved at both N- and C-termini, while the central region resisted proteolysis. Because of multiple effects of PgtE on complement components (cleavage of C3, C3b, B, and H) we assessed its effect on the opsonophagocytosis of Salmonella. In human serum, C3 cleavage was dependent on proteolytically active PgtE. Human neutrophils interacted less with serum-opsonized FITC-stained S. enterica 14028R than with the isogenic ?pgtE strain, as analyzed by flow cytometry. In conclusion, cleavage of B and H by PgtE, together with C3 cleavage, affects the C3-mediated recognition of S. enterica by human neutrophils, thus thwarting the immune protection against Salmonella. PMID:25705210

  11. Lectins and the radioallergosorbent test.

    PubMed

    Barnett, D; Howden, M E

    1987-10-01

    An investigation into the possible role of lectin binding of IgE in RAST of legume and wheat extracts is reported. Lectins from pea, broad bean, lentil, jack bean, soybean, peanut, and wheat germ were coupled to RAST discs. The discs were pretreated with lectin-specific sugars in an attempt to inhibit RAST with sera from 11 sensitive patients. In all cases, RAST was almost unaffected by the inhibitory sugars, indicating that nonimmune binding of IgE by lectins in legume or wheat RAST was not significant when RAST was carried out with allergens bound to the usual paper discs. IgE contains binding sites for all the lectins examined, and five of the sera had high total IgE greater than 300 IU/ml. It is suggested that competition by IgG and other serum glycoproteins may explain the lack of effect by the added sugars. All sera contained endogenous glucose at a level of about 3 mmol/L that may have accounted for some self-inhibition of the lectin binding by pea, broad bean, lentil, and jack bean lectins. There was, however, significant immune binding of some of the lectins by specific IgE, and it is concluded that these lectins may be important in expression of IgE-mediated allergic responses. PMID:3668119

  12. Characterization and expression analysis of a complement component gene in sea cucumber ( Apostichopus japonicus)

    NASA Astrophysics Data System (ADS)

    Chen, Zhong; Zhou, Zunchun; Yang, Aifu; Dong, Ying; Guan, Xiaoyan; Jiang, Bei; Wang, Bai

    2015-12-01

    The complement system plays a crucial role in the innate immune system of animals. It can be activated by distinct yet overlapping classical, alternative and lectin pathways. In the alternative pathway, complement factor B (Bf) serves as the catalytic subunit of complement component 3 (C3) convertase, which plays the central role among three activation pathways. In this study, the Bf gene in sea cucumber ( Apostichopus japonicus), termed AjBf, was obtained by rapid amplification of cDNA ends (RACE). The full-length cDNA of AjBf was 3231 bp in length barring the poly (A) tail. It contained an open reading frame (ORF) of 2742 bp encoding 913 amino acids, a 105 bp 5'-UTR (5'-terminal untranslated region) and a 384 bp 3'-UTR. AjBf was a mosaic protein with six CCP (complement control protein) domains, a VWA (von Willebrand factor A) domain, and a serine protease domain. The deduced molecular weight of AjBf protein was 101 kDa. Quantitative real time PCR (qRT-PCR) analysis indicated that the expression level of AjBf in A. japonicus was obviously higher at larval stage than that at embryonic stage. Expression detection in different tissues showed that AjBf expressed higher in coelomocytes than in other four tissues. In addation, AjBf expression in different tissues was induced significantly after LPS or PolyI:C challenge. These results indicated that AjBf plays an important role in immune responses to pathogen infection.

  13. Functional Characterization of LcpA, a Surface-Exposed Protein of Leptospira spp. That Binds the Human Complement Regulator C4BP▿

    PubMed Central

    Barbosa, Angela S.; Monaris, Denize; Silva, Ludmila B.; Morais, Zenaide M.; Vasconcellos, Sílvio A.; Cianciarullo, Aurora M.; Isaac, Lourdes; Abreu, Patricia A. E.

    2010-01-01

    We have previously shown that pathogenic leptospiral strains are able to bind C4b binding protein (C4BP). Surface-bound C4BP retains its cofactor activity, indicating that acquisition of this complement regulator may contribute to leptospiral serum resistance. In the present study, the abilities of seven recombinant putative leptospiral outer membrane proteins to interact with C4BP were evaluated. The protein encoded by LIC11947 interacted with this human complement regulator in a dose-dependent manner. The cofactor activity of C4BP bound to immobilized recombinant LIC11947 (rLIC11947) was confirmed by detecting factor I-mediated cleavage of C4b. rLIC11947 was therefore named LcpA (for leptospiral complement regulator-acquiring protein A). LcpA was shown to be an outer membrane protein by using immunoelectron microscopy, cell surface proteolysis, and Triton X-114 fractionation. The gene coding for LcpA is conserved among pathogenic leptospiral strains. This is the first characterization of a Leptospira surface protein that binds to the human complement regulator C4BP in a manner that allows this important regulator to control complement system activation mediated either by the classical pathway or by the lectin pathway. This newly identified protein may play a role in immune evasion by Leptospira spp. and may therefore represent a target for the development of a human vaccine against leptospirosis. PMID:20404075

  14. Fungal lectins: a growing family.

    PubMed

    Kobayashi, Yuka; Kawagishi, Hirokazu

    2014-01-01

    Fungi are members of a large group of eukaryotic organisms that include yeasts and molds, as well as the most familiar member, mushrooms. Fungal lectins with unique specificity and structures have been discovered. In general, fungal lectins are classified into specific families based on their amino acid sequences and three-dimensional structures. In this chapter, we provide an overview of the approximately 80 types of mushroom and fungal lectins that have been isolated and studied to date. In particular, we have focused on ten fungal lectins (Agaricus bisporus, Agrocybe cylindracea, Aleuria aurantia, Aspergillus oryzae, Clitocybe nebularis, Marasmius oreades, Psathyrella velutina, Rhizopus stolonifer, Pholiota squarrosa, Polyporus squamosus), many of which are commercially available and their properties, sugar-binding specificities, structural grouping into families, and applications for biological research being described. The sialic acid-specific lectins (Agrocybe cylindracea and Polyporus squamosus) and fucose-specific lectins (Aleuria aurantia, Aspergillus oryzae, Rhizopus stolonifer, and Pholiota squarrosa) each showed potential for use in identifying sialic acid glycoconjugates and fucose glycoconjugates. Although not much is currently known about fungal lectins compared to animal and plant lectins, the knowledge accumulated thus far shows great promise for several applications in the fields of taxonomy, biomedicine, and molecular and cellular biology. PMID:25117221

  15. Role of complement in experiment silicosis

    SciTech Connect

    Callis, A.H.; Sohnle, P.G.; Mandel, G.S.; Mandel, N.S.

    1986-08-01

    The role of the complement system in the pathogenesis of crystal-induced pulmonary inflammation and fibrosis was evaluated using a mouse model of silicosis and congenitally complement-deficient mice. Mice lacking the fifth component of complement (B10.D2/o) were compared to C5-sufficient animals (B10.D2/n) for pulmonary changes following intratracheal instillation of silica crystals. Complement-deficient mice demonstrated a significant reduction compared to complement-sufficient mice in both cell number and protein content of lung lavage fluid throughout the 12 weeks following silica exposure. Lung hydroxyproline content (indicative of collagen deposition) was equivalent for both strains and significantly higher than controls at all times points following silica instillation. Moreover, studies in vitro have shown that silica crystals are capable of activating complement via the alternative pathway. These studies indicate that the complement system may be responsible for some of the pulmonary inflammation, but not fibrosis elicited by silica exposure.

  16. Meningococcal disease and the complement system

    PubMed Central

    Lewis, Lisa A; Ram, Sanjay

    2014-01-01

    Despite considerable advances in the understanding of the pathogenesis of meningococcal disease, this infection remains a major cause of morbidity and mortality globally. The role of the complement system in innate immune defenses against invasive meningococcal disease is well established. Individuals deficient in components of the alternative and terminal complement pathways are highly predisposed to invasive, often recurrent meningococcal infections. Genome-wide analysis studies also point to a central role for complement in disease pathogenesis. Here we review the pathophysiologic events pertinent to the complement system that accompany meningococcal sepsis in humans. Meningococci use several often redundant mechanisms to evade killing by human complement. Capsular polysaccharide and lipooligosaccharide glycan composition play critical roles in complement evasion. Some of the newly described protein vaccine antigens interact with complement components and have sparked considerable research interest. PMID:24104403

  17. The novel complement inhibitor human CUB and Sushi multiple domains 1 (CSMD1) protein promotes factor I-mediated degradation of C4b and C3b and inhibits the membrane attack complex assembly.

    PubMed

    Escudero-Esparza, Astrid; Kalchishkova, Nikolina; Kurbasic, Emila; Jiang, Wen G; Blom, Anna M

    2013-12-01

    CUB and Sushi multiple domains 1 (CSMD1) is a transmembrane protein containing 15 consecutive complement control protein (CCP) domains, which are characteristic for complement inhibitors. We expressed a membrane-bound fragment of human CSMD1 composed of the 15 C-terminal CCP domains and demonstrated that it inhibits deposition of C3b by the classical pathway on the surface of Chinese hamster ovary cells by 70% at 6% serum and of C9 (component of membrane attack complex) by 90% at 1.25% serum. Furthermore, this fragment of CSMD1 served as a cofactor to factor I-mediated degradation of C3b. In all functional assays performed, well-characterized complement inhibitors were used as positive controls, whereas Coxsackie adenovirus receptor, a protein with no effect on complement, was a negative control. Moreover, attenuation of expression in human T47 breast cancer cells that express endogenous CSMD1 significantly increased C3b deposition on these cells by 45% at 8% serum compared with that for the controls. Furthermore, by expressing a soluble 17-21 CCP fragment of CSMD1, we found that CSMD1 inhibits complement by promoting factor I-mediated C4b/C3b degradation and inhibition of MAC assembly at the level of C7. Our results revealed a novel complement inhibitor for the classical and lectin pathways. PMID:23964079

  18. Immune evasion by acquisition of complement inhibitors: the mould Aspergillus binds both factor H and C4b binding protein.

    PubMed

    Vogl, G; Lesiak, I; Jensen, D B; Perkhofer, S; Eck, R; Speth, C; Lass-Flrl, C; Zipfel, P F; Blom, A M; Dierich, M P; Wrzner, R

    2008-03-01

    Pathogenic fungi represent a major threat particularly to immunocompromised hosts, leading to severe, and often lethal, systemic opportunistic infections. Although the impaired immune status of the host is clearly the most important factor leading to disease, virulence factors of the fungus also play a role. Factor H (FH) and its splice product FHL-1 represent the major fluid phase inhibitors of the alternative pathway of complement, whereas C4b-binding protein (C4bp) is the main fluid phase inhibitor of the classical and lectin pathways. Both proteins can bind to the surface of various human pathogens conveying resistance to complement destruction and thus contribute to their pathogenic potential. We have recently shown that Candida albicans evades complement by binding both Factor H and C4bp. Here we show that moulds such as Aspergillus spp. bind Factor H, the splicing variant FHL-1 and also C4bp. Immunofluorescence and flow cytometry studies show that the binding of Factor H and C4bp to Aspergillus spp. appears to be even stronger than to Candida spp. and that different, albeit possibly nearby, binding moieties mediate this surface attachment. PMID:17915330

  19. Analysis of common bean expressed sequence tags identifies sulfur metabolic pathways active in seed and sulfur-rich proteins highly expressed in the absence of phaseolin and major lectins

    PubMed Central

    2011-01-01

    Background A deficiency in phaseolin and phytohemagglutinin is associated with a near doubling of sulfur amino acid content in genetically related lines of common bean (Phaseolus vulgaris), particularly cysteine, elevated by 70%, and methionine, elevated by 10%. This mostly takes place at the expense of an abundant non-protein amino acid, S-methyl-cysteine. The deficiency in phaseolin and phytohemagglutinin is mainly compensated by increased levels of the 11S globulin legumin and residual lectins. Legumin, albumin-2, defensin and albumin-1 were previously identified as contributing to the increased sulfur amino acid content in the mutant line, on the basis of similarity to proteins from other legumes. Results Profiling of free amino acid in developing seeds of the BAT93 reference genotype revealed a biphasic accumulation of gamma-glutamyl-S-methyl-cysteine, the main soluble form of S-methyl-cysteine, with a lag phase occurring during storage protein accumulation. A collection of 30,147 expressed sequence tags (ESTs) was generated from four developmental stages, corresponding to distinct phases of gamma-glutamyl-S-methyl-cysteine accumulation, and covering the transitions to reserve accumulation and dessication. Analysis of gene ontology categories indicated the occurrence of multiple sulfur metabolic pathways, including all enzymatic activities responsible for sulfate assimilation, de novo cysteine and methionine biosynthesis. Integration of genomic and proteomic data enabled the identification and isolation of cDNAs coding for legumin, albumin-2, defensin D1 and albumin-1A and -B induced in the absence of phaseolin and phytohemagglutinin. Their deduced amino acid sequences have a higher content of cysteine than methionine, providing an explanation for the preferential increase of cysteine in the mutant line. Conclusion The EST collection provides a foundation to further investigate sulfur metabolism and the differential accumulation of sulfur amino acids in seed of common bean. Identification of sulfur-rich proteins whose levels are elevated in seed lacking phaseolin and phytohemagglutinin and sulfur metabolic genes may assist the improvement of protein quality. PMID:21615926

  20. Mannan-Binding Lectin Is Involved in the Protection against Renal Ischemia/Reperfusion Injury by Dietary Restriction

    PubMed Central

    Shushimita, Shushimita; van der Pol, Pieter; W.F. de Bruin, Ron; N. M. Ijzermans, Jan; van Kooten, Cees; Dor, Frank J. M. F.

    2015-01-01

    Preoperative fasting and dietary restriction offer robust protection against renal ischemia/reperfusion injury (I/RI) in mice. We recently showed that Mannan-binding lectin (MBL), the initiator of the lectin pathway of complement activation, plays a pivotal role in renal I/RI. Based on these findings, we investigated the effect of short-term DR (30% reduction of total food intake) or three days of water only fasting on MBL in 10–12 weeks old male C57/Bl6 mice. Both dietary regimens significantly reduce the circulating levels of MBL as well as its mRNA expression in liver, the sole production site of MBL. Reconstitution of MBL abolished the protection afforded by dietary restriction, whereas in the fasting group the protection persisted. These data show that modulation of MBL is involved in the protection against renal I/RI induced by dietary restriction, and suggest that the mechanisms of protection induced by dietary restriction and fasting may be different. PMID:26367533

  1. Mannan-Binding Lectin Is Involved in the Protection against Renal Ischemia/Reperfusion Injury by Dietary Restriction.

    PubMed

    Shushimita, Shushimita; van der Pol, Pieter; W F de Bruin, Ron; N M Ijzermans, Jan; van Kooten, Cees; Dor, Frank J M F

    2015-01-01

    Preoperative fasting and dietary restriction offer robust protection against renal ischemia/reperfusion injury (I/RI) in mice. We recently showed that Mannan-binding lectin (MBL), the initiator of the lectin pathway of complement activation, plays a pivotal role in renal I/RI. Based on these findings, we investigated the effect of short-term DR (30% reduction of total food intake) or three days of water only fasting on MBL in 10-12 weeks old male C57/Bl6 mice. Both dietary regimens significantly reduce the circulating levels of MBL as well as its mRNA expression in liver, the sole production site of MBL. Reconstitution of MBL abolished the protection afforded by dietary restriction, whereas in the fasting group the protection persisted. These data show that modulation of MBL is involved in the protection against renal I/RI induced by dietary restriction, and suggest that the mechanisms of protection induced by dietary restriction and fasting may be different. PMID:26367533

  2. Complement C5a-C5aR interaction enhances MAPK signaling pathway activities to mediate renal injury in trichloroethylene sensitized BALB/c mice.

    PubMed

    Zhang, Jia-Xiang; Zha, Wan-Sheng; Ye, Liang-Ping; Wang, Feng; Wang, Hui; Shen, Tong; Wu, Chang-Hao; Zhu, Qi-Xing

    2016-02-01

    We have previously shown complement activation as a possible mechanism for trichloroethylene (TCE) sensitization, leading to multi-organ damage including the kidneys. In particular, excessive deposition of C5 and C5b-9-the membrane attack complex, which can generate significant tissue damage, was observed in the kidney tissue after TCE sensitization. The present study tested the hypothesis that anaphylatoxin C5a binding to its receptor C5aR mediates renal injury in TCE-sensitized BALB/c mice. BALB/c mice were sensitized through skin challenge with TCE, with or without pretreatment by the C5aR antagonist W54011. Kidney histopathology and the renal functional test were performed to assess renal injury, and immunohistochemistry and fluorescent labeling were carried out to assess C5a and C5aR expressions. TCE sensitization up-regulated C5a and C5aR expressions in kidney tissue, generated inflammatory infiltration, renal tubule damage, glomerular hypercellularity and impaired renal function. Antagonist pretreatment blocked C5a binding to C5aR and attenuated TCE-induced tissue damage and renal dysfunction. TCE sensitization also caused the deposition of major pro-inflammatory cytokines IL-2, TNF-? and IFN-? in the kidney tissue (P?complement-mediated renal injury by sensitization with TCE or other environmental chemicals. Copyright 2015 John Wiley & Sons, Ltd. PMID:26095957

  3. Lectins in the investigation of receptors

    NASA Astrophysics Data System (ADS)

    Lakhtin, V. M.; Yamskov, Igor A.

    1991-08-01

    Problems of the purification and characterisation are considered for approximately 270 receptors (including cell surface and organelle enzymes), which are glycoconjugates (mainly glycoproteins) from animals, plants and microorganisms, using various lectins (mainly lectin sorbents). An analysis has been carried out of the stages of lectin affinity chromatography of receptors (choice of detergent, use of organic solvents, elution with carbohydrates, etc.). Examples are given of procedures for the purification of receptors, including the use of paired columns and combination chromatography on lectins. The possibility of separating sub-populations of receptors using lectins has been demonstrated. Examples are given of the use of lectins in the analysis of the oligosaccharide structure of receptors. Cases are recorded of the interaction of receptors with endogenous lectins and of receptor lectins with endogenous glycoconjugates. It has been shown that lectins, in combination with glycosidases and antibodies, may be useful in the investigation of receptors. The bibliography contains 406 references.

  4. Use of lectins in immunohematology

    PubMed Central

    Gorakshakar, Ajit C.; Ghosh, Kanjaksha

    2016-01-01

    Lectins are carbohydrate binding proteins present in seeds of many plants, especially corals and beans, in fungi and bacteria, and in animals. Apart from their hemagglutinating property, a wide range of functions have been attributed to them. Their importance in the area of immunohematology is immense. They are used to detect specific red cell antigens, to activate different types of lymphocytes, in order to resolve problems related to polyagglutination and so on. The introduction of advanced biotechnological tools generates new opportunities to exploit the properties of lectins, which were not used earlier. Stem cell research is a very important area in transplant medicine. Certain lectins detect surface markers of stem cell. Hence, they are used to understand the developmental biology of stem cells. The role of various lectins in the areas of transfusion and transplant medicine is discussed in detail in this review. PMID:27011665

  5. A review of fish lectins.

    PubMed

    Ng, Tzi Bun; Fai Cheung, Randy Chi; Wing Ng, Charlene Cheuk; Fang, Evandro Fei; Wong, Jack Ho

    2015-01-01

    Lectins have been reported from various tissues of a diversity of fish species including Japanese eel, conger eel, electric eel, bighead carp, gibel carp, grass carp, Arabian Gulf catfish, channel catfish, blue catfish, catfish, pike perch, perch, powan, zebrafish, toxic moray, cobia fish, steelhead trout, Japanese trout, Atlantic salmon, chinook salmon, olive rainbow smelt, rainbow smelt, white-spotted charr, tilapia, blue gourami, ayu, Potca fish, Spanish mackerel, gilt head bream, tench, roach, rudd, common skate, and sea lamprey. The tissues from which the lectins were isolated comprise gills, eggs, electric organ, stomach, intestine, and liver. Lectins have also been isolated from skin, mucus serum, and plasma. The lectins differ in molecular weight, number of subunits, glycosylation, sugar binding specificity and amino acid sequence. Their activities include antimicrobial, antitumor, immunoregulatory and a role in development. PMID:25929869

  6. Infectious diseases associated with complement deficiencies.

    PubMed Central

    Figueroa, J E; Densen, P

    1991-01-01

    The complement system consists of both plasma and membrane proteins. The former influence the inflammatory response, immune modulation, and host defense. The latter are complement receptors, which mediate the cellular effects of complement activation, and regulatory proteins, which protect host cells from complement-mediated injury. Complement activation occurs via either the classical or the alternative pathway, which converge at the level of C3 and share a sequence of terminal components. Four aspects of the complement cascade are critical to its function and regulation: (i) activation of the classical pathway, (ii) activation of the alternative pathway, (iii) C3 convertase formation and C3 deposition, and (iv) membrane attack complex assembly and insertion. In general, mechanisms evolved by pathogenic microbes to resist the effects of complement are targeted to these four steps. Because individual complement proteins subserve unique functional activities and are activated in a sequential manner, complement deficiency states are associated with predictable defects in complement-dependent functions. These deficiency states can be grouped by which of the above four mechanisms they disrupt. They are distinguished by unique epidemiologic, clinical, and microbiologic features and are most prevalent in patients with certain rheumatologic and infectious diseases. Ethnic background and the incidence of infection are important cofactors determining this prevalence. Although complement undoubtedly plays a role in host defense against many microbial pathogens, it appears most important in protection against encapsulated bacteria, especially Neisseria meningitidis but also Streptococcus pneumoniae, Haemophilus influenzae, and, to a lesser extent, Neisseria gonorrhoeae. The availability of effective polysaccharide vaccines and antibiotics provides an immunologic and chemotherapeutic rationale for preventing and treating infection in patients with these deficiencies. PMID:1889047

  7. Effects of MASP-1 of the Complement System on Activation of Coagulation Factors and Plasma Clot Formation

    PubMed Central

    Hess, Katharina; Ajjan, Ramzi; Phoenix, Fladia; Dob, Jzsef; Gl, Pter; Schroeder, Verena

    2012-01-01

    Background Numerous interactions between the coagulation and complement systems have been shown. Recently, links between coagulation and mannan-binding lectin-associated serine protease-1 (MASP-1) of the complement lectin pathway have been proposed. Our aim was to investigate MASP-1 activation of factor XIII (FXIII), fibrinogen, prothrombin, and thrombin-activatable fibrinolysis inhibitor (TAFI) in plasma-based systems, and to analyse effects of MASP-1 on plasma clot formation, structure and lysis. Methodology/Principal Findings We used a FXIII incorporation assay and specific assays to measure the activation products prothrombin fragment F1+2, fibrinopeptide A (FPA), and activated TAFI (TAFIa). Clot formation and lysis were assessed by turbidimetric assay. Clot structure was studied by scanning electron microscopy. MASP-1 activated FXIII and, contrary to thrombin, induced FXIII activity faster in the Val34 than the Leu34 variant. MASP-1-dependent generation of F1+2, FPA and TAFIa showed a dose-dependent response in normal citrated plasma (NCP), albeit MASP-1 was much less efficient than FXa or thrombin. MASP-1 activation of prothrombin and TAFI cleavage were confirmed in purified systems. No FPA generation was observed in prothrombin-depleted plasma. MASP-1 induced clot formation in NCP, affected clot structure, and prolonged clot lysis. Conclusions/Significance We show that MASP-1 interacts with plasma clot formation on different levels and influences fibrin structure. Although MASP-1-induced fibrin formation is thrombin-dependent, MASP-1 directly activates prothrombin, FXIII and TAFI. We suggest that MASP-1, in concerted action with other complement and coagulation proteins, may play a role in fibrin clot formation. PMID:22536427

  8. Complement activating cryoglobulins in the nephritis of systemic lupus erythematosus.

    PubMed Central

    Adu, D; Williams, D G

    1984-01-01

    Complement activation in vitro by cryoglobulins isolated from the sera of 28 patients with systemic lupus erythematosus (SLE) was examined by incubating the cryoglobulin with normal human serum and performing crossed-immunoelectrophoresis of the mixture to detect C3 conversion. Eighteen of the 28 SLE cryoglobulins activated complement; eight by the classical pathway, four by the alternative pathway exclusively, and six by both pathways. In contrast only two out of 20 cryoglobulins isolated from the sera of normal subjects activated complement and both did so by the classical pathway. Twenty-three of the 28 SLE sera activated complement and complement activating cryoglobulins were isolated from 15 of these 23 sera. The parent sera of cryoglobulins activating complement had lower C4 and C3 concentrations than sera whose cryoglobulins did not split complement but these differences were not significant. The ability of SLE cryoglobulins to activate complement in vitro suggests that these immune complexes activate complement in vivo and thus may contribute to tissue damage in this disease. The activation of both classical and alternative complement pathways is in keeping with other evidence that both pathways are involved in SLE. PMID:6705265

  9. Critical Roles of Complement and Antibodies in Host Defense Mechanisms against Neisseria meningitidis as Revealed by Human Complement Genetic Deficiencies ▿

    PubMed Central

    Hellerud, Bernt Christian; Aase, Audun; Herstad, Tove Karin; Næss, Lisbeth Meyer; Kristiansen, Lisa Høyem; Trøseid, Anne-Marie Siebke; Harboe, Morten; Lappegård, Knut Tore; Brandtzæg, Petter; Høiby, E. Arne; Mollnes, Tom Eirik

    2010-01-01

    Certain complement defects are associated with an increased propensity to contract Neisseria meningitidis infections. We performed detailed analyses of complement-mediated defense mechanisms against N. meningitidis 44/76 with whole blood and serum from two adult patients who were completely C2 or C5 deficient. The C5-deficient patient and the matched control were also deficient in mannose-binding lectin (MBL). The proliferation of meningococci incubated in freshly drawn whole blood was estimated by CFU and quantitative DNA real-time PCR. The serum bactericidal activity and opsonophagocytic activity by granulocytes were investigated, including heat-inactivated postvaccination sera, to examine the influence of antimeningococcal antibodies. The meningococci proliferated equally in C2- and C5-deficient blood, with a 2 log10 increase of CFU and 4- to 5-log10 increase in DNA copies. Proliferation was modestly decreased in reconstituted C2-deficient and control blood. After reconstitution of C5-deficient blood, all meningococci were killed, which is consistent with high antibody titers being present. The opsonophagocytic activity was strictly C2 dependent, appeared with normal serum, and increased with postvaccination serum. Serum bactericidal activity was strictly dependent on C2, C5, and high antibody titers. MBL did not influence any of the parameters observed. Complement-mediated defense against meningococci was thus dependent on the classical pathway. Some opsonophagocytic activity occurred despite low levels of antimeningococcal antibodies but was more efficient with immune sera. Serum bactericidal activity was dependent on C2, C5, and immune sera. MBL did not influence any of the parameters observed. PMID:19933829

  10. Complement-targeted therapeutics

    PubMed Central

    Ricklin, Daniel; Lambris, John D

    2010-01-01

    The complement system is a central component of innate immunity and bridges the innate to the adaptive immune response. However, it can also turn its destructive capabilities against host cells and is involved in numerous diseases and pathological conditions. Modulation of the complement system has been recognized as a promising strategy in drug discovery, and a large number of therapeutic modalities have been developed. However, successful marketing of complement-targeted drugs has proved to be more difficult than initially expected, and many strategies have been discontinued. The US Food and Drug Administrations approval of the first complement-specific drug, an antibody against complement component C5 (eculizumab; Soliris), in March 2007, was a long-awaited breakthrough in the field. Approval of eculizumab validates the complement system as therapeutic target and might facilitate clinical development of other promising drug candidates. PMID:17989689

  11. Complement activation in progressive renal disease

    PubMed Central

    Fearn, Amy; Sheerin, Neil Stephen

    2015-01-01

    Chronic kidney disease (CKD) is common and the cause of significant morbidity and mortality. The replacement of functioning nephrons by fibrosis is characteristic of progressive disease. The pathways that lead to fibrosis are not fully understood, although chronic non-resolving inflammation in the kidney is likely to drive the fibrotic response that occurs. In patients with progressive CKD there is histological evidence of inflammation in the interstitium and strategies that reduce inflammation reduce renal injury in pre-clinical models of CKD. The complement system is an integral part of the innate immune system but also augments adaptive immune responses. Complement activation is known to occur in many diverse renal diseases, including glomerulonephritis, thrombotic microangiopathies and transplant rejection. In this review we discuss current evidence that complement activation contributes to progression of CKD, how complement could cause renal inflammation and whether complement inhibition would slow progression of renal disease. PMID:25664245

  12. [Structure and function of complement protein C1q and its role in the development of autoimmune diseases].

    PubMed

    Smykał-Jankowiak, Katarzyna; Niemir, Zofia I

    2009-01-01

    Complement plays an important role in the immune system. Three different pathways of complement activation are known: the classical, alternative, and lectin dependent. They involve more than 30 serum peptides. C1q is the fi rst subcomponent of the classical pathway of complement activation.It is composed of three types of chains, A, B, and C, which form a molecule containing 18 peptides. Each of the chains has a short amino-terminal region followed by a collagen-like region(playing a role in the activation of C1r2C1s2) and a carboxy-terminal head, which binds to immune complexes. Recent studies have shown a great number of ligands for C1q, including aggregated IgG, IgM, human T-cell lymphotropic virus-I (HTLV-I), gp21 peptide, human immunodeficiency virus-1 (HIV-1) gp21 peptide, beta-amyloid, fragments of bacterial walls, apoptotic cells, and many others. However, the role of C1q is not only associated with complement activation.It also helps in the removal of immune complexes and necrotic cells, stimulates the production of some cytokines, and modulates the function of lymphocytes. Complete C1q deficiency is a rare genetic disorder. The C1q gene is located on the short arm of chromosome 1. So far, only a few mutations in C1q gene have been reported. The presence of these mutations is strongly associated with recurrent bacterial infections and the development of systemic lupus erythematosus(SLE). Recent clinical studies point to the significance of anti-C1q antibodies in the diagnosis and assessment of lupus nephritis activity. PMID:19373194

  13. Structural insights on complement activation.

    PubMed

    Alcorlo, Martn; Lpez-Perrote, Andrs; Delgado, Sandra; Ybenes, Hugo; Subas, Marta; Rodrguez-Gallego, Csar; Rodrguez de Crdoba, Santiago; Llorca, Oscar

    2015-10-01

    The proteolytic cleavage of C3 to generate C3b is the central and most important step in the activation of complement, a major component of innate immunity. The comparison of the crystal structures of C3 and C3b illustrates large conformational changes during the transition from C3 to C3b. Exposure of a reactive thio-ester group allows C3b to bind covalently to surfaces such as pathogens or apoptotic cellular debris. The displacement of the thio-ester-containing domain (TED) exposes hidden surfaces that mediate the interaction with complement factor B to assemble the C3-convertase of the alternative pathway (AP). In addition, the displacement of the TED and its interaction with the macroglobulin 1 (MG1) domain generates an extended surface in C3b where the complement regulators factor H (FH), decay accelerating factor (DAF), membrane cofactor protein (MCP) and complement receptor 1 (CR1) can bind, mediating accelerated decay of the AP C3-convertase and proteolytic inactivation of C3b. In the last few years, evidence has accumulated revealing that the structure of C3b in solution is significantly more flexible than anticipated. We review our current knowledge on C3b structural flexibility to propose a general model where the TED can display a collection of conformations around the MG ring, as well as a few specialized positions where the TED is held in one of several fixed locations. Importantly, this conformational heterogeneity in C3b impacts complement regulation by affecting the interaction with regulators. PMID:26250513

  14. Sentential Complementation in Romanian.

    ERIC Educational Resources Information Center

    Sauer, Keith

    This paper explores the syntactic properties, in Romanian, of one kind of subordination, namely Sentential Predicate Complementation. Some generalizations are offered concerning the relationship between the meaning and the syntactic properties of these constructions. The complement structures are isolated into groups according to verb selection:

  15. Lectin histochemistry of rabbit nephron.

    PubMed

    Castagnaro, M

    In order to investigate the usefulness of lectin histochemistry to detail nephronal segmentation we used 12 different biotinylated lectins (Con-A, DBA, GS-I, LCA, PNA, PWN, RCA-I, RCA-II, SWGA, SBA, UEA-I, and WGA) and Avidin-Biotin-Peroxidase (ABC) system on formalin-fixed and paraffin-embedded rabbit kidney sections. Each lectin, except UEA-I which did not stain any nephron structure, shows a different staining pattern along the nephron. Con-A, LCA, and RCA-I display a diffuse staining, while BS-I, RCA-II, SWGA, PWN, DBA, SBA and PNA are selective markers for specific nephron tracts. Furthermore, it is possible, according to the WGA binding pattern, to differentiate the convoluted part of the proximal tubule into two parts, named Segment A and Segment B. Lectin histochemistry on formalin-fixed and paraffin-embedded rabbit kidney sections displays a specific binding pattern along the rabbit nephron and shows interesting morphofunctional correlations. PMID:2091802

  16. Role of complement and complement regulatory proteins in the complications of diabetes.

    PubMed

    Ghosh, Pamela; Sahoo, Rupam; Vaidya, Anand; Chorev, Michael; Halperin, Jose A

    2015-06-01

    It is well established that the organ damage that complicates human diabetes is caused by prolonged hyperglycemia, but the cellular and molecular mechanisms by which high levels of glucose cause tissue damage in humans are still not fully understood. The prevalent hypothesis explaining the mechanisms that may underlie the pathogenesis of diabetes complications includes overproduction of reactive oxygen species, increased flux through the polyol pathway, overactivity of the hexosamine pathway causing intracellular formation of advanced glycation end products, and activation of protein kinase C isoforms. In addition, experimental and clinical evidence reported in past decades supports a strong link between the complement system, complement regulatory proteins, and the pathogenesis of diabetes complications. In this article, we summarize the body of evidence that supports a role for the complement system and complement regulatory proteins in the pathogenesis of diabetic vascular complications, with specific emphasis on the role of the membrane attack complex (MAC) and of CD59, an extracellular cell membrane-anchored inhibitor of MAC formation that is inactivated by nonenzymatic glycation. We discuss a pathogenic model of human diabetic complications in which a combination of CD59 inactivation by glycation and hyperglycemia-induced complement activation increases MAC deposition, activates pathways of intracellular signaling, and induces the release of proinflammatory, prothrombotic cytokines and growth factors. Combined, complement-dependent and complement-independent mechanisms induced by high glucose promote inflammation, proliferation, and thrombosis as characteristically seen in the target organs of diabetes complications. PMID:25859860

  17. Molecular cloning and characterisation of two homologues of Mannose-Binding Lectin in rainbow trout.

    PubMed

    Nikolakopoulou, Konstantina; Zarkadis, Ioannis K

    2006-09-01

    Mannose-binding lectin (MBL) is a C-type lectin which participates in the innate immune system as an activator of the complement system and as opsonin after binding to certain carbohydrate structures on microorganisms and pathogens. C-type lectins are all Ca(2+)-dependent molecules and they share a tightly folded carbohydrate recognition domain (CRD). In this report the isolation and characterisation of cDNA transcripts encoding two mannose-binding lectin isoforms MBL-1 and MBL-2 from rainbow trout (Oncorhynchus mykiss) is presented. The deduced amino acid sequences of trout MBL-1 and MBL-2 (185 and 186 aa, respectively) present 83% identity to each other, exhibiting the highest identity score 46, 46 and 42% with the Atlantic salmon, shishamo smelt and zebrafish counterparts, respectively. The identity to birds and mammalian MBLs ranges from 25 to 33%. The trout MBL-1 and MBL-2 contain the EPN motif of mannose-binding C-type lectins, important for mannose specificity and they are expressed exclusively in liver and spleen, respectively. PMID:16542855

  18. Cytotoxic ribosome-inactivating lectins from plants.

    PubMed

    Hartley, M R; Lord, J M

    2004-09-01

    A class of heterodimeric plant proteins consisting of a carbohydrate-binding B-chain and an enzymatic A-chain which act on ribosomes to inhibit protein synthesis are amongst the most toxic substances known. The best known example of such a toxic lectin is ricin, produced by the seeds of the castor oil plant, Ricinnus communis. For ricin to reach its substrate in the cytosol, it must be endocytosed, transported through the endomembrane system to reach the compartment from which it is translocated into the cytosol, and there avoid degradation making it possible for a few molecules to inactivate a large proportion of the ribosomes and hence kill the cell. Cell entry by ricin involves the following steps: (i) binding to cell-surface glycolipids and glycoproteins bearing beta-1,4-linked galactose residues through the lectin activity of the B-chain (RTB); (ii) uptake by endocytosis and entry into early endosomes; (iii) transfer by vesicular transport to the trans-Golgi network; (iv) retrograde vesicular transport through the Golgi complex and into the endoplasmic reticulum (ER); (v) reduction of the disulfide bond connecting the A- and B-chains; (vi) a partial unfolding of the A-chain (RTA) to enable it to translocate across the ER membrane via the Sec61p translocon using the pathway normally followed by misfolded ER proteins for targeting to the ER-associated degradation (ERAD) machinery; (vi) refolding in the cytosol into a protease-resistant, enzymatically active structure; (vii) interaction with the sarcin-ricin domain (SRD) of the large ribosome subunit RNA followed by cleavage of a single N-glycosidic bond in the RNA to generate a depurinated, inactive ribosome. In addition to the highly specific action on ribosomes, ricin and related ribosome-inactivating proteins (RIPs) have a less specific action in vitro on DNA and RNA substrates releasing multiple adenine, and in some instances, guanine residues. This polynucleotide:adenosine glycosidase activity has been implicated in the general antiviral, and specifically, the anti HIV-1 activity of several single-chain RIPs which are homologous to the A-chains of the heterodimeric lectins. However, in the absence of clear cause and effect evidence in vivo, such claims should be regarded with caution. PMID:15450171

  19. Lectin cDNA and transgenic plants derived therefrom

    DOEpatents

    Raikhel, Natasha V.

    2000-10-03

    Transgenic plants containing cDNA encoding Gramineae lectin are described. The plants preferably contain cDNA coding for barley lectin and store the lectin in the leaves. The transgenic plants, particularly the leaves exhibit insecticidal and fungicidal properties.

  20. Interaction of lectins with Cryptosporidium parvum.

    PubMed

    Llovo, J; Lopez, A; Fabregas, J; Muñoz, A

    1993-06-01

    Cell surface carbohydrates from four clinical isolates of Cryptosporidium parvum were analyzed by agglutination assays using a battery of 20 highly purified lectins with affinity for receptor molecules containing N-acetyl-D-glucosamine (GlcNAc), N-acetyl-D-galactosamine, galactose, mannose, glucose, fucose, and N-acetyl-neuraminic acid. Tomentine, a lectin from the green seaweed Codium tomentosum, and UEA-II lectin, from Ulex europeus, both of them GlcNAc-specific lectins, agglutinated the oocysts. Subsequent inhibition assay confirmed the presence of this sugar on the surface of Cryptosporidium parvum oocysts. Codium fragile lectin, from another green seaweed, also exhibited agglutination activity against the oocysts. This is the first published demonstration of such an interaction between a human coccidian and lectins from seaweeds. PMID:8501345

  1. Recombinant expression of the autocatalytic complement protease MASP-1 is crucially dependent on co-expression with its inhibitor, C1 inhibitor.

    PubMed

    Degn, Sren E; Thiel, Steffen; Jensenius, Jens C

    2013-04-01

    MASP-1 is a protease of the lectin pathway of complement. It is homologous with MASP-2, previously thought both necessary and sufficient for lectin pathway activation. Recently MASP-1 has taken centre stage with the observation that it is crucial to the activation of MASP-2 and thus central to complement activation. Numerous additional functions have been suggested for MASP-1 and its importance is obvious. Yet, thorough analyses of proteolytic activities and physiological roles in the human scenario have been hampered by difficulties in purifying or producing full-length human MASP-1. We present the successful expression of full-length recombinant human MASP-1 entirely in the zymogen form in a mammalian expression system. We found that the catalytic activity of MASP-1 suppresses its expression through rapid auto-activation and auto-degradation. This auto-degradation was not inhibited by the addition of inhibitors to the culture medium, and it was subsequently found to occur intracellularly. Numerous mutations aimed at attenuating auto-activation or preventing auto-degradation failed to rescue expression, as did also attempts at stabilizing the protease by co-expression with MBL or ficolins or expression in hepatocyte cell lines, representing the natural site of synthesis. The active protease was finally produced through co-expression with the serine protease inhibitor C1 inhibitor. We demonstrate that the expressed protease is capable of binding MBL and auto-activating, and is catalytically active. We have generalized the concept to the expression also of MASP-2 entirely in its zymogen form and with improved yields. We suggest a general advantage of expressing aggressive, autocatalytic proteases with their cognate inhibitors. PMID:23314348

  2. Plant as a plenteous reserve of lectin

    PubMed Central

    Hivrale, AU; Ingale, AG

    2013-01-01

    Lectins are clusters of glycoproteins of nonimmune foundation that combine specifically and reversibly to carbohydrates, mainly the sugar moiety of glycoconjugates, resulting in cell agglutination and precipitation of glycoconjugates. They are universally distributed in nature, being established in plants, fungi, viruses, bacteria, crustacea, insects, and animals, but leguminacae plants are rich source of lectins. The present review reveals the structure, biological properties, and application of plant lectins. PMID:24084524

  3. Lectins and their application to clinical microbiology.

    PubMed Central

    Slifkin, M; Doyle, R J

    1990-01-01

    Lectins are generally associated with plant or animal components, selectively bind carbohydrates, and interact with procaryotic and eucaryotic cells. Lectins have various specificities that are associated with their ability to interact with acetylaminocarbohydrates, aminocarbohydrates, sialic acids, hexoses, pentoses, and as other carbohydrates. Microbial surfaces generally contain many of the sugar residues that react with lectins. Lectins are presently used in the clinical laboratory to type blood cells and are used in a wide spectrum of applications, including, in part, as carriers of chemotherapeutic agents, as mitogens, for fractionation of animal cells, and for investigations of cellular surfaces. Numerous studies have shown that lectins can be used to identify rapidly certain microorganisms isolated from a clinical specimen or directly in a clinical specimen. Lectins have been demonstrated to be important diagnostic reagents in the major realms of clinical microbiology. Thus, they have been applied in bacteriology, mycology, mycobacteriology, and virology for the identification and/or differentiation of various microorganisms. Lectins have been used successfully as epidemiologic as well as taxonomic markers of specific microorganisms. Lectins provide the clinical microbiologist with cost-effective and potential diagnostic reagents. This review describes the applications of lectins in clinical microbiology. Images PMID:2200603

  4. Extreme High Prevalence of a Defective Mannose-Binding Lectin (MBL2) Genotype in Native South American West Andean Populations

    PubMed Central

    Sandoval, Jos Raul; Madsen, Hans O.; De Stefano, Gianfranco; Descailleaux-Dulanto, Jaime; Velazquez-Reinoso, Margarita; ique, Cesar; Fujita, Ricardo; Garred, Peter

    2014-01-01

    Mannose-binding lectin (MBL) is one of the five recognition molecules in the lectin complement pathway. Common variant alleles in the promoter and structural regions of the human MBL gene (MBL2) influence the stability and serum concentration of the protein. Epidemiological studies have shown that MBL2 variant alleles are associated with susceptibility to and the course of different types of infectious and inflammatory conditions. However, it has been suggested that these alleles are maintained in different populations due to selected advantages for carriers. We investigated the MBL2 allelic variation in indigenous individuals from 12 different West Central South America localities spanning from the desert coast, high altitude Andean plates and the Amazon tropical forest within the territories of Peru (n?=?249) (Departments of Loreto, Ucayali, Lambayeque, Junin, Ayacucho, Huancayo and Puno), and Ecuador (n?=?182) (Region of Esmeraldas and Santo Domingo de los Colorados). The distribution of MBL2 genotypes among the populations showed that the defective variant LYPB haplotype was very common. It showed the highest frequencies in Puno (Taquile (0.80), Amantani (0.80) and Anapia (0.58) islander communities of the Lake Titicaca), but lower frequencies of 0.22 in Junin (Central Andean highland) and Ucayali (Central Amazonian forest), as well as 0.27 and 0.24 in the Congoma and Cayapa/Chachis populations in the Amazonian forest in Ecuador were also observed. Our results suggest that the high prevalence of the MBL2 LYPB variant causing low levels of functional MBL in serum may mainly reflect a random distribution due to a population bottleneck in the founder populations. PMID:25313559

  5. Enhanced complement consumption in neuromyelitis optica and Behet's disease patients.

    PubMed

    Tzn, Erdem; Krtnc, Murat; Trko?lu, Recai; Iz, Sema; Pehlivan, Mnevver; Biri?ik, Omer; Eraksoy, Mefkre; Akman-Demir, Gulsen

    2011-04-01

    The complement system is essential in the pathogenesis of inflammatory central nervous system disorders. To investigate the involvement of complement pathways in neuromyelitis optica (NMO), levels of breakdown products for classical (C4d), alternative (FBb) and common (sC5b-9) pathways were measured in the sera of 28 NMO and control patients (30 Behet's disease (BD), 29 multiple sclerosis (MS)) and 31 healthy controls by ELISA. Classical and/or alternative pathway consumption was enhanced in NMO and BD patients as compared to MS patients and healthy controls. Our results suggest that NBD and NMO differ from MS by the predominance of complement system involvement. PMID:21215465

  6. Outline of Hungarian Complementation.

    ERIC Educational Resources Information Center

    Szamosi, Michael

    This study presents a preliminary analysis of Hungarian complement constructions and the syntactic operations needed to account for them. The expository framework (and the implicit framework of the research itself) is based upon that of Rosenbaum (1967). The aim of the paper is to arrive at a rough picture of the kinds of structures and syntactic…

  7. Targeting C-type lectin-like molecule-1 for antibody-mediated immunotherapy in acute myeloid leukemia

    PubMed Central

    Zhao, Xiaoxian; Singh, Shweta; Pardoux, Cecile; Zhao, Jingsong; Hsi, Eric D.; Abo, Arie; Korver, Wouter

    2010-01-01

    Background C-type lectin-like molecule-1 is a transmembrane receptor expressed on myeloid cells, acute myeloid leukemia blasts and leukemic stem cells. To validate the potential of this receptor as a therapeutic target in acute myeloid leukemia, we generated a series of monoclonal antibodies against the extracellular domain of C-type lectin-like molecule-1 and used them to extend the expression profile analysis of acute myeloid leukemia cells and to select cytotoxic monoclonal antibodies against acute myeloid leukemia cells in preclinical models. Design and Methods C-type lectin-like molecule-1 expression was analyzed in acute myeloid leukemia cell lines, and in myeloid derived cells from patients with acute myeloid leukemia and healthy donors. Anti-C-type lectin-like molecule-1 antibody-mediated in vitro cytotoxic activity against acute myeloid leukemia blasts/cell lines and in vivo anti-cancer activity in a mouse xenograft model were assessed. Internalization of C-type lectin-like molecule-1 monoclonal antibodies upon receptor ligation was also investigated. Results C-type lectin-like molecule-1 was expressed in 86.5% (45/52) of cases of acute myeloid leukemia, in 54.5% (12/22) of acute myeloid leukemia CD34+/CD38? stem cells, but not in acute lymphoblastic leukemia blasts (n=5). Selected anti-C-type lectin-like molecule-1 monoclonal antibodies mediated dose-dependent complement-dependent cytotoxicity and antibody-dependent cellular cytotoxicity specifically against acute myeloid leukemia-derived cell lines. Exogenous expression of the transmembrane receptor in HEK293 cells rendered the cells susceptible to antibody-mediated killing by monoclonal antibodies to the receptor. Furthermore, these monoclonal antibodies demonstrated strong complement-dependent cytotoxicity against freshly isolated acute myeloid leukemia blasts (15/16 cases; 94%). The monoclonal antibodies were efficiently internalized upon binding to C-type lectin-like molecule-1 in HL-60 cells. Moreover, a lead chimeric C-type lectin-like molecule-1 monoclonal antibody reduced the tumor size in xenograft mice implanted with HL-60 cells. Conclusions Our results demonstrate that targeting C-type lectin-like molecule-1 with specific cytotoxic monoclonal antibodies is an attractive approach which could lead to novel therapies for acute myeloid leukemia. PMID:19648166

  8. Novel roles for complement receptors in T cell regulation and beyond.

    PubMed

    Kemper, Claudia; Köhl, Jörg

    2013-12-15

    Complement receptors are expressed on cells of the innate and the adaptive immune system. They play important roles in pathogen and danger sensing as they translate the information gathered by complement fluid phase sensors into cellular responses. Further, they control complement activation on viable and apoptotic host cells, clearance of immune complexes and mediate opsonophagocytosis. More recently, evidence has accumulated that complement receptors form a complex network with other innate receptors systems such as the Toll-like receptors, the Notch signaling system, IgG Fc receptors and C-type lectin receptors contributing to the benefit and burden of innate and adaptive immune responses in autoimmune and allergic diseases as well as in cancer and transplantation. Here, we will discuss recent developments and emerging concepts of complement receptor activation and regulation with a particular focus on the differentiation, maintenance and contraction of effector and regulatory T cells. PMID:23796748

  9. Recent insights into structures and functions of C-type lectins in the immune system

    PubMed Central

    Drickamer, Kurt; Taylor, Maureen E

    2015-01-01

    The majority of the C-type lectin-like domains in the human genome likely to bind sugars have been investigated structurally, although novel mechanisms of sugar binding are still being discovered. In the immune system, adhesion and endocytic receptors that bind endogenous mammalian glycans are often conserved, while pathogen-binding C-type lectins on cells of the innate immune system are more divergent. Lack of orthology between some human and mouse receptors, as well as overlapping specificities of many receptors and formation of receptor hetero-oligomers, can make it difficult to define the roles of individual receptors. There is good evidence that C-type lectins initiate signalling pathways in several different ways, but this function remains the least well understood from a mechanistic perspective. PMID:26163333

  10. Endogenous lectins shape the function of dendritic cells and tailor adaptive immunity: mechanisms and biomedical applications.

    PubMed

    Mascanfroni, Ivn D; Cerliani, Juan P; Dergan-Dylon, Sebastin; Croci, Diego O; Ilarregui, Juan M; Rabinovich, Gabriel A

    2011-07-01

    In spite of their central role in orchestrating immunity, dendritic cells (DCs) can also limit harmful reactions and promote immune tolerance by inducing T cell anergy or favoring the differentiation of T regulatory (T(reg)) cells. Several factors may influence the 'decision' of DCs to become immunogenic or tolerogenic including the nature of antigenic challenge, the engagement of selective pathogen recognition receptors (PRRs) and the balance of cytokines and growth factors. In addition, mounting evidence indicates a key role of endogenous lectins including C-type lectins, siglecs and galectins in shaping DC immunogenicity and tailoring adaptive immune responses, through recognition of specific 'glycan signatures' on invading pathogens or host cells. While galectins are in general secreted proteins that act in a paracrine or autocrine manner, all known siglecs and most C-type lectins are membrane-bound receptors that convey glycan-containing information into DC differentiation or maturation programs. Yet, some of the signaling pathways triggered by endogenous lectins converge in similar functional outcomes regardless of divergences in their structure, homology or glycan-binding specificity. To gain a more complete understanding on the role of protein-glycan interactions in DC biology, here we will integrate scattered information on these structurally-divergent but functionally-related lectins and their potential biomedical applications. PMID:21296197

  11. Effect of chum salmon egg lectin on tight junctions in Caco-2 cell monolayers.

    PubMed

    Nemoto, Ryo; Yamamoto, Shintaro; Ogawa, Tomohisa; Naude, Ryno; Muramoto, Koji

    2015-01-01

    The effect of a chum salmon egg lectin (CSL3) on tight junction (TJ) of Caco-2 cell monolayers was investigated. The lectin opened TJ as indicated by the decrease of the transepithelial electrical resistance (TER) value and the increase of the permeation of lucifer yellow, which is transported via the TJ-mediated paracellular pathway. The effects of CSL3 were inhibited by the addition of 10 mM L-rhamnose or D-galactose which were specific sugars for CSL3. The lectin increased the intracellular Ca2+ of Caco-2 cell monolayers, that could be inhibited by the addition of L-rhamnose. The fluorescence immunostaining of ?-actin in Caco-2 cell monolayers revealed that the cytoskeleton was changed by the CSL3 treatment, suggesting that CSL3 depolymerized ?-actin to cause reversible TJ structural and functional disruption. Although Japanese jack bean lectin and wheat germ lectin showed similar effects in the decrease of the TER values and the increase of the intracellular Ca2+, they could not be inhibited by the same concentrations of simple sugars, such as D-glucose and N-acetyl-D-glucosamine. PMID:25951005

  12. Agglutination of Helicobacter pylori coccoids by lectins

    PubMed Central

    Khin, Mar Mar; Hua, Jie Song; Ng, Han Cong; Wadström, Torkel; Ho, Bow

    2000-01-01

    AIM: To study the agglutination pattern of Helicobacter pylori coccoid and spiral forms. METHODS: Assays of agglutination and agglutination inhibition were applied using fifteen commercial lectins. RESULTS: Strong agglutination was observed with mannose-specific Concanavalin A (Con A), fucose-specific Tetragonolobus purpureas (Lotus A) and N-acetyl glucosamine-specific Triticum vulgaris (WGA) lectins. Mannose and fucose specific lectins were reactive with all strains of H. pylori coccoids as compared to the spirals. Specific carbohydrates, glycoproteins and mucin were shown to inhibit H. pylori lectin-agglutination reactions. Pre-treatment of the bacterial cells with formalin and sulphuric acid did not alter the agglutination patterns with lectins. However, sodium periodate treatment of bacterial cells were shown to inhibit agglutination reaction with Con A, Lotus A and WGA lectins. On the contrary, enzymatic treatment of coccoids and spirals did not show marked inhibition of H. pylori lectin agglutination. Interes tingly, heating of H. pylori cells at 60 °C for 1 h was shown to augment the agglutination with all of the lectins tested. CONCLUSION: The considerable differences in lectin agglutination patterns seen among the two differentiated forms of H. pylori might be attributable to the structural changes during the events of morphological transformation, resulting in exposing or masking some of the sugar residues on the cell surface. Possibility of various sugar residues on the cell wall of the coccoids may allow them to bind to different carbohydrate receptors on gastric mucus and epithelial cells. The coccoids with adherence characteristics like the spirals could aid in the pathogenic process of Helicobacter infection. This may probably lead to different clinical outcome of H. pylori associated gastroduodenal disease. PMID:11819557

  13. The X-ray Crystal Structure of Mannose-binding Lectin-associated Serine Proteinase-3 Reveals the Structural Basis for Enzyme Inactivity Associated with the Carnevale, Mingarelli, Malpuech, and Michels (3MC) Syndrome*

    PubMed Central

    Yongqing, Tang; Wilmann, Pascal G.; Reeve, Shane B.; Coetzer, Theresa H.; Smith, A. Ian; Whisstock, James C.; Pike, Robert N.; Wijeyewickrema, Lakshmi C.

    2013-01-01

    The mannose-binding lectin associated-protease-3 (MASP-3) is a member of the lectin pathway of the complement system, a key component of human innate and active immunity. Mutations in MASP-3 have recently been found to be associated with Carnevale, Mingarelli, Malpuech, and Michels (3MC) syndrome, a severe developmental disorder manifested by cleft palate, intellectual disability, and skeletal abnormalities. However, the molecular basis for MASP-3 function remains to be understood. Here we characterize the substrate specificity of MASP-3 by screening against a combinatorial peptide substrate library. Through this approach, we successfully identified a peptide substrate that was 20-fold more efficiently cleaved than any other identified to date. Furthermore, we demonstrated that mutant forms of the enzyme associated with 3MC syndrome were completely inactive against this substrate. To address the structural basis for this defect, we determined the 2.6-? structure of the zymogen form of the G666E mutant of MASP-3. These data reveal that the mutation disrupts the active site and perturbs the position of the catalytic serine residue. Together, these insights into the function of MASP-3 reveal how a mutation in this enzyme causes it to be inactive and thus contribute to the 3MC syndrome. PMID:23792966

  14. Neutrophil recruitment in skin window chambers--activation by complement.

    PubMed

    Elmgreen, J

    1985-06-01

    Complement was studied in skin window chambers, a human model of neutrophil recruitment in acute aseptic inflammation. Autologous plasma activated by the alternative pathway served as attractant; control chambers were filled with a balanced salt solution or with non-activated plasma samples. Neutrophil accumulation during a 24-hour period was consistently enhanced by activated complement in all of 15 healthy volunteers. Control chambers showed negligible cell counts. Reference assays revealed 1) consumption of the centrally placed complement component, C3, 2) generation of chemotactic activity as assessed in Boyden chambers by the standard complement activation procedure. Simultaneously obtained responses to activated complement in skin window chambers and in the Boyden assay of chemotaxis showed a highly significant, positive correlation. Our results demonstrate that the biological capacity of complement includes stimulation of neutrophil migration during simulated in vivo conditions and thus extends previous observations in animals. PMID:4036613

  15. Mannose-binding Lectin (MBL) as a susceptible host factor influencing Indian Visceral Leishmaniasis.

    PubMed

    Mishra, Anshuman; Antony, Justin S; Gai, Prabhanjan; Sundaravadivel, Pandarisamy; Van, Tong Hoang; Jha, Aditya Nath; Singh, Lalji; Velavan, Thirumalaisamy P; Thangaraj, Kumarasamy

    2015-12-01

    Visceral Leishmaniasis (VL), caused by Leishmania donovani is endemic in the Indian sub-continent. Mannose-binding Lectin (MBL) is a complement lectin protein that binds to the surface of Leishmania promastigotes and results in activation of the complement lectin cascade. We utilized samples of 218 VL patients and 215 healthy controls from an Indian population. MBL2 functional variants were genotyped and the circulating MBL serum levels were measured. MBL serum levels were elevated in patients compared to the healthy controls (adjusted P=0.007). The MBL2 promoter variants -78C/T and +4P/Q were significantly associated with relative protection to VL (-78C/T, OR=0.7, 95% CI=0.5-0.96, adjusted P=0.026 and +4P/Q, OR=0.66, 95% CI=0.48-0.9, adjusted P=0.012). MBL2*LYQA haplotypes occurred frequently among controls (OR=0.69, 95% CI=0.5-0.97, adjusted P=0.034). MBL recognizes Leishmania and plays a relative role in establishing L. donovani infection and subsequent disease progression. In conclusion, MBL2 functional variants were associated with VL. PMID:26297290

  16. Activated Complement Factors as Disease Markers for Sepsis

    PubMed Central

    Charchaflieh, Jean; Rushbrook, Julie; Worah, Samrat; Zhang, Ming

    2015-01-01

    Sepsis is a leading cause of death in the United States and worldwide. Early recognition and effective management are essential for improved outcome. However, early recognition is impeded by lack of clinically utilized biomarkers. Complement factors play important roles in the mechanisms leading to sepsis and can potentially serve as early markers of sepsis and of sepsis severity and outcome. This review provides a synopsis of recent animal and clinical studies of the role of complement factors in sepsis development, together with their potential as disease markers. In addition, new results from our laboratory are presented regarding the involvement of the complement factor, mannose-binding lectin, in septic shock patients. Future clinical studies are needed to obtain the complete profiles of complement factors/their activated products during the course of sepsis development. We anticipate that the results of these studies will lead to a multipanel set of sepsis biomarkers which, along with currently used laboratory tests, will facilitate earlier diagnosis, timely treatment, and improved outcome. PMID:26420913

  17. Mannose-binding dietary lectins induce adipogenic differentiation of the marrow-derived mesenchymal cells via an active insulin-like signaling mechanism.

    PubMed

    Bajaj, Manmohan; Hinge, Ashwini; Limaye, Lalita S; Gupta, Rajesh Kumar; Surolia, Avadhesha; Kale, Vaijayanti P

    2011-04-01

    We have recently demonstrated that the mannose-binding lectins, namely banana lectin (BL) and garlic lectin (GL), interacted with the insulin receptors on M210B4 cells--an established mesenchymal cell line of murine marrow origin--and initiate mitogen-activated protein kinase kinase (MEK)-dependent extracellular signal-regulated kinase (ERK) signaling in them. In this study, we show that this lectin-mediated active ERK signaling culminates into an adipogenic differentiation of these cells. Gene expression studies indicate that the effect takes place at the transcriptional level. Experiments carried out with pharmacological inhibitors show that MEK-dependent ERK and phosphatidylinositol 3-kinase-dependent AKT pathways are positive regulators of the lectin- and insulin-mediated adipogenic differentiation, while stress-activated kinase/c-jun N-terminal kinase pathway acts as a negative one. Since both lectins could efficiently substitute for insulin in the standard adipogenic induction medium, they may perhaps serve as molecular tools to study the mechanistic aspects of the adipogenic process that are independent of cell proliferation. Our study clearly demonstrates the ability of BL and GL to activate insulin-like signaling in the mesenchymal cells in vitro leading to their adipocytic differentiation. The dietary origin of these lectins underscores an urgent need to examine their in vivo effects on tissue homeostasis. PMID:21106560

  18. Disabling complement regulatory activities of vaccinia virus complement control protein reduces vaccinia virus pathogenicity

    PubMed Central

    Bernet, John; Ahmad, Muzammil; Mullick, Jayati; Panse, Yogesh; Singh, Akhilesh K.; Parab, Pradeep B.; Sahu, Arvind

    2011-01-01

    Poxviruses encode a repertoire of immunomodulatory proteins to thwart the host immune system. One among this array is a homolog of the host complement regulatory proteins that is conserved in various poxviruses including vaccinia (VACV) and variola. The vaccinia virus complement control protein (VCP), which inhibits complement by decaying the classical pathway C3-convertase (decay-accelerating activity), and by supporting inactivation of C3b and C4b by serine protease factor I (cofactor activity), was shown to play a role in viral pathogenesis. However, the role its individual complement regulatory activities impart in pathogenesis, have not yet been elucidated. Here, we have generated monoclonal antibodies (mAbs) that block the VCP functions and utilized them to evaluate the relative contribution of complement regulatory activities of VCP in viral pathogenesis by employing a rabbit intradermal model for VACV infection. Targeting VCP by mAbs that inhibited the decay-accelerating activity as well as cofactor activity of VCP or primarily the cofactor activity of VCP, by injecting them at the site of infection, significantly reduced VACV lesion size. This reduction however was not pronounced when VCP was targeted by a mAb that inhibited only the decay-accelerating activity. Further, the reduction in lesion size by mAbs was reversed when host complement was depleted by injecting cobra venom factor. Thus, our results suggest that targeting VCP by antibodies reduces VACV pathogenicity and that principally the cofactor activity of VCP appears to contribute to the virulence. PMID:21803094

  19. Trichinella spiralis Paramyosin Binds Human Complement C1q and Inhibits Classical Complement Activation

    PubMed Central

    Sun, Ran; Zhao, Xi; Wang, Zixia; Yang, Jing; Zhao, Limei; Zhan, Bin; Zhu, Xinping

    2015-01-01

    Background Trichinella spiralis expresses paramyosin (Ts-Pmy) as a defense mechanism. Ts-Pmy is a functional protein with binding activity to human complement C8 and C9 and thus plays a role in evading the attack of the host’s immune system. In the present study, the binding activity of Ts-Pmy to human complement C1q and its ability to inhibit classical complement activation were investigated. Methods and Findings The binding of recombinant and natural Ts-Pmy to human C1q were determined by ELISA, Far Western blotting and immunoprecipitation, respectively. Binding of recombinant Ts-Pmy (rTs-Pmy) to C1q inhibited C1q binding to IgM and consequently inhibited C3 deposition. The lysis of antibody-sensitized erythrocytes (EAs) elicited by the classical complement pathway was also inhibited in the presence of rTs-Pmy. In addition to inhibiting classical complement activation, rTs-Pmy also suppressed C1q binding to THP-1-derived macrophages, thereby reducing C1q-induced macrophages migration. Conclusion Our results suggest that T. spiralis paramyosin plays an important role in immune evasion by interfering with complement activation through binding to C1q in addition to C8 and C9. PMID:26720603

  20. Incorporation of Host Complement Regulatory Proteins into Newcastle Disease Virus Enhances Complement Evasion

    PubMed Central

    Biswas, Moanaro; Johnson, John B.; Kumar, Sandeep R. P.; Parks, Griffith D.

    2012-01-01

    Newcastle disease virus (NDV), an avian paramyxovirus, is inherently tumor selective and is currently being considered as a clinical oncolytic virus and vaccine vector. In this study, we analyzed the effect of complement on the neutralization of NDV purified from embryonated chicken eggs, a common source for virus production. Fresh normal human serum (NHS) neutralized NDV by multiple pathways of complement activation, independent of neutralizing antibodies. Neutralization was associated with C3 deposition and the activation of C2, C3, C4, and C5 components. Interestingly, NDV grown in mammalian cell lines was resistant to complement neutralization by NHS. To confirm whether the incorporation of regulators of complement activity (RCA) into the viral envelope afforded complement resistance, we grew NDV in CHO cells stably transfected with CD46 or HeLa cells, which strongly express CD46 and CD55. NDV grown in RCA-expressing cells was resistant to complement by incorporating CD46 and CD55 on virions. Mammalian CD46 and CD55 molecules on virions exhibited homologous restriction, since chicken sera devoid of neutralizing antibodies to NDV were able to effectively neutralize these virions. The incorporation of chicken RCA into NDV produced in embryonated eggs similarly provided species specificity toward chicken sera. PMID:22973037

  1. Identification of mycobacterial lectins from genomic data.

    PubMed

    Abhinav, K V; Sharma, Alok; Vijayan, M

    2013-04-01

    Sixty-four sequences containing lectin domains with homologs of known three-dimensional structure were identified through a search of mycobacterial genomes. They appear to belong to the β-prism II, the C-type, the Microcystis virdis (MV), and the β-trefoil lectin folds. The first three always occur in conjunction with the LysM, the PI-PLC, and the β-grasp domains, respectively while mycobacterial β-trefoil lectins are unaccompanied by any other domain. Thirty heparin binding hemagglutinins (HBHA), already annotated, have also been included in the study although they have no homologs of known three-dimensional structure. The biological role of HBHA has been well characterized. A comparison between the sequences of the lectin from pathogenic and nonpathogenic mycobacteria provides insights into the carbohydrate binding region of the molecule, but the structure of the molecule is yet to be determined. A reasonable picture of the structural features of other mycobacterial proteins containing one or the other of the four lectin domains can be gleaned through the examination of homologs proteins, although the structure of none of them is available. Their biological role is also yet to be elucidated. The work presented here is among the first steps towards exploring the almost unexplored area of the structural biology of mycobacterial lectins. PMID:23180653

  2. Complementing Gender Analysis Methods.

    PubMed

    Kumar, Anant

    2016-01-01

    The existing gender analysis frameworks start with a premise that men and women are equal and should be treated equally. These frameworks give emphasis on equal distribution of resources between men and women and believe that this will bring equality which is not always true. Despite equal distribution of resources, women tend to suffer and experience discrimination in many areas of their lives such as the power to control resources within social relationships, and the need for emotional security and reproductive rights within interpersonal relationships. These frameworks believe that patriarchy as an institution plays an important role in women's oppression, exploitation, and it is a barrier in their empowerment and rights. Thus, some think that by ensuring equal distribution of resources and empowering women economically, institutions like patriarchy can be challenged. These frameworks are based on proposed equality principle which puts men and women in competing roles. Thus, the real equality will never be achieved. Contrary to the existing gender analysis frameworks, the Complementing Gender Analysis framework proposed by the author provides a new approach toward gender analysis which not only recognizes the role of economic empowerment and equal distribution of resources but suggests to incorporate the concept and role of social capital, equity, and doing gender in gender analysis which is based on perceived equity principle, putting men and women in complementing roles that may lead to equality. In this article the author reviews the mainstream gender theories in development from the viewpoint of the complementary roles of gender. This alternative view is argued based on existing literature and an anecdote of observations made by the author. While criticizing the equality theory, the author offers equity theory in resolving the gender conflict by using the concept of social and psychological capital. PMID:25941756

  3. Complement activation promotes muscle inflammation during modified muscle use

    NASA Technical Reports Server (NTRS)

    Frenette, J.; Cai, B.; Tidball, J. G.

    2000-01-01

    Modified muscle use can result in muscle inflammation that is triggered by unidentified events. In the present investigation, we tested whether the activation of the complement system is a component of muscle inflammation that results from changes in muscle loading. Modified rat hindlimb muscle loading was achieved by removing weight-bearing from the hindlimbs for 10 days followed by reloading through normal ambulation. Experimental animals were injected with the recombinant, soluble complement receptor sCR1 to inhibit complement activation. Assays for complement C4 or factor B in sera showed that sCR1 produced large reductions in the capacity for activation of the complement system through both the classical and alternative pathways. Analysis of complement C4 concentration in serum in untreated animals showed that the classical pathway was activated during the first 2 hours of reloading. Analysis of factor B concentration in untreated animals showed activation of the alternative pathway at 6 hours of reloading. Administration of sCR1 significantly attenuated the invasion of neutrophils (-49%) and ED1(+) macrophages (-52%) that occurred in nontreated animals after 6 hours of reloading. The presence of sCR1 also reduced significantly the degree of edema by 22% as compared to untreated animals. Together, these data show that increased muscle loading activated the complement system which then briefly contributes to the early recruitment of inflammatory cells during modified muscle loading.

  4. Complement Activation and Inhibition in Retinal Diseases.

    PubMed

    Kleinman, Mark E; Ambati, Jayakrishna

    2016-01-01

    Within the past several decades, a brigade of dedicated researchers from around the world has provided essential insights into the critical niche of immune-mediated inflammation in the pathogenesis of age-related macular degeneration (AMD). Yet, the question has lingered as to whether disease-initiating events are more or less dependent on isolated immune-related responses, unimpeded inflammation, endogenous pathways of age-related cell senescence and oxidative stress, or any of the other numerous molecular derangements that have been identified in the natural history of AMD. There is now an abundant cache of data signifying immune system activation as an impetus in the pathogenesis of this devastating condition. Furthermore, recent rigorous investigations have revealed multiple inciting factors, including several important complement-activating components, thus creating a new array of disease-modulating targets for the research and development of molecular therapeutic interventions. While the precise in vivo effects of complement activation and inhibition in the progression and treatment of AMD remain to be determined, ongoing clinical trials of the first generation of complement-targeted therapeutics are hoped to yield critical data on the contribution of this pathway to the disease process. PMID:26501209

  5. Complement and HIV-I infection/ HIV-associated neurocognitive disorders

    PubMed Central

    Liu, Fengming; Dai, Shen; Gordon, Jennifer; Qin, Xuebin

    2014-01-01

    The various neurological complications associated with HIV-1 infection, specifically HIV-associated neurocognitive disorders (HAND) persist as a major public health burden worldwide. Despite the widespread use of anti-retroviral therapy, the prevalence of HAND is significantly high. HAND results from the direct effects of an HIV-1 infection as well as secondary effects of HIV-1-induced immune reaction and inflammatory response. Complement, a critical mediator of innate and acquired immunity, plays important roles in defeating many viral infections by the formation of a lytic pore or indirectly by opsonization and recruitment of phagocytes. While the role of complement in the pathogenesis of HIV-1 infection and HAND has been previously recognized for over fifteen years, it has been largely underestimated thus far. Complement can be activated through HIV-1 envelope proteins, mannose binding lectins (MBL) and anti-HIV-1 antibodies. Complement not only fights against HIV-1 infection but also enhances HIV-1 infection. Also, HIV-1 can hijack complement regulators such as CD59 and CD55 and can utilize these regulators and factor H to escape from complement attack. Normally, complement levels in brain are much lower than plasma levels and there is no or little complement deposition in brain cells. Interestingly, local production and deposition of complement are dramatically increased in HIV-1-infected brain, indicating that complement may contribute to the pathogenesis of HAND. Here, we review the current understanding of the role of complement in HIV-1 infection and HAND as well as potential therapeutic approaches targeting to the complement system for the treatment and eradications of HIV-1 infection. PMID:24639397

  6. Mannan-binding lectin may facilitate the clearance of circulating immune complexes--implications from a study on C2-deficient individuals.

    PubMed

    Saevarsdottir, S; Steinsson, K; Ludviksson, B R; Grondal, G; Valdimarsson, H

    2007-05-01

    Deficiency of both mannan-binding lectin (MBL) and complement components C4 and C2 has been associated with increased risk of systemic lupus erythematosus (SLE). MBL can activate the complement system either through C4 and C2 or directly through C3. Circulating immune complexes (CICs) are believed to play a pathogenic role in SLE and MBL has been shown to bind certain forms of immunoglobulins, including IgM, IgG and IgA. Thus, MBL might promote CIC clearance. In order to evaluate this, six individuals with non-functional classical pathway due to the rare homozygous C2 deficiency were chosen, as the classical pathway is known to have a fundamental role in CIC clearance. Four of the six C2-deficient individuals had SLE, two of whom also had MBL deficiency. MBL serum levels and genotypes were compared with the serum levels of CICs, as measured by their content of kappa, lambda, IgM, IgA, IgG and C3 opsonization. The C2-deficient individuals had higher serum levels of CICs than 16 healthy controls (P < 0.0001). Furthermore, an inverse association was observed between MBL and CIC levels in the C2-deficient individuals, which was strongest for IgM-CICs (r = - 0.84, P = 0.037). Moreover, C3 opsonization of the CICs correlated positively with MBL levels in the C2-deficient individuals (r = 0.89, P = 0.017). In conclusion, individuals with C2 deficiency have increased levels of CICs and MBL may facilitate their clearance. Defective CIC clearance might partly explain the increased risk of SLE associated with low MBL. PMID:17335556

  7. Evasion of Complement-Mediated Lysis and Complement C3 Deposition Are Regulated by Francisella tularensis Lipopolysaccharide O Antigen1

    PubMed Central

    Clay, Corey D.; Soni, Shilpa; Gunn, John S.; Schlesinger, Larry S.

    2009-01-01

    The bacterium Francisella tularensis (Ft) is a potential weapon of bioterrorism when aerosolized. Macrophage infection is necessary for disease progression and efficient phagocytosis by human macrophages requires serum opsonization by complement. Microbial complement activation leads to surface deposition of a highly regulated protein complex resulting in opsonization or membrane lysis. The nature of complement component C3 deposition, i.e., C3b (opsonization and lysis) or C3bi (opsonization only) fragment deposition, is central to the outcome of activation. In this study, we examine the mechanisms of Ft resistance to complement-mediated lysis, C3 component deposition on the Ft surface, and complement activation. Upon incubation in fresh nonimmune human serum, Schu S4 (Ft subsp. tularensis), Fn (Ft subsp. novicida), and LVS (Ft subsp. holarctica live vaccine strain) were resistant to complement-mediated lysis, but LVSG and LVSR (LVS strains altered in surface carbohydrate structures) were susceptible. C3 deposition, however, occurred on all strains. Complement-susceptible strains had markedly increased C3 fragment deposition, including the persistent presence of C3b compared with C3bi, which indicates that C3b inactivation results in survival of complement-resistant strains. C1q, an essential component of the classical activation pathway, was necessary for lysis of complement-susceptible strains and optimal C3 deposition on all strains. Finally, use of Francisella LPS mutants confirmed O Ag as a major regulator of complement resistance. These data provide evidence that pathogenic Francisella activate complement, but are resistant to complement-mediated lysis in part due to limited C3 deposition, rapid conversion of surface-bound C3b to C3bi, and the presence of LPS O Ag. PMID:18832715

  8. A family with complement factor D deficiency

    PubMed Central

    Biesma, Douwe H.; Hannema, Andr J.; van Velzen-Blad, Heleen; Mulder, Leontine; van Zwieten, Rob; Kluijt, Irma; Roos, Dirk

    2001-01-01

    A complement factor D deficiency was found in a young woman who had experienced a serious Neisseria meningitidis infection, in a deceased family member with a history of meningitis, and in three relatives without a history of serious infections. The patient and these three relatives showed a normal activity of the classical complement pathway, but a very low activity of the alternative complement pathway and a very low capacity to opsonize Escherichia coli and N. meningitidis (isolated from the patient) for phagocytosis by normal human neutrophils. The alternative pathway-dependent hemolytic activity and the opsonizing capacity of these sera were restored by addition of purified factor D. The family had a high degree of consanguinity, and several other family members exhibited decreased levels of factor D. The gene encoding factor D was found to contain a point mutation that changed the TCG codon for serine 42 into a TAG stop codon. This mutation was found in both alleles of the five completely factor Ddeficient family members and in one allele of 21 other members of the same family who had decreased or low-normal factor D levels in their serum. The gene sequence of the signal peptide of human factor D was also identified. Our report is the first, to our knowledge, to document a Factor D gene mutation. The mode of inheritance of factor D deficiency is autosomal recessive, in accordance with the localization of the Factor D gene on chromosome 19. Increased susceptibility for infections in individuals with a partial factor D deficiency is unlikely. PMID:11457876

  9. Anti-Gal binds to pili of Neisseria meningitidis: the immunoglobulin A isotype blocks complement-mediated killing.

    PubMed Central

    Hamadeh, R M; Estabrook, M M; Zhou, P; Jarvis, G A; Griffiss, J M

    1995-01-01

    alpha 1,3-Galactosyl antibodies (anti-Gal) are ubiquitous natural human serum and secretory polyclonal antibodies that bind to terminal galactose-alpha 1,3-galactose (alpha-galactosyl) residues. Serum immunoglobulin G (IgG) anti-Gal can block alternative complement pathway-mediated lysis of representative gram-negative enteric bacteria that bind it to lipopolysaccharide alpha-galactosyl structures, thereby promoting survival of such bacteria in the nonimmune host. We wanted to know whether anti-Gal also could bind to the lipooligosaccharides (LOS) of Neisseria meningitidis. To our surprise, we found that serum and secretory anti-Gal bound to pili but not to LOS of certain strains. This suggested the presence of an immunogenic pilus carbohydrate epitope. Mild periodate oxidation of sodium dodecyl sulfate-polyacrylamide gel electrophoresis-separated outer membrane preparations from strains that bound anti-Gal followed by labeling of the neoaldehyde groups resulted in the labeling of bands that corresponded to pilin and LOS, confirming that pilin contains carbohydrate structures. A Bandeiraea simplicifolia lectin that also binds terminal alpha 1,3-galactosyl residues also bound to pilin. Serum IgG, IgA, and IgM anti-Gal as well as colostral secretory IgA anti-Gal bound to pilin, as judged by immunoblotting, and to the pili of intact piliated organisms, as judged by immunoelectron microscopy. Total serum anti-Gal (IgG, IgA, and IgM) and purified serum IgA1 anti-Gal, but not its purified IgG isotype, blocked complement-mediated lysis of a piliated meningococcal strain that bound anti-Gal to its pili. Colostral anti-Gal secretory IgA blocked killing of the same strain. Thus, anti-Gal IgA may promote disease when it binds to the pili of N. meningitidis strains. PMID:7591153

  10. Purification and characterization of Dolichos lablab lectin.

    PubMed

    Mo, H; Meah, Y; Moore, J G; Goldstein, I J

    1999-02-01

    The mannose/glucose-binding Dolichos lablab lectin (designated DLL) has been purified from seeds of Dolichos lablab (hyacinth bean) to electrophoretic homogeneity by affinity chromatography on an ovalbumin-Sepharose 4B column. The purified lectin gave a single symmetric protein peak with an apparent molecular mass of 67 kDa on gel filtration chromatography, and five bands ranging from 10 kDa to 22 kDa upon SDS-PAGE. N-Terminal sequence analysis of these bands revealed subunit heterogeneity due to posttranslational proteolytic truncation at different sites mostly at the carboxyl terminus. The carbohydrate binding properties of the purified lectin were investigated by three different approaches: hemagglutination inhibition assay, quantitative precipitation inhibition assay, and ELISA. On the basis of these studies, it is concluded that the Dolichos lablab lectin has neither an extended carbohydrate combining site, nor a hydrophobic binding site adjacent to it. The carbohydrate combining site of DLL appears to most effectively accommodate a nonreducing terminal alpha-d-mannosyl unit, and to be complementary to the C-3, C-4, and C-6 equatorial hydroxyl groups of alpha-d-mannopyranosyl and alpha-d-glucopyranosyl residues. DLL strongly precipitates murine IgM but not IgG, and the recent finding that this lectin interacts specifically with NIH 3T3 fibroblasts transfected with the Flt3 tyrosine kinase receptor and preserves human cord blood stem cells and progenitors in a quiescent state for prolonged periods in culture, make this lectin a valuable tool in biomedical research. PMID:9949194

  11. Leishmania donovani metacyclic promastigotes: transformation in vitro, lectin agglutination, complement resistance, and infectivity.

    PubMed

    Howard, M K; Sayers, G; Miles, M A

    1987-10-01

    Freshly transformed Leishmania donovani amastigotes from hamster spleen were used to establish axenic cultures at high density in a modified Grace's medium, which was only partly replenished when cultures were fed. Small, free-swimming, highly active stationary phase promastigotes with a short cell body and long flagellum were induced in this medium. The freshly transformed stationary phase promastigotes so induced were less able to bind peanut agglutinin, had more than 40-fold increased resistance to killing by normal human serum, and 15-fold increased infectivity both in vivo and in vitro when compared to freshly transformed logarithmic phase or long term culture promastigotes. These short form promastigotes may correspond to the metacyclic promastigote forms in the sand fly vector. PMID:3653336

  12. Role of Lectins in Plant-Microorganism Interactions

    PubMed Central

    Pueppke, Steven G.; Bauer, Wolfgang D.; Keegstra, Kenneth; Ferguson, Ardene L.

    1978-01-01

    Three different assay procedures have been used to quantitate the levels of soybean (Glycine max [L.] Merr.) lectin in various tissues of soybean plants. The assays used were a standard hemagglutination assay, a radioimmunoassay, and an isotope dilution assay. Most of the lectin in seeds was found in the cotyledons, but lectin was also detected in the embryo axis and the seed coat. Soybean lectin was present in all of the tissues of young seedlings, but decreased as the plants matured and was not detectable in plants older than 2 to 3 weeks. Soybean lectin isolated from seeds of several soybean varieties were identical when compared by several methods. PMID:16660384

  13. Mushroom Lectins: Specificity, Structure and Bioactivity Relevant to Human Disease

    PubMed Central

    Hassan, Mohamed Ali Abol; Rouf, Razina; Tiralongo, Evelin; May, Tom W.; Tiralongo, Joe

    2015-01-01

    Lectins are non-immunoglobulin proteins that bind diverse sugar structures with a high degree of selectivity. Lectins play crucial role in various biological processes such as cellular signaling, scavenging of glycoproteins from the circulatory system, cell–cell interactions in the immune system, differentiation and protein targeting to cellular compartments, as well as in host defence mechanisms, inflammation, and cancer. Among all the sources of lectins, plants have been most extensively studied. However, more recently fungal lectins have attracted considerable attention due to their antitumor, antiproliferative and immunomodulatory activities. Given that only 10% of mushroom species are known and have been taxonomically classified, mushrooms represent an enormous unexplored source of potentially useful and novel lectins. In this review we provide an up-to-date summary on the biochemical, molecular and structural properties of mushroom lectins, as well as their versatile applications specifically focusing on mushroom lectin bioactivity. PMID:25856678

  14. Mushroom lectins: specificity, structure and bioactivity relevant to human disease.

    PubMed

    Hassan, Mohamed Ali Abol; Rouf, Razina; Tiralongo, Evelin; May, Tom W; Tiralongo, Joe

    2015-01-01

    Lectins are non-immunoglobulin proteins that bind diverse sugar structures with a high degree of selectivity. Lectins play crucial role in various biological processes such as cellular signaling, scavenging of glycoproteins from the circulatory system, cell-cell interactions in the immune system, differentiation and protein targeting to cellular compartments, as well as in host defence mechanisms, inflammation, and cancer. Among all the sources of lectins, plants have been most extensively studied. However, more recently fungal lectins have attracted considerable attention due to their antitumor, antiproliferative and immunomodulatory activities. Given that only 10% of mushroom species are known and have been taxonomically classified, mushrooms represent an enormous unexplored source of potentially useful and novel lectins. In this review we provide an up-to-date summary on the biochemical, molecular and structural properties of mushroom lectins, as well as their versatile applications specifically focusing on mushroom lectin bioactivity. PMID:25856678

  15. Identification of Complement Regulatory Domains in Vaccinia Virus Complement Control Protein

    PubMed Central

    Mullick, Jayati; Bernet, John; Panse, Yogesh; Hallihosur, Sharanabasava; Singh, Akhilesh K.; Sahu, Arvind

    2005-01-01

    Vaccinia virus encodes a homolog of the human complement regulators named vaccinia virus complement control protein (VCP). It is composed of four contiguous complement control protein (CCP) domains. Previously, VCP has been shown to bind to C3b and C4b and to inactivate the classical and alternative pathway C3 convertases by accelerating the decay of the classical pathway C3 convertase and (to a limited extent) the alternative pathway C3 convertase, as well as by supporting the factor I-mediated inactivation of C3b and C4b (the subunits of C3 convertases). In this study, we have mapped the CCP domains of VCP important for its cofactor activities, decay-accelerating activities, and binding to the target proteins by utilizing a series of deletion mutants. Our data indicate the following. (i) CCPs 1 to 3 are essential for cofactor activity for C3b and C4b; however, CCP 4 also contributes to the optimal activity. (ii) CCPs 1 to 2 are enough to mediate the classical pathway decay-accelerating activity but show very minimal activity, and all the four CCPs are necessary for its efficient activity. (iii) CCPs 2 to 4 mediate the alternative pathway decay-accelerating activity. (iv) CCPs 1 to 3 are required for binding to C3b and C4b, but the presence of CCP 4 enhances the affinity for both the target proteins. These results together demonstrate that the entire length of the protein is required for VCP's various functional activities and suggests why the four-domain structure of viral CCP is conserved in poxviruses. PMID:16160165

  16. Structural analysis of a Dioclea sclerocarpa lectin: Study on the vasorelaxant properties of Dioclea lectins.

    PubMed

    Barroso-Neto, Ito L; Delatorre, Plinio; Teixeira, Claudener S; Correia, Jorge L A; Cajazeiras, João B; Pereira, Ronniery I; Nascimento, Kyria S; Laranjeira, Eva P P; Pires, Alana F; Assreuy, Ana M S; Rocha, Bruno A M; Cavada, Benildo S

    2016-01-01

    Lectins are proteins that show a variety of biological activities. However, they share in common at least one domain capable of recognizing specific carbohydrates reversibly without changing its structure. The legume lectins family is the most studied family of plant lectins, in particular the Diocleinae subtribe, which possesses high degree of structural similarity, but variable biological activities. This variability lies in small differences that can be analyzed in studies based on structures. In particular, Dioclea sclerocarpa seed lectin (DSL) presents low ability to relax endothelialized rat aorta in comparison with other Dioclea lectins such as Dioclea violacea (DVL), Dioclea virgata (DvirL) and Dioclea rostrata (DRL). The DSL relaxation mechanism relies on nitric oxide production and carbohydrate recognition domain (CRD). This feature can be explained by structural differences, since DSL has a carbohydrate recognition domain design less favorable. In addition, the presence of a glutamate residue at position 205 proved to be a decisive factor for the low relaxant effect of Dioclea lectins. PMID:26499084

  17. Complement activation by neutrophil granulocytes.

    PubMed

    Asberg, A E; Mollnes, T E; Videm, V

    2008-04-01

    Complement plays a vital role in the body's defence systems. Cardiopulmonary bypass induces a detrimental inflammatory reaction in which the complement system is known to participate through direct effects as well as through activation of neutrophils, platelets and endothelial cells. On the other hand, it has been suggested that in the setting of cardiopulmonary bypass, complement may be activated by neutrophils, perhaps due to fragmentation caused by the heart-lung machine. We therefore investigated whether intact or fragmented neutrophils were able to activate the complement system, and whether neutrophil-platelet interaction could influence such complement activation. Lepirudin-anticoagulated plasma was incubated at 37 degrees C with resting or activated intact neutrophils or neutrophils combined with platelets, or increasing amounts of fragmented neutrophils. Complement activation was evaluated by measurement of C1rs-C1 inhibitor complexes, C4bc, C3bBbP, C3bc, C5a and sC5b-9. We found significant activation of complement only by unphysiological doses of fragmented neutrophils or supernatant from fragmented neutrophils, consistent with a limited clinical significance related to neutrophil destruction during cardiopulmonary bypass. Unstimulated neutrophils induced C3bPBb formation but little formation of other activation products, indicating an increased C3 hydrolysis which was kept under control by regulatory mechanisms. Neutrophils and platelets combined increased classical activation and decreased alternative activation, similar to the findings with platelets alone. Our data confirm that in the setting of acute neutrophil fragmentation or activation, complement activation is much more important in the inflammatory network as an event upstream to neutrophil activation than vice versa. PMID:18248527

  18. Displacement phenomena in lectin affinity chromatography.

    PubMed

    Cho, Wonryeon

    2015-10-01

    The work described here examines displacement phenomena that play a role in lectin affinity chromatography and their potential to impact reproducibility. This was achieved using Lycopersicon esculentum lectin (LEL), a lectin widely used in monitoring cancer. Four small identical LEL columns were coupled in series to form a single affinity chromatography system with the last in the series connected to an absorbance detector. The serial affinity column set (SACS) was then loaded with human plasma proteins. At the completion of loading, the column set was disassembled, the four columns were eluted individually, the captured proteins were trypsin digested, the peptides were deglycosylated with PNGase F, and the parent proteins were identified through mass spectral analyses. Significantly different sets of glycoproteins were selected by each column, some proteins appearing to be exclusively bound to the first column while others were bound further along in the series. Clearly, sample displacement chromatography (SDC) occurs. Glycoproteins were bound at different places in the column train, identifying the presence of glycoforms with different affinity on a single glycoprotein. It is not possible to see these phenomena in the single column mode of chromatography. Moreover, low abundance proteins were enriched, which facilitates detection. The great advantage of this method is that it differentiates between glycoproteins on the basis of their binding affinity. Displacement phenomena are concluded to be a significant component of the separation mechanism in heavily loaded lectin affinity chromatography columns. This further suggests that care must be exercised in sample loading of lectin columns to prevent analyte displacement with nonretained proteins. PMID:26348026

  19. Complement involvement in periodontitis: molecular mechanisms and rational therapeutic approaches

    PubMed Central

    Hajishengallis, George; Maekawa, Tomoki; Abe, Toshiharu; Hajishengallis, Evlambia; Lambris, John D.

    2015-01-01

    The complement system is a network of interacting fluid-phase and cell surface-associated molecules that trigger, amplify, and regulate immune and inflammatory signaling pathways. Dysregulation of this finely balanced network can destabilize host-microbe homeostasis and cause inflammatory tissue damage. Evidence from clinical and animal model-based studies suggests that complement is implicated in the pathogenesis of periodontitis, a polymicrobial community-induced chronic inflammatory disease that destroys the tooth-supporting tissues. This review discusses molecular mechanisms of complement involvement in the dysbiotic transformation of the periodontal microbiome and the resulting destructive inflammation, culminating in loss of periodontal bone support. These mechanistic studies have additionally identified potential therapeutic targets. In this regard, interventional studies in preclinical models have provided proof-of-concept for using complement inhibitors for the treatment of human periodontitis. PMID:26306443

  20. Complement - a key system for immune surveillance and homeostasis

    PubMed Central

    Ricklin, Daniel; Hajishengallis, George; Yang, Kun; Lambris, John D.

    2010-01-01

    Nearly a century after the significance of the human complement system was recognized we have come to realize that its versatile functions extend far beyond the elimination of microbes. Indeed, complement acts as a rapid and efficient immune surveillance system that has distinct effects on healthy and altered host cells and foreign intruders. By eliminating cellular debris and infectious microbes, orchestrating immune responses, and sending `danger' signals, complement contributes substantially to homeostasis, but it may also take action against healthy cells if not properly controlled. This review describes our updated view of the function, structure, and dynamics of the complement network, highlights its interconnection with immunity at large and with other endogenous pathways, and illustrates its dual role in homeostasis and disease. PMID:20720586

  1. Genetics Home Reference: Complement component 2 deficiency

    MedlinePLUS

    ... literature OMIM Genetic disorder catalog Conditions > Complement component 2 deficiency On this page: Description Genetic changes Inheritance ... definitions Reviewed June 2014 What is complement component 2 deficiency? Complement component 2 deficiency is a disorder ...

  2. Genetics Home Reference: Complement component 2 deficiency

    MedlinePLUS

    ... other areas of the world is unknown. What genes are related to complement component 2 deficiency? Complement ... deficiency is caused by mutations in the C2 gene. This gene provides instructions for making the complement ...

  3. Genetics Home Reference: Complement component 8 deficiency

    MedlinePLUS

    ... Recent literature OMIM Genetic disorder catalog Conditions > Complement component 8 deficiency On this page: Description Genetic changes ... Glossary definitions Reviewed December 2015 What is complement component 8 deficiency? Complement component 8 deficiency is a ...

  4. Sensing lectin-glycan interactions using lectin super-microarrays and glycans labeled with dye-doped silica nanoparticles

    PubMed Central

    Wang, Xin; Matei, Elena; Deng, Lingquan; Koharudin, Leonardus; Gronenborn, Angela M.; Ramström, Olof; Yan, Mingdi

    2013-01-01

    A new microarray platform, based on lectin super-microarrays and glycans labeled with dye-doped nanoparticles, has been developed to study glycan-lectin interactions. Glycan ligands were conjugated onto fluorescein-doped silica nanoparticles (FSNPs) using a general photocoupling chemistry to afford FSNP-labeled glycan probes. Lectins were printed on epoxy slides in duplicate sets to generate lectin super-microarrays where multiple assays could be carried out simultaneously in each lectin microarray. Thus, the lectin super-microarray was treated with FSNP-labeled glycans to screen for specific binding pairs. Furthermore, a series of ligand competition assays were carried out on a single lectin super-microarray to generate the dose-response curve for each glycan-lectin pair, from which the apparent affinity constants were obtained. Results showed 4~7 orders of magnitude increase in affinity over the free glycans with the corresponding lectins. Thus, the glycan epitope structures having weaker affinity than the parent glycans could be readily identified and analyzed from the lectin super-microarrays. PMID:23584388

  5. Two Chitotriose-Specific Lectins Show Anti-Angiogenesis, Induces Caspase-9-Mediated Apoptosis and Early Arrest of Pancreatic Tumor Cell Cycle

    PubMed Central

    Sarkar, Dhiman; Suresh, C. G.

    2016-01-01

    The antiproliferative activity of two chito- specific agglutinins purified from Benincasa hispida (BhL) and Datura innoxia (DiL9) of different plant family origin was investigated on various cancer cell lines. Both lectins showed chitotriose specificity, by inhibiting lectin hemagglutinating activity. On further studies, it was revealed that these agglutinins caused remarkable concentration-dependent antiproliferative effect on human pancreatic cancerous cells but not on the normal human umbilical vein endothelial cells even at higher doses determined using MTT assay. The GI50 values were approximately 8.4 μg ml-1 (0.247 μM) and 142 μg ml-1(14.8 μM) for BhL and DiL9, respectively, against PANC-1 cells. The growth inhibitory effect of these lectins on pancreatic cancer cells were shown to be a consequence of lectin cell surface binding and triggering G0/G1 arrest, mitochondrial membrane depolarization, sustained increase of the intracellular calcium release and the apoptotic signal is amplified by activation of caspases executing cell death. Interestingly, these lectins also showed anti-angiogenic activity by disrupting the endothelial tubulogenesis. Therefore, we report for the first time two chito-specific lectins specifically binding to tumor glycans; they can be considered to be a class of molecules with antitumor activity against pancreatic cancer cells mediated through caspase dependent mitochondrial apoptotic pathway. PMID:26795117

  6. Depletion of specific cell populations by complement depletion.

    PubMed

    Dittel, Bonnie N

    2010-01-01

    The purification of immune cell populations is often required in order to study their unique functions. In particular, molecular approaches such as real-time PCR and microarray analysis require the isolation of cell populations with high purity. Commonly used purification strategies include fluorescent activated cell sorting (FACS), magnetic bead separation and complement depletion. Of the three strategies, complement depletion offers the advantages of being fast, inexpensive, gentle on the cells and a high cell yield. The complement system is composed of a large number of plasma proteins that when activated initiate a proteolytic cascade culminating in the formation of a membrane-attack complex that forms a pore on a cell surface resulting in cell death(1). The classical pathway is activated by IgM and IgG antibodies and was first described as a mechanism for killing bacteria. With the generation of monoclonal antibodies (mAb), the complement cascade can be used to lyse any cell population in an antigen-specific manner. Depletion of cells by the complement cascade is achieved by the addition of complement fixing antigen-specific antibodies and rabbit complement to the starting cell population. The cells are incubated for one hour at 37 degrees C and the lysed cells are subsequently removed by two rounds of washing. MAb with a high efficiency for complement fixation typically deplete 95-100% of the targeted cell population. Depending on the purification strategy for the targeted cell population, complement depletion can be used for cell purification or for the enrichment of cell populations that then can be further purified by a subsequent method. PMID:20139864

  7. Algal lectins as promising biomolecules for biomedical research.

    PubMed

    Singh, Ram Sarup; Thakur, Shivani Rani; Bansal, Parveen

    2015-02-01

    Lectins are natural bioactive ubiquitous proteins or glycoproteins of non-immune response that bind reversibly to glycans of glycoproteins, glycolipids and polysaccharides possessing at least one non-catalytic domain causing agglutination. Some of them consist of several carbohydrate-binding domains which endow them with the properties of cell agglutination or precipitation of glycoconjugates. Lectins are rampant in nature from plants, animals and microorganisms. Among microorganisms, algae are the potent source of lectins with unique properties specifically from red algae. The demand of peculiar and neoteric biologically active substances has intensified the developments on isolation and biomedical applications of new algal lectins. Comprehensively, algal lectins are used in biomedical research for antiviral, antinociceptive, anti-inflammatory, anti-tumor activities, etc. and in pharmaceutics for the fabrication of cost-effective protein expression systems and nutraceutics. In this review, an attempt has been made to collate the information on various biomedical applications of algal lectins. PMID:23855360

  8. Lectin cDNA and transgenic plants derived therefrom

    DOEpatents

    Raikhel, N.V.

    1994-01-04

    Transgenic plants containing cDNA encoding Gramineae lectin are described. The plants preferably contain cDNA coding for barley lectin and store the lectin in the leaves. The transgenic plants, particularly the leaves exhibit insecticidal and fungicidal properties. GOVERNMENT RIGHTS This application was funded under Department of Energy Contract DE-AC02-76ER01338. The U.S. Government has certain rights under this application and any patent issuing thereon. .

  9. Lectin cDNA and transgenic plants derived therefrom

    DOEpatents

    Raikhel, Natasha V.

    1994-01-04

    Transgenic plants containing cDNA encoding Gramineae lectin are described. The plants preferably contain cDNA coding for barley lectin and store the lectin in the leaves. The transgenic plants, particularly the leaves exhibit insecticidal and fungicidal properties. GOVERNMENT RIGHTS This application was funded under Department of Energy Contract DE-AC02-76ER01338. The U.S. Government has certain rights under this application and any patent issuing thereon.

  10. Towards functional glycomics by localization of binding sites for tissue lectins: lectin histochemical reactivity for galectins during diethylstilbestrol-induced kidney tumorigenesis in male Syrian hamster.

    PubMed

    Saussez, Sven; Lorfevre, Francois; Nonclercq, Denis; Laurent, Guy; Andr, Sabine; Journ, Fabrice; Kiss, Robert; Toubeau, Grard; Gabius, Hans-Joachim

    2006-07-01

    Endogenous lectins act as effectors of cellular activities such as growth regulation, migration, and adhesion. Following their immunohistochemical localization in our previous study (Saussez et al. in Histochem Cell Biol 123:29-41, 2005) we purified several galectins and used them as tools for monitoring accessible binding sites. Herein, we report the use of galectin histochemistry for the analysis of diethylstilbestrol (DES)-induced renal tumors in male Syrian hamster kidney (SHKT). Sections of normal kidney and DES-treated kidney were analyzed with biotinylated galectins-1, -3 (full-length and truncated), and -7. Accessible binding sites were detected, localization was predominantly extracellular and confined to medium-sized and large tumors. Monitoring the SHKT-derived HKT-1097 line, processed in vitro or as xenograft material, cytoplasmic and nuclear staining for galectins-1, -3, and -3tr could be observed. Adaptation of SHKT cells to long-term growth in culture is thus associated with emergence of this signal. Our data set illustrates the feasibility to complement immunohistochemical data by application of the tissue lectins as probes, and to detect regulation of galectin reactivity with differential characteristics within tumor progression in vivo and unique features of the tumor cell line in vitro and in vivo. PMID:16435123

  11. Archeal lectins: An identification through a genomic search.

    PubMed

    Abhinav, K V; Samuel, Ebenezer; Vijayan, M

    2016-01-01

    Forty-six lectin domains which have homologues among well established eukaryotic and bacterial lectins of known three-dimensional structure, have been identified through a search of 165 archeal genomes using a multipronged approach involving domain recognition, sequence search and analysis of binding sites. Twenty-one of them have the 7-bladed ?-propeller lectin fold while 16 have the ?-trefoil fold and 7 the legume lectin fold. The remainder assumes the C-type lectin, the ?-prism I and the tachylectin folds. Acceptable models of almost all of them could be generated using the appropriate lectins of known three-dimensional structure as templates, with binding sites at one or more expected locations. The work represents the first comprehensive bioinformatic study of archeal lectins. The presence of lectins with the same fold in all domains of life indicates their ancient origin well before the divergence of the three branches. Further work is necessary to identify archeal lectins which have no homologues among eukaryotic and bacterial species. Proteins 2016; 84:21-30. 2015 Wiley Periodicals, Inc. PMID:26492087

  12. Lectins discriminate between pathogenic and nonpathogenic South American trypanosomes

    SciTech Connect

    de Miranda Santos, I.K.; Pereira, M.E.

    1984-09-01

    Cell surface carbohydrates of Trypanosoma cruzi, Trypanosoma rangeli, and Trypanosoma conorhini were analyzed by a micro-agglutination assay employing 27 highly purified lectins and by binding assays using various /sup 125/I-labeled lectins. The following seven lectins discriminated between the trypanosomes: 1) tomato lectin (an N-acetyl-D-glucosamine-binding protein), both in purified form and as crude tomato juice; 2) Bauhinea purpurea and Sophora japonica lectins (both N-acetyl-D-galactosamine-binding proteins), which selectively agglutinated T. cruzi; 3) Vicia villosa (an N-acetyl-D-galactosamine-binding protein) which was specific for T. rangeli; 4) peanut lectin (a D-galactose-binding protein) both in purified form and as crude saline extract; and 5) Ulex europaeus and Lotus tetragonolobus (both L-fucose-binding proteins) lectins which reacted only with T. conorhini. Binding studies with 125I-labeled lectins were performed to find whether unagglutinated cells of the three different species of trypanosomes might have receptors for these lectins, in which case absence of agglutination could be due to a peculiar arrangement of the receptors. These assays essentially confirmed the agglutination experiments.

  13. Microbial Hijacking of ComplementToll-like Receptor Crosstalk*

    PubMed Central

    Wang, Min; Krauss, Jennifer L.; Domon, Hisanori; Hosur, Kavita B.; Liang, Shuang; Magotti, Paola; Triantafilou, Martha; Triantafilou, Kathy; Lambris, John D.; Hajishengallis, George

    2010-01-01

    Recent evidence suggests that complement and Toll-like receptors (TLRs) crosstalk to coordinate innate immunity. We report a novel immune subversion mechanism involving microbial exploitation of the ability of complement and TLRs for communication. Porphyromonas gingivalis, a major oral and systemic pathogen expressing complement C5 convertase-like activity, was shown to synergize with C5a for cAMP elevation resulting in macrophage immunosuppression and enhanced pathogen survival in vitro and in vivo. The cAMP synergy strictly required TLR2 signaling and a pertussis toxin- and thapsigargin-sensitive C5a receptor pathway, whereas protein kinase A and glycogen synthase kinase-3? acted as downstream effectors. Antagonistic blockade of the C5a receptor abrogated this evasive strategy and may thus have important therapeutic implications in periodontitis and atherosclerosis, where P. gingivalis is implicated. This first demonstration of complement-TLR crosstalk for immunosuppressive cAMP signaling indicates that pathogens may not simply undermine complement and/or TLRs as separate entities, but may also exploit their crosstalk pathways. PMID:20159852

  14. Complement activation by Coccidioides immitis: in vitro and clinical studies.

    PubMed Central

    Galgiani, J N; Yam, P; Petz, L D; Williams, P L; Stevens, D A

    1980-01-01

    Mycelial- or spherule-phase derivatives of Coccidioides immitis caused a decrease in vitro of total hemolytic complement in serum from a nonsensitized person. Activation involved both classic and alternative pathways as shown by deprssion of hemolytic C4 and by generation of products of activation of components C3, C4, and factor B. In addition, functional complement activity or immunoreactive levels of complement components or both were measured in 23 patients with self-limited or disseminated coccidioidomycosis. Low total hemolytic complement was found in nine, usually during the early phase of primary illness, and was transient. Hemolytic C4 was low, and the effect of inulin to decrease complement levels was blunted, suggested both classic and alternative pathways may be deficient. However, associated depression of immunoreactive levels of components assayed (C3, C4, C5, factor B, and properdin) was not consistently found. This disparity raises the possibility of enhanced in vitro inactivation analogous to activation by immune complexes. Images Fig. 2 PMID:6901703

  15. Algal Lectins as Potential HIV Microbicide Candidates

    PubMed Central

    Huskens, Dana; Schols, Dominique

    2012-01-01

    The development and use of topical microbicides potentially offers an additional strategy to reduce the spread of the Human Immunodeficiency Virus (HIV). Carbohydrate-binding agents (CBAs) that show specificity for high mannose carbohydrates on the surface of the heavily glycosylated envelope of HIV are endowed with potent anti-HIV activity. In fact, a number of algal lectins such as cyanovirin-N, microvirin, microcystis viridis lectin, scytovirin, Oscillatoria agardhii agglutinin and griffithsin are considered as potential microbicide candidates to prevent the sexual transmission of HIV through topical applications. They not only inhibit infection of cells by cell-free virus but they can also efficiently prevent virus transmission from virus-infected cells to uninfected CD4+ target T-lymphocytes and DC-SIGN-directed capture of HIV-1 and transmission to CD4+ T lymphocytes. This review focuses on the structural properties and carbohydrate specificity of these algal lectins, their antiviral activity against HIV and several other enveloped viruses, their safety profile and viral resistance patterns. PMID:22851920

  16. Lectin-mediated drug delivery: influence of mucin on cytoadhesion of plant lectins in vitro.

    PubMed

    Wirth, M; Gerhardt, K; Wurm, C; Gabor, F

    2002-02-19

    As the mucous layer represents the first barrier to peroral lectin-mediated drug delivery, the influence of mucin on the cytoadhesive properties of lectins was studied in vitro by establishing a rapid and simple microplate format assay using pig gastric mucin (PGM) for coating the wells. The lectin-binding capacity of mucin followed the order WGA>UEA-I>LCA=STL>PNA>DBA. The PGM-binding of wheat germ agglutinin (WGA) was strongly dependent on pH being highest at pH 5.0. In comparison, PGM-binding of WGA was about 15% at gastric pH and 60-70% at intestinal pH. This points to unimpeded gastric transit of WGA-grafted formulations and favorable conditions within the intestine for binding to mucus coated enterocytes. Moreover the WGA-PGM interaction was concentration-dependent, specific and fully reversible. According to a competitive assay in the presence of Caco-2 monolayers, the PGM-binding of WGA was saturated and influenced by the lectin-concentration yielding 28% Caco-2 bound WGA (125 ng WGA/0.29 cm(2) monolayer) and 68% Caco-2 bound WGA (4 microg WGA/0.29 cm(2) monolayer), respectively. Following on from these results, lectins are expected to suffer at least partially from premature inactivation by shed off mucus like bioadhesives of the first generation, however initial but reversible mucus-binding of lectins offers partititioning to the cell membrane followed by uptake into the enterocyte. PMID:11853930

  17. Sentential Complementation--An Overview.

    ERIC Educational Resources Information Center

    Nuessel, Frank H., Jr.

    A review of traditional and transformational studies on the phenomenon of sentential complementation (noun clauses) reveals many areas of agreement. Although some adherents of generative grammar may have occasionally obscured this aspect because of the offensive nature of their criticism of other modes of analysis, it is seen that, in several…

  18. Improvisation: A Complement to Curriculum

    ERIC Educational Resources Information Center

    Ronald, Green A.

    2006-01-01

    With the growth of standardized assessment benchmarks in both the public and private paradigms, testing performance matters to institutions more than ever. In an attempt to take as many hindering variables out of this process, such as test anxiety, socioeconomic influences, and latency in cognition, Improvisation: A Complement to Curriculum seeks…

  19. Cytolytic complement activity in otitis media with effusion.

    PubMed

    Nrki-Mkel, M; Meri, S

    2001-06-01

    Otitis media with effusion (OME) is a chronic inflammation persisting in the middle ear cavity of at least 8 weeks duration. Middle ear effusion (MEE; n = 38), samples from children suffering from OME were investigated for their direct cytolytic activity or an ability to enhance complement lysis of unsensitized bystander cells. Thirteen of the 38 MEEs had direct endogenous haemolytic activity and 27 samples had an ability to enhance serum-initiated lysis. Using an enzyme immunoassay, high levels of terminal complement complexes (TCC) were detected in the MEE samples (mean 34.1 microg/ml, range 5--89 microg/ml). This indicated strong local complement activation that had progressed to the terminal stage. As one potential factor promoting complement activation we identified both monomeric and trimeric properdin in MEE by Western blotting. By stabilizing C3 and C5 convertases properdin accelerates the alternative and terminal pathways of complement. On the other hand, the membrane attack complex (MAC) inhibitor CD59, which was found to be extensively shed into the MEE in a functionally active form, may control excessive cytotoxicity of the MEE. In conclusion, intense complement activation, up to the terminal level, maintains ongoing inflammation in the middle ear cavity and can pose a threat to the local epithelium. PMID:11472396

  20. Cytolytic complement activity in otitis media with effusion

    PubMed Central

    Nrki-Mkel, M; Meri, S

    2001-01-01

    Otitis media with effusion (OME) is a chronic inflammation persisting in the middle ear cavity of at least 8 weeks duration. Middle ear effusion (MEE; n = 38), samples from children suffering from OME were investigated for their direct cytolytic activity or an ability to enhance complement lysis of unsensitized bystander cells. Thirteen of the 38 MEEs had direct endogenous haemolytic activity and 27 samples had an ability to enhance serum-initiated lysis. Using an enzyme immunoassay, high levels of terminal complement complexes (TCC) were detected in the MEE samples (mean 341 g/ml, range 589 g/ml). This indicated strong local complement activation that had progressed to the terminal stage. As one potential factor promoting complement activation we identified both monomeric and trimeric properdin in MEE by Western blotting. By stabilizing C3 and C5 convertases properdin accelerates the alternative and terminal pathways of complement. On the other hand, the membrane attack complex (MAC) inhibitor CD59, which was found to be extensively shed into the MEE in a functionally active form, may control excessive cytotoxicity of the MEE. In conclusion, intense complement activation, up to the terminal level, maintains ongoing inflammation in the middle ear cavity and can pose a threat to the local epithelium. PMID:11472396

  1. Role of C5b-9 complement complex and response gene to complement-32 (RGC-32) in cancer.

    PubMed

    Vlaicu, Sonia I; Tegla, Cosmin A; Cudrici, Cornelia D; Danoff, Jacob; Madani, Hassan; Sugarman, Adam; Niculescu, Florin; Mircea, Petru A; Rus, Violeta; Rus, Horea

    2013-05-01

    Complement system activation plays an important role in both innate and acquired immunity, with the activation of complement and the subsequent formation of C5b-9 terminal complement complex on cell membranes inducing target cell death. Recognition of this role for C5b-9 leads to the assumption that C5b-9 might play an antitumor role. However, sublytic C5b-9 induces cell cycle progression by activating signal transduction pathways and transcription factors in cancer cells, indicating a role in tumor promotion for this complement complex. The induction of the cell cycle by C5b-9 is dependent upon the activation of the phosphatidylinositol 3-kinase (PI3K)/Akt/FOXO1 and ERK1 pathways in a Gi protein-dependent manner. C5b-9 also induces response gene to complement (RGC)-32, a gene that plays a role in cell cycle promotion through activation of Akt and the CDC2 kinase. RGC-32 is expressed by tumor cells and plays a dual role in cancers, in that it has both a tumor suppressor role and tumor-promoting activity. Thus, through the activation of tumor cells, the C5b-9-mediated induction of the cell cycle plays an important role in tumor proliferation and oncogenesis. PMID:23247987

  2. Complement activation in the context of stem cells and tissue repair

    PubMed Central

    Schraufstatter, Ingrid U; Khaldoyanidi, Sophia K; DiScipio, Richard G

    2015-01-01

    The complement pathway is best known for its role in immune surveillance and inflammation. However, its ability of opsonizing and removing not only pathogens, but also necrotic and apoptotic cells, is a phylogenetically ancient means of initiating tissue repair. The means and mechanisms of complement-mediated tissue repair are discussed in this review. There is increasing evidence that complement activation contributes to tissue repair at several levels. These range from the chemo-attraction of stem and progenitor cells to areas of complement activation, to increased survival of various cell types in the presence of split products of complement, and to the production of trophic factors by cells activated by the anaphylatoxins C3a and C5a. This repair aspect of complement biology has not found sufficient appreciation until recently. The following will examine this aspect of complement biology with an emphasis on the anaphylatoxins C3a and C5a. PMID:26435769

  3. Complement activation in the context of stem cells and tissue repair.

    PubMed

    Schraufstatter, Ingrid U; Khaldoyanidi, Sophia K; DiScipio, Richard G

    2015-09-26

    The complement pathway is best known for its role in immune surveillance and inflammation. However, its ability of opsonizing and removing not only pathogens, but also necrotic and apoptotic cells, is a phylogenetically ancient means of initiating tissue repair. The means and mechanisms of complement-mediated tissue repair are discussed in this review. There is increasing evidence that complement activation contributes to tissue repair at several levels. These range from the chemo-attraction of stem and progenitor cells to areas of complement activation, to increased survival of various cell types in the presence of split products of complement, and to the production of trophic factors by cells activated by the anaphylatoxins C3a and C5a. This repair aspect of complement biology has not found sufficient appreciation until recently. The following will examine this aspect of complement biology with an emphasis on the anaphylatoxins C3a and C5a. PMID:26435769

  4. Reincarnation of ancient links between coagulation and complement.

    PubMed

    Conway, E M

    2015-06-01

    Throughout evolution, organisms have developed means to contain wounds by simultaneously limiting bleeding and eliminating pathogens and damaged host cells via the recruitment of innate defense mechanisms. Disease emerges when there is unchecked activation of innate immune and/or coagulation responses. A key component of innate immunity is the complement system. Concurrent excess activation of coagulation and complement - two major blood-borne proteolytic pathways - is evident in numerous diseases, including atherosclerosis, diabetes, venous thromboembolic disease, thrombotic microangiopathies, arthritis, cancer, and infectious diseases. Delineating the cross-talk between these two cascades will uncover novel therapeutic insights. PMID:26149013

  5. Exploring the Innate Immune System: Using Complement-Medicated Cell Lysis in the Classroom

    ERIC Educational Resources Information Center

    Fuller, Kevin G.

    2008-01-01

    The protein complement pathway comprises an important part of the innate immunity. The use of serum to demonstrate complement-mediated destruction across a series of bacterial dilutions allows an instructor to introduce a number of important biological concepts such as bacterial growth, activation cascades, and adaptive versus innate immunity.

  6. Characterization of the complement sensitivity of paroxysmal nocturnal hemoglobinuria erythrocytes.

    PubMed Central

    Parker, C J; Wiedmer, T; Sims, P J; Rosse, W F

    1985-01-01

    The affected erythrocytes of paroxysmal nocturnal hemoglobinuria (PNH II and PNH III cells) are abnormally sensitive to complement-mediated lysis. Normal human erythrocytes chemically modified by treatment with 2-amino-ethylisothiouronium bromide (AET) have been used as models for PNH cells inasmuch as they also exhibit an enhanced susceptibility to complement. To investigate the bases for the greater sensitivity of these abnormal cells to complement-mediated lysis, we compared binding of C3 and constituents of the membrane attack complex to normal, PNH II, PNH III, and AET-treated cells after classical pathway activation by antibody and fluid-phase activation by cobra venom factor complexes. When whole serum complement was activated by antibody, there was increased binding of C3 and C9 to PNH II, PNH III, and AET-treated cells, although the binding of these complement components to PNH II and PNH III cells was considerably greater than their binding to the AET-treated cells. In addition, all of the abnormal cell types showed a greater degree of lysis per C9 bound than did the normal erythrocytes. PNH III and AET-treated cells were readily lysed by fluid-phase activation of complement, whereas normal and PNH II erythrocytes were not susceptible to bystander lysis. The greater hemolysis of PNH III and AET-treated cells in this reactive lysis system was due to a quantitative increase in binding of constituents of the membrane attack complex. This more efficient binding of the terminal components after fluid-phase activation of whole serum complement was not mediated by cell-bound C3 fragments. These investigations demonstrate that the molecular events that characterize the enhanced susceptibility of PNH II, PNH III, and AET-treated erythrocytes to complement-mediated lysis are heterogeneous. Images PMID:4008653

  7. Complement activation patterns in atypical haemolytic uraemic syndrome during acute phase and in remission.

    PubMed

    Volokhina, E B; Westra, D; van der Velden, T J A M; van de Kar, N C A J; Mollnes, T E; van den Heuvel, L P

    2015-08-01

    Atypical haemolytic uraemic syndrome (aHUS) is associated with (genetic) alterations in alternative complement pathway. Nevertheless, comprehensive evidence that the complement system in aHUS patients is more prone to activation is still lacking. Therefore, we performed a thorough analysis of complement activation in acute phase and in remission of this disease. Complement activation patterns of the aHUS patients in acute phase and in remission were compared to those of healthy controls. Background levels of complement activation products C3b/c, C3bBbP and terminal complement complex (TCC) were measured using enzyme-linked immunosorbent assay (ELISA) in ethylenediamine tetraacetic acid (EDTA) plasma. In vitro-triggered complement activation in serum samples was studied using zymosan-coating and pathway-specific assay. Furthermore, efficiencies of the C3b/c, C3bBbP and TCC generation in fluid phase during spontaneous activation were analysed. Patients with acute aHUS showed elevated levels of C3b/c (P?complement factor B mutation we illustrated normalization of complement activation during aHUS recovery. Serum samples from patients in remission showed normal in vitro patterns of complement activation and demonstrated normal kinetics of complement activation in the fluid phase. Our data indicate that while aHUS patients have clearly activated complement in acute phase of the disease, this is not the case in remission of aHUS. This knowledge provides important insight into complement regulation in aHUS and may have an impact on monitoring of these patients, particularly when using complement inhibition therapy. PMID:25079699

  8. The lectin like domain of thrombomodulin is involved in the defence against pyelonephritis.

    PubMed

    Lattenist, Lionel; Teske, Gwendoline; Claessen, Nike; Florquin, Sandrine; Conway, Edward M; Roelofs, Joris J T H

    2015-12-01

    Pyelonephritis, a common complication of urinary tract infections, is frequently associated with kidney scarring and may lead to end-stage renal disease. During bacterial infections inflammatory and coagulation pathways and their mutual interaction are playing pivotal roles in the host response. Given that thrombomodulin (TM) is crucially involved in the interplay between coagulation and inflammation, we aimed to investigate the roles of its EGF and lectin-like domains in inflammation during acute pyelonephritis. Indeed, the EGF-like and the lectin-like domains of TM, are especially known to orchestrate inflammation and coagulation in different ways. Acute pyelonephritis was induced by intravesical inoculation of 110(8) CFU of uropathogenic Escherichia coli in two strains of TM transgenic mice. TM(pro/pro) mice carry a mutation in the EGF-like domain making them unable to activate protein C, an anticoagulant and anti-inflammatory protein. TM(LeD/LeD) mice lack the lectin-like domain of TM, which is critical for its anti-inflammatory and cytoprotective properties. Mice were sacrificed 24 and 48h after inoculation. Bacterial loads, the immune response and the activation of coagulation were evaluated in the kidney and the bladder. TM(LeD/LeD) mice showed elevated bacterial load in bladder and kidneys compared to WT mice, whereas TM(pro/pro) had similar bacterial load as WT mice. TM(LeD/LeD) mice displayed a reduced local production of pro-inflammatory cytokines and neutrophil renal infiltration. Activation of coagulation was comparable in TM(LeD/LeD) and WT mice. From these data, we conclude that the lectin-like domain of thrombomodulin is critically involved in host defence against E. coli induced acute pyelonephritis. PMID:26573396

  9. L-Type lectin from the kuruma shrimp Marsupenaeus japonicus promotes hemocyte phagocytosis.

    PubMed

    Xu, Sen; Wang, Lei; Wang, Xian-Wei; Zhao, Yan-Ran; Bi, Wen-Jie; Zhao, Xiao-Fan; Wang, Jin-Xing

    2014-06-01

    L-Type lectins (LTLs) contain a luminal carbohydrate recognition domain, which exhibits homology to leguminous lectins. These type I membrane proteins are involved in the early secretory pathway of animals, and have functions in glycoprotein sorting, trafficking and targeting. Recent studies suggest that LTLs may be involved in immune responses in vertebrates, but no functional studies have been reported. This study reports an LTL, designated as MjLTL1, from the kuruma shrimp Marsupenaeus japonicus. MjLTL consists of a signal peptide, leguminous lectin domain, and transmembrane region. It was upregulated following challenge of shrimp with Vibrio anguillarum. MjLTL1 could agglutinate several bacteria with the presence of calcium, and bind to several Gram-positive and Gram-negative bacteria through lipopolysaccharide and peptidoglycan binding. MjLTL1 could enhance the clearance of V. anguillarum in vivo. MjLTL1 silencing by RNA interference could impair bacterial clearance ability. Further study suggested that MjLTL1 promoted hemocyte phagocytosis. To analyze the possible mechanism, a disintegrin and metalloprotease-like protein (MjADAM) mediating the proteolytic release of extracellular domains from the membrane-bound precursors was also studied in the shrimp. MjADAM exhibited similar tissue location and expression profiles to MjLTL1. After knockdown of MjADAM, the hemocyte phagocytosis rate also declined significantly. ADAM was reported to have an ectodomain shedding function to LTL and release the ectodomain of the lectin from cell membrane. Therefore, our results suggest that the extracellular domain of MjLTL1 might be released from the cell surface as a soluble protein by MjADAM, and function as an opsonin involved in the antibacterial immune responses in shrimp. PMID:24508102

  10. Identification of hot spots in the variola virus complement inhibitor (SPICE) for human complement regulation.

    PubMed

    Yadav, Viveka Nand; Pyaram, Kalyani; Mullick, Jayati; Sahu, Arvind

    2008-04-01

    Variola virus, the causative agent of smallpox, encodes a soluble complement regulator named SPICE. Previously, SPICE has been shown to be much more potent in inactivating human complement than the vaccinia virus complement control protein (VCP), although they differ only in 11 amino acid residues. In the present study, we have expressed SPICE, VCP, and mutants of VCP by substituting each or more of the 11 non-variant VCP residues with the corresponding residue of SPICE to identify hot spots that impart functional advantage to SPICE over VCP. Our data indicate that (i) SPICE is approximately 90-fold more potent than VCP in inactivating human C3b, and the residues Y98, Y103, K108 and K120 are predominantly responsible for its enhanced activity; (ii) SPICE is 5.4-fold more potent in inactivating human C4b, and residues Y98, Y103, K108, K120 and L193 mainly dictate this increase; (iii) the classical pathway decay-accelerating activity of activity is only twofold higher than that of VCP, and the 11 mutations in SPICE do not significantly affect this activity; (iv) SPICE possesses significantly greater binding ability to human C3b compared to VCP, although its binding to human C4b is lower than that of VCP; (v) residue N144 is largely responsible for the increased binding of SPICE to human C3b; and (vi) the human specificity of SPICE is dictated primarily by residues Y98, Y103, K108, and K120 since these are enough to formulate VCP as potent as SPICE. Together, these results suggest that principally 4 of the 11 residues that differ between SPICE and VCP partake in its enhanced function against human complement. PMID:18216095

  11. A Molecular Insight into Complement Evasion by the Staphylococcal Complement Inhibitor Protein Family1

    PubMed Central

    Ricklin, Daniel; Tzekou, Apostolia; Garcia, Brandon L.; Hammel, Michal; McWhorter, William J.; Sfyroera, Georgia; Wu, You-Qiang; Holers, V. Michael; Herbert, Andrew P.; Barlow, Paul N.; Geisbrecht, Brian V.; Lambris, John D.

    2010-01-01

    Staphylococcus aureus possesses an impressive arsenal of complement evasion proteins that help the bacterium escape attack of the immune system. The staphylococcal complement inhibitor (SCIN) protein exhibits a particularly high potency and was previously shown to block complement by acting at the level of the C3 convertases. However, many details about the exact binding and inhibitory mechanism remained unclear. In this study, we demonstrate that SCIN directly binds with nanomolar affinity to a functionally important area of C3b that lies near the C terminus of its β-chain. Direct competition of SCIN with factor B for C3b slightly decreased the formation of surface-bound convertase. However, the main inhibitory effect can be attributed to an entrapment of the assembled convertase in an inactive state. Whereas native C3 is still able to bind to the blocked convertase, no generation and deposition of C3b could be detected in the presence of SCIN. Furthermore, SCIN strongly competes with the binding of factor H to C3b and influences its regulatory activities: the SCIN-stabilized convertase was essentially insensitive to decay acceleration by factor H and the factor I- and H-mediated conversion of surface-bound C3b to iC3b was significantly reduced. By targeting a key area on C3b, SCIN is able to block several essential functions within the alternative pathway, which explains the high potency of the inhibitor. Our findings provide an important insight into complement evasion strategies by S. aureus and may act as a base for further functional studies. PMID:19625656

  12. Membrane attack by complement: the assembly and biology of terminal complement complexes.

    PubMed

    Tegla, Cosmin A; Cudrici, Cornelia; Patel, Snehal; Trippe, Richard; Rus, Violeta; Niculescu, Florin; Rus, Horea

    2011-10-01

    Complement system activation plays an important role in both innate and acquired immunity. Activation of the complement and the subsequent formation of C5b-9 channels (the membrane attack complex) on the cell membranes lead to cell death. However, when the number of channels assembled on the surface of nucleated cells is limited, sublytic C5b-9 can induce cell cycle progression by activating signal transduction pathways and transcription factors and inhibiting apoptosis. This induction by C5b-9 is dependent upon the activation of the phosphatidylinositol 3-kinase/Akt/FOXO1 and ERK1 pathways in a Gi protein-dependent manner. C5b-9 induces sequential activation of CDK4 and CDK2, enabling the G1/S-phase transition and cellular proliferation. In addition, it induces RGC-32, a novel gene that plays a role in cell cycle activation by interacting with Akt and the cyclin B1-CDC2 complex. C5b-9 also inhibits apoptosis by inducing the phosphorylation of Bad and blocking the activation of FLIP, caspase-8, and Bid cleavage. Thus, sublytic C5b-9 plays an important role in cell activation, proliferation, and differentiation, thereby contributing to the maintenance of cell and tissue homeostasis. PMID:21850539

  13. Assessment of lectin inactivation by heat and digestion.

    PubMed

    Pusztai, A; Grant, G

    1998-01-01

    Proteins/glycoproteins from plants, particularly lectins, are more resistant to heat denaturation than animal proteins (1, 2). With legume seeds, whose lectin content is appreciable, this presents potentially serious problems in nutritional practice. Therefore, before they can be used safely, legume-based food/ feeds usually require thorough and expensive heat processing to inactivate antinutritive components. Indeed, dry or moist heating of seeds at 70°C for several h has little or no effect on their lectin activity (Fig. 1) and treatment at much higher temperatures is needed to inactivate the biological and antinutritional effects of legume lectins (1, 2). The safety aspect is even more serious with some monocot lectins, such as wheatgerm agglutinin or a number of oilseed lectins, such as peanut agglutinin and many others because they are extremely heat stable and normal cooking or other conventional heat treatments may fail to inactivate them (3) Thus, the best way to avoid potential harmful effects of these heat-resistant lectins is to limit their dietary intake to a minimum. Fig. 1. Loss of lectin activity during aqueous heat treatment of soybean at various temperatures. PMID:21374488

  14. Plant Lectins: Wheat Defense Strategy Against Hessian Fly

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Plants produce a variety of defense proteins, including lectins in response to attack by phytophagous insects. Ultrastructural studies reveal that binding to insect gut structures and resistance to proteolytic degradation by insect digestive enzymes are the two main prerequisites for the lectins to...

  15. 21 CFR 864.9550 - Lectins and protectins.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Lectins and protectins. 864.9550 Section 864.9550 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Products Used In Establishments That Manufacture Blood and Blood Products 864.9550 Lectins...

  16. 21 CFR 864.9550 - Lectins and protectins.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Lectins and protectins. 864.9550 Section 864.9550 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Products Used In Establishments That Manufacture Blood and Blood Products 864.9550 Lectins...

  17. 21 CFR 864.9550 - Lectins and protectins.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Lectins and protectins. 864.9550 Section 864.9550 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Products Used In Establishments That Manufacture Blood and Blood Products 864.9550 Lectins...

  18. Porifera Lectins: Diversity, Physiological Roles and Biotechnological Potential

    PubMed Central

    Gardères, Johan; Bourguet-Kondracki, Marie-Lise; Hamer, Bojan; Batel, Renato; Schröder, Heinz C.; Müller, Werner E. G.

    2015-01-01

    An overview on the diversity of 39 lectins from the phylum Porifera is presented, including 38 lectins, which were identified from the class of demosponges, and one lectin from the class of hexactinellida. Their purification from crude extracts was mainly performed by using affinity chromatography and gel filtration techniques. Other protocols were also developed in order to collect and study sponge lectins, including screening of sponge genomes and expression in heterologous bacterial systems. The characterization of the lectins was performed by Edman degradation or mass spectrometry. Regarding their physiological roles, sponge lectins showed to be involved in morphogenesis and cell interaction, biomineralization and spiculogenesis, as well as host defense mechanisms and potentially in the association between the sponge and its microorganisms. In addition, these lectins exhibited a broad range of bioactivities, including modulation of inflammatory response, antimicrobial and cytotoxic activities, as well as anticancer and neuromodulatory activity. In view of their potential pharmacological applications, sponge lectins constitute promising molecules of biotechnological interest. PMID:26262628

  19. Porifera Lectins: Diversity, Physiological Roles and Biotechnological Potential.

    PubMed

    Gardres, Johan; Bourguet-Kondracki, Marie-Lise; Hamer, Bojan; Batel, Renato; Schrder, Heinz C; Mller, Werner E G

    2015-08-01

    An overview on the diversity of 39 lectins from the phylum Porifera is presented, including 38 lectins, which were identified from the class of demosponges, and one lectin from the class of hexactinellida. Their purification from crude extracts was mainly performed by using affinity chromatography and gel filtration techniques. Other protocols were also developed in order to collect and study sponge lectins, including screening of sponge genomes and expression in heterologous bacterial systems. The characterization of the lectins was performed by Edman degradation or mass spectrometry. Regarding their physiological roles, sponge lectins showed to be involved in morphogenesis and cell interaction, biomineralization and spiculogenesis, as well as host defense mechanisms and potentially in the association between the sponge and its microorganisms. In addition, these lectins exhibited a broad range of bioactivities, including modulation of inflammatory response, antimicrobial and cytotoxic activities, as well as anticancer and neuromodulatory activity. In view of their potential pharmacological applications, sponge lectins constitute promising molecules of biotechnological interest. PMID:26262628

  20. 21 CFR 864.9550 - Lectins and protectins.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ...) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Products Used In Establishments That Manufacture Blood and Blood Products 864.9550 Lectins and protectins. (a) Identification. Lectins and protectins are proteins derived from plants and lower animals that cause cell agglutination in the presence of...

  1. 21 CFR 864.9550 - Lectins and protectins.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ...) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Products Used In Establishments That Manufacture Blood and Blood Products 864.9550 Lectins and protectins. (a) Identification. Lectins and protectins are proteins derived from plants and lower animals that cause cell agglutination in the presence of...

  2. Lectin-binding properties of Aeromonas caviae strains

    PubMed Central

    Rocha-de-Souza, Cláudio M.; Hirata-Jr, Raphael; Mattos-Guaraldi, Ana L.; Freitas-Almeida, Angela C.; Andrade, Arnaldo F. B.

    2008-01-01

    The cell surface carbohydrates of four strains of Aeromonas caviae were analyzed by agglutination and lectin-binding assays employing twenty highly purified lectins encompassing all sugar specificities. With the exception of L-fucose and sialic acid, the sugar residues were detected in A. caviae strains. A marked difference, however, in the pattern of cell surface carbohydrates in different A. caviae isolates was observed. Specific receptors for Tritricum vulgaris (WGA), Lycopersicon esculentum (LEL) and Solanum tuberosum (STA) (D-GlcNAc-binding lectins) were found only in ATCC 15468 strain, whereas Euonymus europaeus (EEL, D-Gal-binding lectin) sites were present exclusively in AeQ32 strain, those for Helix pomatia (HPA, D-GalNAc-binding lectin) in AeC398 and AeV11 strains, and for Canavalia ensiformes (Con A, D-Man-binding lectin) in ATCC 15468, AeC398, AeQ32 and AeV11 strains, after bacterial growing at 37°C. On the other hand, specific receptors for WGA and EEL were completely abrogated growing the bacteria at 22°C. Binding studies with 125I- labeled lectins from WGA, EEL and Con A were performed. These assays essentially confirmed the selectivity, demonstrated in the agglutination assays of these lectins for the A. caviae strains. PMID:24031204

  3. Decidual expression and localization of human surfactant protein SP-A and SP-D, and complement protein C1q.

    PubMed

    Madhukaran, Shanmuga Priyaa; Kishore, Uday; Jamil, Kaiser; Choolani, Mahesh; Lu, Jinhua

    2015-08-01

    Surfactant proteins SP-A and SP-D, and complement protein C1q are soluble innate immune pattern recognizing molecules. SP-A, SP-D and C1q have an overall similar structure composed of an N-terminal triple-helical collagen region that is followed by a trimeric globular domain. While SP-A and SP-D belong to the collectin family (collagen containing lectin), C1q is the first recognition subcomponent of the classical pathway of the complement system. Recently, SP-A, SP-D and C1q have been considered to play important roles in early and late pregnancy. However, their expression in early human decidua has not been examined. Here, we investigated whether SP-A, SP-D and C1q are expressed within first trimester decidua in humans and their expression is associated with trophoblasts and decidual stromal cells. Decidual samples from women undergoing elective vaginal termination of pregnancy during first trimester were obtained from 25 subjects. Immunohistochemical studies using anti-human SP-A, anti-human SP-D and anti-human C1q antibodies were performed on decidual tissue sections along with anti-vimentin and cytokeratin-7 antibodies to identify stromal cells and trophoblasts. The expression was also examined by immunostaining and PCR using decidual and stromal cells. C1q expression was significantly higher when compared to SP-A and SP-D in the first trimester human decidua. Double immunostaining revealed that all stromal cells and trophoblasts expressed SP-A, SP-D and C1q, while only few invasive trophoblasts expressed C1q. Thus, expression of SP-A, SP-D and C1q in human decidua during first trimester suggests potential role of SP-A, SP-D and C1q during the early stages of pregnancy including implantation, trophoblast invasion and placental development. PMID:25829244

  4. Complement Activation in Placental Malaria

    PubMed Central

    McDonald, Chloe R.; Tran, Vanessa; Kain, Kevin C.

    2015-01-01

    Sixty percent of all pregnancies worldwide occur in malaria endemic regions. Pregnant women are at greater risk of malaria infection than their non-pregnant counterparts and have a higher risk of adverse birth outcomes including low birth weight resulting from intrauterine growth restriction and/or preterm birth. The complement system plays an essential role in placental and fetal development as well as the host innate immune response to malaria infection. Excessive or dysregulated complement activation has been associated with the pathobiology of severe malaria and with poor pregnancy outcomes, dependent and independent of infection. Here we review the role of complement in malaria and pregnancy and discuss its part in mediating altered placental angiogenesis, malaria-induced adverse birth outcomes, and disruptions to the in utero environment with possible consequences on fetal neurodevelopment. A detailed understanding of the mechanisms underlying adverse birth outcomes, and the impact of maternal malaria infection on fetal neurodevelopment, may lead to biomarkers to identify at-risk pregnancies and novel therapeutic interventions to prevent these complications. PMID:26733992

  5. Complement system activation in ANCA vasculitis: A translational success story?

    PubMed

    Kallenberg, Cees G M; Heeringa, Peter

    2015-11-01

    The ANCA-associated vasculitides (AAV) are characterized by pauci-immune necrotizing small to medium size vessel vasculitis frequently including necrotizing crescentric glomerulonephritis. Neutrophil activation by ANCA appears a primary pathogenic event. More recently, the complement system has been shown to be involved as well. Activation of the alternative pathway of complement, at least in part via activated neutrophils, results, amongst others, in the generation of C5a, a strong chemoattractant for neutrophils. C5a is also effective in neutrophil priming, a process leading to surface expression of the ANCA antigens so enabling neutrophils to be further activated by ANCA. Both in vitro and in vivo experimental data and histopathological studies from AAV patients underscore the role of complement, and particularly of C5a, in the pathophysiology of AAV. Preliminary data show that blocking of the C5a-receptor is a promising approach in the treatment of AAV. PMID:26597208

  6. Biotoxicity assays for fruiting body lectins and other cytoplasmic proteins.

    PubMed

    Künzler, Markus; Bleuler-Martinez, Silvia; Butschi, Alex; Garbani, Mattia; Lüthy, Peter; Hengartner, Michael O; Aebi, Markus

    2010-01-01

    Recent studies suggest that a specific class of fungal lectins, commonly referred to as fruiting body lectins, play a role as effector molecules in the defense of fungi against predators and parasites. Hallmarks of these fungal lectins are their specific expression in reproductive structures, fruiting bodies, and/or sclerotia and their synthesis on free ribosomes in the cytoplasm. Fruiting body lectins are released upon damage of the fungal cell and bind to specific carbohydrate structures of predators and parasites, which leads to deterrence, inhibition of growth, and development or even killing of these organisms. Here, we describe assays to assess the toxicity of such lectins and other cytoplasmic proteins toward three different model organisms: the insect Aedes aegypti, the nematode Caenorhabditis elegans, and the amoeba Acanthamoeba castellanii. All three assays are based on heterologous expression of the examined proteins in the cytoplasm of Escherichia coli and feeding of these recombinant bacteria to omnivorous and bacterivorous organisms. PMID:20816208

  7. Structure-function relationship of monocot mannose-binding lectins.

    PubMed Central

    Barre, A; Van Damme, E J; Peumans, W J; Rougé, P

    1996-01-01

    The monocot mannose-binding lectins are an extended superfamily of structurally and evolutionarily related proteins, which until now have been isolated from species of the Amaryllidaceae, Alliaceae, Araceae, Orchidaceae, and Liliaceae. To explain the obvious differences in biological activities, the structure-function relationships of the monocot mannose-binding lectins were studied by a combination of glycan-binding studies and molecular modeling using the deduced amino acid sequences of the currently known lectins. Molecular modeling indicated that the number of active mannose-binding sites per monomer varies between three and zero. Since the number of binding sites is fairly well correlated with the binding activity measured by surface plasmon resonance, and is also in good agreement with the results of previous studies of the biological activities of the mannose-binding lectins, molecular modeling is of great value for predicting which lectins are best suited for a particular application. PMID:8972598

  8. Isolation and characterization of Lentinus edodes (Berk.) singer extracellular lectins.

    PubMed

    Tsivileva, O M; Nikitina, V E; Loshchinina, E A

    2008-10-01

    Lectin preparations have been isolated and purified from the culture liquid of the xylotrophic basidiomycete Lentinus edodes (Berk.) Singer [Lentinula edodes (Berk.) Pegler]. The culture of L. edodes F-249 synthesizes two extracellular lectins different in composition and physicochemical properties. Extracellular lectin L1 from L. edodes is a glycoprotein of mono-subunit structure with molecular weight of 43 kD. L1 is comprised of 10.5 +/- 1.0% (w/w) carbohydrates represented by glucose (Glc). Extracellular lectin L2 is a proteoglycan of mono-subunit structure with molecular weight of 37 kD. L2 is comprised of 90.3 +/- 1.0% (w/w) carbohydrates represented by Glc (73% of the total mass of the carbohydrate moiety of the lectin molecule) and galactose (Gal) (27% of the total mass of the carbohydrate part of the lectin molecule). The content of Asn in L2 is high, i.e. 42% (w/w) of total amino acids. This fact along with the composition of the carbohydrate part of the molecule (Glc + Gal) allows one to assign L2 to N-asparagine-bound proteins. Both lectins are specific to D-Gal and lactose (Lac) at an equal for L1 and L2 minimal inhibiting concentration of these carbohydrates (2.08 mM Gal and 8.33 mM Lac). Other carbohydrates to which the lectins show affinity are different for the two lectins: Rha (4.16 mM) for L1 and Ara (4.16 mM) and mannitol (8.33 mM) for L2. The purified extracellular lectins of L. edodes are highly selective at recognition of definite structures on the surface of trypsinized rabbit erythrocytes and do not react with the erythrocytes of other animals and humans. PMID:18991563

  9. Quantitative evaluation of lectin-reactive glycoforms of ?(1)-acid glycoprotein using lectin affinity capillary electrophoresis with fluorescence detection.

    PubMed

    Shimura, Kiyohito; Tamura, Mayumi; Toda, Tosifusa; Yazawa, Shin; Kasai, Ken-ichi

    2011-08-01

    ?(1)-Acid glycoprotein (AGP) was previously shown to be a marker candidate of disease progression and prognosis of patients with malignancies by analysis of its glycoforms via lectins. Herein, affinity capillary electrophoresis of fluorescein-labeled AGP using lectins with the aid of laser-induced fluorescence detection was developed for quantitative evaluation of the fractional ratios of concanavalin A-reactive or Aleuria aurantia lectin-reactive AGP. Labeled AGP was applied at the anodic end of a fused-silica capillary (50??m id, 360??m od, 27?cm long) coated with linear polyacryloyl-?-alanyl-?-alanine, and electrophoresis was carried out for about 10?min in 60?mM 3-morpholinopropane-1-sulfonic acid-NaOH buffer (pH 7.35). Addition of the lectins to the anode buffer resulted in the separation of lectin-reactive glycoform peaks from lectin-non-reactive glycoform peaks. Quantification of the peak area of each group revealed that the percent of lectin-reactive AGP is independent of a labeling ratio ranging from 0.4 to 1.5?mol fluorescein/mol AGP, i.e. the standard deviation of 0.5% for an average of 59.9% (n=3). In combination with a facile procedure for micro-purification of AGP from serum, the present procedure, marking the reactivity of AGP with lectins, should be useful in determining the prognosis for a large number of patients with malignancies. PMID:21766474

  10. The Liverwort Contains a Lectin That Is Structurally and Evolutionary Related to the Monocot Mannose-Binding Lectins1

    PubMed Central

    Peumans, Willy J.; Barre, Annick; Bras, Julien; Roug, Pierre; Proost, Paul; Van Damme, Els J.M.

    2002-01-01

    A mannose (Man)-binding lectin has been isolated and characterized from the thallus of the liverwort Marchantia polymorpha. N-terminal sequencing indicated that the M. polymorpha agglutinin (Marpola) shares sequence similarity with the superfamily of monocot Man-binding lectins. Searches in the databases yielded expressed sequence tags encoding Marpola. Sequence analysis, molecular modeling, and docking experiments revealed striking structural similarities between Marpola and the monocot Man-binding lectins. Activity and specificity studies further indicated that Marpola is a much stronger agglutinin than the Galanthus nivalis agglutinin and exhibits a preference for methylated Man and glucose, which is unprecedented within the family of monocot Man-binding lectins. The discovery of Marpola allows us, for the first time, to corroborate the evolutionary relationship between a lectin from a lower plant and a well-established lectin family from flowering plants. In addition, the identification of Marpola sheds a new light on the molecular evolution of the superfamily of monocot Man-binding lectins. Beside evolutionary considerations, the occurrence of a G. nivalis agglutinin homolog in a lower plant necessitates the rethinking of the physiological role of the whole family of monocot Man-binding lectins. PMID:12114560

  11. MMBL proteins: from lectin to bacteriocin.

    PubMed

    Ghequire, Maarten G K; Loris, Remy; De Mot, Ren

    2012-12-01

    Arguably, bacteriocins deployed in warfare among related bacteria are among the most diverse proteinacous compounds with respect to structure and mode of action. Identification of the first prokaryotic member of the so-called MMBLs (monocot mannose-binding lectins) or GNA (Galanthus nivalis agglutinin) lectin family and discovery of its genus-specific killer activity in the Gram-negative bacteria Pseudomonas and Xanthomonas has added yet another kind of toxin to this group of allelopathic molecules. This novel feature is reminiscent of the protective function, on the basis of antifungal, insecticidal, nematicidal or antiviral activity, assigned to or proposed for several of the eukaryotic MMBL proteins that are ubiquitously distributed among monocot plants, but also occur in some other plants, fish, sponges, amoebae and fungi. Direct bactericidal activity can also be effected by a C-type lectin, but this is a mammalian protein that limits mucosal colonization by Gram-positive bacteria. The presence of two divergent MMBL domains in the novel bacteriocins raises questions about task distribution between modules and the possible role of carbohydrate binding in the specificity of target strain recognition and killing. Notably, bacteriocin activity was also demonstrated for a hybrid MMBL protein with an accessory protease-like domain. This association with one or more additional modules, often with predicted peptide-hydrolysing or -binding activity, suggests that additional bacteriotoxic proteins may be found among the diverse chimaeric MMBL proteins encoded in prokaryotic genomes. A phylogenetic survey of the bacterial MMBL modules reveals a mosaic pattern of strongly diverged sequences, mainly occurring in soil-dwelling and rhizosphere bacteria, which may reflect a trans-kingdom acquisition of the ancestral genes. PMID:23176516

  12. Fluorescent carbohydrate probes for cell lectins

    NASA Astrophysics Data System (ADS)

    Galanina, Oxana; Feofanov, Alexei; Tuzikov, Alexander B.; Rapoport, Evgenia; Crocker, Paul R.; Grichine, Alexei; Egret-Charlier, Marguerite; Vigny, Paul; Le Pendu, Jacques; Bovin, Nicolai V.

    2001-09-01

    Fluorescein labeled carbohydrate (Glyc) probes were synthesized as analytical tools for the study of cellular lectins, i.e. SiaLe x-PAA-flu, Sia 2-PAA-flu, GlcNAc 2-PAA-flu, LacNAc-PAA-flu and a number of similar ones, with PAA a soluble polyacrylamide carrier. The binding of SiaLe x-PAA-flu was assessed using CHO cells transfected with E-selectin, and the binding of Sia 2-PAA-flu was assessed by COS cells transfected with siglec-9. In flow cytometry assays, the fluorescein probes demonstrated a specific binding to the lectin-transfected cells that was inhibited by unlabeled carbohydrate ligands. The intense binding of SiaLe x-PAA- 3H to the E-selectin transfected cells and the lack of binding to both native and permeabilized control cells lead to the conclusion that the polyacrylamide carrier itself and the spacer arm connecting the carbohydrate moiety with PAA did not contribute anymore to the binding. Tumors were obtained from nude mice by injection of CHO E-selectin or mock transfected cells. The fluorescent SiaLe x-PAA-flu probe could bind to the tumor sections from E-selectin positive CHO cells, but not from the control ones. Thus, these probes can be used to reveal specifically the carbohydrate binding sites on cells in culture as well as cells in tissue sections. The use of the confocal spectral imaging technique with Glyc-PAA-flu probes offered the unique possibility to detect lectins in different cells, even when the level of lectin expression was rather low. The confocal mode of spectrum recording provided an analysis of the probe localization with 3D submicron resolution. The spectral analysis (as a constituent part of the confocal spectral imaging technique) enabled interfering signals of the probe and intrinsic cellular fluorescence to be accurately separated, the distribution of the probe to be revealed and its local concentration to be measured.

  13. Herbal complement inhibitors in the treatment of neuroinflammation: future strategy for neuroprotection.

    PubMed

    Kulkarni, Amod P; Kellaway, Laurie A; Kotwal, Girish J

    2005-11-01

    The upregulated complement system plays a damaging role in disorders of the central nervous system (CNS). The classical and alternate pathways are two major pathways activated in neuroinflammatory disorders such as Alzheimer's disease, multiple sclerosis, traumatic brain injury, spinal cord injury, HIV-associated dementia, Parkinson's disease, and mad cow disease. Failure of currently available anti-inflammatory agents, especially cyclooxygenase inhibitors, in offering significant neuroprotection in large epidemiologic clinical trials of CNS disorders suggests an urgent need for the development of new neuroprotective agents. The positive preclinical outcomes in treating CNS disorders by complement regulatory molecules, such as vaccinia virus complement control protein, suggest the possibility of using complement-inhibitory molecules as neuroprotective agents. Several active ingredients of herbal origin are found to have complement-inhibitory activity. These herbal ingredients along with other anti-inflammatory roles might be useful in treating neuroinflammation associated with CNS disorders. Active ingredients of herbal origin with complement inhibitory ingredients are summarized and classified according to their chemical nature and specificity towards the major pathways activating the complement system. The structure activity relationship of some specific examples is also discussed in this report. This information might be helpful in formulating a natural panacea against complement-mediated neuroinflammation. PMID:16387706

  14. Gain-of-function mutations in complement factor B are associated with atypical hemolytic uremic syndrome

    PubMed Central

    de Jorge, Elena Goicoechea; Harris, Claire L.; Esparza-Gordillo, Jorge; Carreras, Luis; Arranz, Elena Aller; Garrido, Cynthia Abarrategui; López-Trascasa, Margarita; Sánchez-Corral, Pilar; Morgan, B. Paul; de Córdoba, Santiago Rodríguez

    2007-01-01

    Hemolytic uremic syndrome (HUS) is an important cause of acute renal failure in children. Mutations in one or more genes encoding complement-regulatory proteins have been reported in approximately one-third of nondiarrheal, atypical HUS (aHUS) patients, suggesting a defect in the protection of cell surfaces against complement activation in susceptible individuals. Here, we identified a subgroup of aHUS patients showing persistent activation of the complement alternative pathway and found within this subgroup two families with mutations in the gene encoding factor B (BF), a zymogen that carries the catalytic site of the complement alternative pathway convertase (C3bBb). Functional analyses demonstrated that F286L and K323E aHUS-associated BF mutations are gain-of-function mutations that result in enhanced formation of the C3bBb convertase or increased resistance to inactivation by complement regulators. These data expand our understanding of the genetic factors conferring predisposition to aHUS, demonstrate the critical role of the alternative complement pathway in the pathogenesis of aHUS, and provide support for the use of complement-inhibition therapies to prevent or reduce tissue damage caused by dysregulated complement activation. PMID:17182750

  15. Complement C3c as a Biomarker in Heart Failure

    PubMed Central

    Frey, A.; Ertl, G.; Angermann, C. E.; Hofmann, U.; Strk, S.; Frantz, S.

    2013-01-01

    Introduction. Experimental data indicates an important role of the innate immune system in cardiac remodeling and heart failure (HF). Complement is a central effector pathway of the innate immune system. Animals lacking parts of the complement system are protected from adverse remodeling. Based on these data, we hypothesized that peripheral complement levels could be a good marker for adverse remodeling and prognosis in patients with HF. Methods and Results. Since complement activation converges on the complement factor C3, we measured serum C3c, a stable C3-conversion product, in 197 patients with stable systolic HF. Subgroups with normal and elevated C3c levels were compared. C3c levels were elevated in 17% of the cohort. Patients with elevated C3c levels exhibited a trend to better survival, slightly higher LVEF, and lower NTpro-BNP values in comparison to patients with normal C3c values. No differences were found regarding NYHA functional class. Significantly more patients with elevated C3c had preexisting diabetes. The prevalence of CAD, arterial hypertension, and atrial fibrillation was not increased in patients with elevated C3c. Conclusion. Elevated C3c levels are associated with less adverse remodeling and improved survival in patients with stable systolic heart failure. PMID:24489446

  16. Collectins and fungal pathogens: roles of surfactant proteins and mannose binding lectin in host resistance.

    PubMed

    Brummer, Elmer; Stevens, David A

    2010-02-01

    Collectins, collagenous carbohydrate-binding proteins (C-type lectins), are recognized as important factors in non-specific innate immune responses to pathogens. By binding surface carbohydrate structures of pathogens, collectins modify the interaction between pathogens and the immune system. We review the structure of the lung surfactant proteins (SP) SP-A and SP-D, and the serum collectins, mannose binding lectins; the binding of these collectins to pathogen associated molecular patterns or ligands on pathogenic fungi; and the effect of collectin binding to opportunistic and primary fungal pathogens on the interaction with host defense cells, which can result in enhancement or inhibition of resistance. The result of collectin binding to opportunistic fungal pathogens (Aspergillus fumigatus, Candida albicans, Cryptococcus neoformans, Pneumocystis) or primary fungal pathogens (Blastomyces dermatitidis, Coccidioides, Histoplasma capsulatum, Paracoccidioides brasiliensis) on interaction with host defense cells relative to complement fixation, phagocytosis, and stimulation of cytokine/chemokine production is reviewed. Increased understanding of these relationships in future will help understand fungal pathogenesis and host defenses against mycoses. PMID:19639514

  17. Unusual complement activation properties of serum immunoglobulins of the pigeon Columba livia.

    PubMed Central

    Koppenheffer, T L; Russell, B A

    1986-01-01

    Complement-mediated lysis of mouse erythrocytes (MRBC) by whole pigeon antisera was found to occur in the presence of magnesium ions alone. The underlying basis for this observation was demonstrated to be the ability of IgM antibodies to activate the alternative pathway of pigeon complement, whereas IgG activates a calcium-dependent pathway possessing an unusually low lytic capacity. In addition to differing from the alternative pathway by requiring both calcium and magnesium ions, the calcium-dependent pathway exhibited higher activity at low temperature and more rapid kinetics of haemolysis. The presence of early acting C1 in pigeon serum was inferred by the selective depletion of calcium-dependent activity which occurred as a result of incubating serum containing only calcium ions with MRBC sensitized with IgG. Under the same conditions, MRBC sensitized with IgM failed to deplete complement activity, indicating that C1 does not participate in complement activation by this isotype. Interestingly, the calcium-dependent pathway detected in pigeon serum appears to more closely resemble the C1-bypass pathway rather than the classical pathway of mammalian complement. PMID:3957409

  18. Complement modulation in solid-organ transplantation.

    PubMed

    Touzot, Maxime; Obada, Erika Nnang; Beaudreuil, Severine; Franois, Hlne; Durrbach, Antoine

    2014-07-01

    The complement system is a major constituent of the innate immune system. It has a critical role in defense against pathogens but dysregulation of complement activation may lead to tissue injury and modulate the adaptive immune response. In organ transplantation, local complement activation is involved in hyper-acute rejection and antibody-mediated rejection. This last decade, interest in complement activation has increased due to new insights into the pathophysiology of antibody-mediated rejection, but also since the availability of news drugs that target terminal complement activation. In this review, we discuss our current understanding of how local complement activation induces acute and chronic graft injury, and review recent advances in clinical trials that block complement activation using the anti-C5 monoclonal antibody, eculizumab. Finally, we discuss how complement-targeted therapy may be integrated into our current immunosuppressive regimen and what type of patient will benefit most from this therapy. PMID:24996770

  19. Genetics Home Reference: Complement factor I deficiency

    MedlinePLUS

    ... have been reported in the medical literature. What genes are related to complement factor I deficiency? Complement ... deficiency is caused by mutations in the CFI gene. This gene provides instructions for making a protein ...

  20. Molecular modeling of lectin-like protein from Acacia farnesiana reveals a possible anti-inflammatory mechanism in Carrageenan-induced inflammation.

    PubMed

    Abrantes, Vanessa Erika Ferreira; Matias da Rocha, Bruno Anderson; Batista da Nbrega, Raphael; Silva-Filho, Jos Caetano; Teixeira, Claudener Souza; Cavada, Benildo Sousa; Gadelha, Carlos Alberto de Almeida; Ferreira, Sergio Henrique; Figueiredo, Jozi Godoy; Santi-Gadelha, Tatiane; Delatorre, Plinio

    2013-01-01

    Acacia farnesiana lectin-like protein (AFAL) is a chitin-binding protein and has been classified as phytohaemagglutinin from Phaseolus vulgaris (PHA). Legume lectins are examples for structural studies, and this family of proteins shows a remarkable conservation in primary, secondary, and tertiary structures. Lectins have ability to reduce the effects of inflammation caused by phlogistic agents, such as carrageenan (CGN). This paper explains the anti-inflammatory activity of AFAL through structural comparison with anti-inflammatory legume lectins. The AFAL model was obtained by molecular modeling and molecular docking with glycan and carrageenan were performed to explain the AFAL structural behavior and biological activity. Pisum sativum lectin was the best template for molecular modeling. The AFAL structure model is folded as a ? sandwich. The model differs from template in loop regions, number of ? strands and carbohydrate-binding site. Carrageenan and glycan bind to different sites on AFAL. The ability of AFAL binding to carrageenan can be explained by absence of the sixth ? -strand (posterior ? sheets) and two ? strands in frontal region. AFAL can inhibit pathway inflammatory process by carrageenan injection by connecting to it and preventing its entry into the cell and triggers the reaction. PMID:24490151

  1. Molecular Modeling of Lectin-Like Protein from Acacia farnesiana Reveals a Possible Anti-Inflammatory Mechanism in Carrageenan-Induced Inflammation

    PubMed Central

    Abrantes, Vanessa Erika Ferreira; Matias da Rocha, Bruno Anderson; Batista da Nbrega, Raphael; Silva-Filho, Jos Caetano; Teixeira, Claudener Souza; Cavada, Benildo Sousa; Gadelha, Carlos Alberto de Almeida; Ferreira, Sergio Henrique; Figueiredo, Jozi Godoy; Santi-Gadelha, Tatiane; Delatorre, Plinio

    2013-01-01

    Acacia farnesiana lectin-like protein (AFAL) is a chitin-binding protein and has been classified as phytohaemagglutinin from Phaseolus vulgaris (PHA). Legume lectins are examples for structural studies, and this family of proteins shows a remarkable conservation in primary, secondary, and tertiary structures. Lectins have ability to reduce the effects of inflammation caused by phlogistic agents, such as carrageenan (CGN). This paper explains the anti-inflammatory activity of AFAL through structural comparison with anti-inflammatory legume lectins. The AFAL model was obtained by molecular modeling and molecular docking with glycan and carrageenan were performed to explain the AFAL structural behavior and biological activity. Pisum sativum lectin was the best template for molecular modeling. The AFAL structure model is folded as a ? sandwich. The model differs from template in loop regions, number of ? strands and carbohydrate-binding site. Carrageenan and glycan bind to different sites on AFAL. The ability of AFAL binding to carrageenan can be explained by absence of the sixth ?-strand (posterior ? sheets) and two ? strands in frontal region. AFAL can inhibit pathway inflammatory process by carrageenan injection by connecting to it and preventing its entry into the cell and triggers the reaction. PMID:24490151

  2. Systemic complement activation in psoriasis vulgaris.

    PubMed

    Fleming, C J; Holme, E R; Mackie, R M

    1996-11-01

    Nineteen patients with psoriasis vulgaris and no other cause for systemic complement activation were studied for evidence of such activation. There was a marked elevation in serum C5b-9 complexes with no other significant complement abnormalities, and no correlation between C5b-9 levels and disease activity. This is the most detailed study of complement in psoriasis yet attempted and confirms that complement activation is a feature of psoriasis vulgaris. PMID:9167335

  3. Lectin domains at the frontiers of plant defense

    PubMed Central

    Lannoo, Nausicaä; Van Damme, Els J. M.

    2014-01-01

    Plants are under constant attack from pathogens and herbivorous insects. To protect and defend themselves, plants evolved a multi-layered surveillance system, known as the innate immune system. Plants sense their encounters upon perception of conserved microbial structures and damage-associated patterns using cell-surface and intracellular immune receptors. Plant lectins and proteins with one or more lectin domains represent a major part of these receptors. The whole group of plant lectins comprises an elaborate collection of proteins capable of recognizing and interacting with specific carbohydrate structures, either originating from the invading organisms or from damaged plant cell wall structures. Due to the vast diversity in protein structures, carbohydrate recognition domains and glycan binding specificities, plant lectins constitute a very diverse protein superfamily. In the last decade, new types of nucleocytoplasmic plant lectins have been identified and characterized, in particular lectins expressed inside the nucleus and the cytoplasm of plant cells often as part of a specific plant response upon exposure to different stress factors or changing environmental conditions. In this review, we provide an overview on plant lectin motifs used in the constant battle against pathogens and predators during plant defenses. PMID:25165467

  4. Extensive amino acid sequence homologies between animal lectins

    SciTech Connect

    Paroutaud, P.; Levi, G.; Teichberg, V.I.; Strosberg, A.D.

    1987-09-01

    The authors have established the amino acid sequence of the ..beta..-D-galactoside binding lectin from the electric eel and the sequences of several peptides from a similar lectin isolated from human placenta. These sequences were compared with the published sequences of peptides derived from the ..beta..-D-galactoside binding lectin from human lung and with sequences deduced from cDNAs assigned to the ..beta..-D-galactoside binding lectins from chicken embryo skin and human hepatomas. Significant homologies were observed. One of the highly conserved regions that contains a tryptophan residue and two glutamic acid resides is probably part of the ..beta..-D-galactoside binding site, which, on the basis of spectroscopic studies of the electric eel lectin, is expected to contain such residues. The similarity of the hydropathy profiles and the predicted secondary structure of the lectins from chicken skin and electric eel, in spite of differences in their amino acid sequences, strongly suggests that these proteins have maintained structural homologies during evolution and together with the other ..beta..-D-galactoside binding lectins were derived form a common ancestor gene.

  5. Cloning and characterization of root-specific barley lectin

    SciTech Connect

    Lerner, D.R.; Raikhel, N.V. )

    1989-09-01

    Cereal lectins are a class of biochemically and antigenically related proteins localized in a tissue-specific manner in embryos and adult plants. To study the specificity of lectin expression, a barley (Hordeum vulgare L.) embryo cDNa library was constructed and a clone (BLc3) for barley lectin was isolated. BLc3 is 972 nucleotides long and includes an open reading frame of 212 amino acids. The deduced amino acid sequence contains a putative signal peptide of 26 amino acid residues followed by a 186 amino acid polypeptide. This polypeptide has 95% sequence identity to the antigenically indistinguishable wheat germ agglutinin isolectin-B (WGA-B) suggesting that BLc3 encodes barley lectin. Further evidence that BLc3 encodes barley lectin was obtained by immunoprecipitation of the in vitro translation products of BLc3 RNA transcripts and barley embryo poly(A{sup +}) RNA. In situ hybridizations with BLc3 showed that barley lectin gene expression is confined to the outermost cell layers of both embryonic and adult root tips. On Northern blots, BLc3 hybridizes to a 1.0 kilobyte mRNA in poly(A{sup +}) RNA from both embryos and root tips. We suggest, on the basis of immunoblot experiments, that barley lectin is synthesized as a glycosylated precursor and processed by removal of a portion of the carboxyl terminus including the single N-linked glycosylation site.

  6. Lectin domains at the frontiers of plant defense.

    PubMed

    Lannoo, Nausicaä; Van Damme, Els J M

    2014-01-01

    Plants are under constant attack from pathogens and herbivorous insects. To protect and defend themselves, plants evolved a multi-layered surveillance system, known as the innate immune system. Plants sense their encounters upon perception of conserved microbial structures and damage-associated patterns using cell-surface and intracellular immune receptors. Plant lectins and proteins with one or more lectin domains represent a major part of these receptors. The whole group of plant lectins comprises an elaborate collection of proteins capable of recognizing and interacting with specific carbohydrate structures, either originating from the invading organisms or from damaged plant cell wall structures. Due to the vast diversity in protein structures, carbohydrate recognition domains and glycan binding specificities, plant lectins constitute a very diverse protein superfamily. In the last decade, new types of nucleocytoplasmic plant lectins have been identified and characterized, in particular lectins expressed inside the nucleus and the cytoplasm of plant cells often as part of a specific plant response upon exposure to different stress factors or changing environmental conditions. In this review, we provide an overview on plant lectin motifs used in the constant battle against pathogens and predators during plant defenses. PMID:25165467

  7. The insecticidal activity of recombinant garlic lectins towards aphids.

    PubMed

    Fitches, Elaine; Wiles, Duncan; Douglas, Angela E; Hinchliffe, Gareth; Audsley, Neil; Gatehouse, John A

    2008-10-01

    The heterodimeric and homodimeric garlic lectins ASAI and ASAII were produced as recombinant proteins in the yeast Pichia pastoris. The proteins were purified as functional dimeric lectins, but underwent post-translational proteolysis. Recombinant ASAII was a single homogenous polypeptide which had undergone C-terminal processing similar to that occurring in planta. The recombinant ASAI was glycosylated and subject to variable and heterogenous proteolysis. Both lectins showed insecticidal effects when fed to pea aphids (Acyrthosiphon pisum) in artificial diet, ASAII being more toxic than ASAI at the same concentration. Acute toxicity (mortality at < or =48 h exposure; similar timescale to starvation) was only apparent at the highest lectin concentrations tested (2.0 mg ml(-)1), but dose-dependent chronic toxicity (mortality at >3d exposure) was observed over the concentration range 0.125-2.0 mg ml(-1). The recombinant lectins caused mortality in both symbiotic and antibiotic-treated aphids, showing that toxicity is not dependent on the presence of the bacterial symbiont (Buchnera aphidicola), or on interaction with symbiont proteins, such as the previously identified lectin "receptor" symbionin. A pull-down assay coupled with peptide mass fingerprinting identified two abundant membrane-associated aphid gut proteins, alanyl aminopeptidase N and sucrase, as "receptors" for lectin binding. PMID:18707000

  8. Complement in the Homeostatic and Ischemic Brain

    PubMed Central

    Alawieh, Ali; Elvington, Andrew; Tomlinson, Stephen

    2015-01-01

    The complement system is a component of the immune system involved in both recognition and response to pathogens, and it is implicated in an increasing number of homeostatic and disease processes. It is well documented that reperfusion of ischemic tissue results in complement activation and an inflammatory response that causes post-reperfusion injury. This occurs following cerebral ischemia and reperfusion and triggers secondary damage that extends beyond the initial infarcted area, an outcome that has rationalized the use of complement inhibitors as candidate therapeutics after stroke. In the central nervous system, however, recent studies have revealed that complement also has essential roles in synaptic pruning, neurogenesis, and neuronal migration. In the context of recovery after stroke, these apparent divergent functions of complement may account for findings that the protective effect of complement inhibition in the acute phase after stroke is not always maintained in the subacute and chronic phases. The development of effective stroke therapies based on modulation of the complement system will require a detailed understanding of complement-dependent processes in both early neurodegenerative events and delayed neuro-reparatory processes. Here, we review the role of complement in normal brain physiology, the events initiating complement activation after cerebral ischemia-reperfusion injury, and the contribution of complement to both injury and recovery. We also discuss how the design of future experiments may better characterize the dual role of complement in recovery after ischemic stroke. PMID:26322048

  9. Upregulation of Glycans Containing 3 Fucose in a Subset of Pancreatic Cancers Uncovered Using Fusion-Tagged Lectins

    PubMed Central

    Singh, Sudhir; Pal, Kuntal; Yadav, Jessica; Tang, Huiyuan; Partyka, Katie; Kletter, Doron; Hsueh, Peter; Ensink, Elliot; Birendra, KC; Hostetter, Galen; Xu, H. Eric; Bern, Marshall; Smith, David F.; Mehta, Anand S.; Brand, Randall; Melcher, Karsten; Haab, Brian B.

    2015-01-01

    The fucose post-translational modification is frequently increased in pancreatic cancer, thus forming the basis for promising biomarkers, but a subset of pancreatic cancer patients does not elevate the known fucose-containing biomarkers. We hypothesized that such patients elevate glycan motifs with fucose in linkages and contexts different from the known fucose-containing biomarkers. We used a database of glycan array data to identify the lectins CCL2 to detect glycan motifs with fucose in a 3 linkage; CGL2 for motifs with fucose in a 2 linkage; and RSL for fucose in all linkages. We used several practical methods to test the lectins and determine the optimal mode of detection, and we then tested whether the lectins detected glycans in pancreatic cancer patients who did not elevate the sialyl-Lewis A glycan, which is upregulated in ~75% of pancreatic adenocarcinomas. Patients who did not upregulate sialyl-Lewis A, which contains fucose in a 4 linkage, tended to upregulate fucose in a 3 linkage, as detected by CCL2, but they did not upregulate total fucose or fucose in a 2 linkage. CCL2 binding was high in cancerous epithelia from pancreatic tumors, including areas negative for sialyl-Lewis A and a related motif containing 3 fucose, sialyl-Lewis X. Thus glycans containing 3 fucose may complement sialyl-Lewis A to contribute to improved detection of pancreatic cancer. Furthermore, the use of panels of recombinant lectins may uncover details about glycosylation that could be important for characterizing and detecting cancer. PMID:25938165

  10. Mesenchymal stem cells are injured by complement after their contact with serum

    PubMed Central

    Li, Yan

    2012-01-01

    Despite the potent immunosuppressive activity that mesenchymal stem cells (MSCs) display in vitro, recent clinical trial results are disappointing, suggesting that MSC viability and/or function are greatly reduced after infusion. In this report, we demonstrated that human MSCs activated complement of the innate immunity after their contact with serum. Although all 3 known intrinsic cell-surface complement regulators were present on MSCs, activated complement overwhelmed the protection of these regulators and resulted in MSCs cytotoxicity and dysfunction. In addition, autologous MSCs suffered less cellular injury than allogeneic MSCs after contacting serum. All 3 complement activation pathways were involved in generating the membrane attack complex to directly injure MSCs. Supplementing an exogenous complement inhibitor, or up-regulating MSC expression levels of CD55, one of the cell-surface complement regulators, helped to reduce the serum-induced MSC cytotoxicity. Finally, adoptively transferred MSCs in complement deficient mice or complement-depleted mice showed reduced cellular injury in vivo compared with those in wild type mice. These results indicate that complement is integrally involved in recognizing and injuring MSCs after their infusion, suggesting that autologous MSCs may have ad-vantages over allogeneic MSCs, and that inhibiting complement activation could be a novel strategy to improve existing MSC-based therapies. PMID:22966167

  11. On the Functional Overlap between Complement and Anti-Microbial Peptides.

    PubMed

    Zimmer, Jana; Hobkirk, James; Mohamed, Fatima; Browning, Michael J; Stover, Cordula M

    2014-01-01

    Intriguingly, activated complement and anti-microbial peptides share certain functionalities; lytic, phagocytic, and chemo-attractant activities and each may, in addition, exert cell instructive roles. Each has been shown to have distinct LPS detoxifying activity and may play a role in the development of endotoxin tolerance. In search of the origin of complement, a functional homolog of complement C3 involved in opsonization has been identified in horseshoe crabs. Horseshoe crabs possess anti-microbial peptides able to bind to acyl chains or phosphate groups/saccharides of endotoxin, LPS. Complement activity as a whole is detectable in marine invertebrates. These are also a source of anti-microbial peptides with potential pharmaceutical applicability. Investigating the locality for the production of complement pathway proteins and their role in modulating cellular immune responses are emerging fields. The significance of local synthesis of complement components is becoming clearer from in vivo studies of parenchymatous disease involving specifically generated, complement-deficient mouse lines. Complement C3 is a central component of complement activation. Its provision by cells of the myeloid lineage varies. Their effector functions in turn are increased in the presence of anti-microbial peptides. This may point to a potentiating range of activities, which should serve the maintenance of health but may also cause disease. Because of the therapeutic implications, this review will consider closely studies dealing with complement activation and anti-microbial peptide activity in acute inflammation (e.g., dialysis-related peritonitis, appendicitis, and ischemia). PMID:25646095

  12. On the Functional Overlap between Complement and Anti-Microbial Peptides

    PubMed Central

    Zimmer, Jana; Hobkirk, James; Mohamed, Fatima; Browning, Michael J.; Stover, Cordula M.

    2015-01-01

    Intriguingly, activated complement and anti-microbial peptides share certain functionalities; lytic, phagocytic, and chemo-attractant activities and each may, in addition, exert cell instructive roles. Each has been shown to have distinct LPS detoxifying activity and may play a role in the development of endotoxin tolerance. In search of the origin of complement, a functional homolog of complement C3 involved in opsonization has been identified in horseshoe crabs. Horseshoe crabs possess anti-microbial peptides able to bind to acyl chains or phosphate groups/saccharides of endotoxin, LPS. Complement activity as a whole is detectable in marine invertebrates. These are also a source of anti-microbial peptides with potential pharmaceutical applicability. Investigating the locality for the production of complement pathway proteins and their role in modulating cellular immune responses are emerging fields. The significance of local synthesis of complement components is becoming clearer from in vivo studies of parenchymatous disease involving specifically generated, complement-deficient mouse lines. Complement C3 is a central component of complement activation. Its provision by cells of the myeloid lineage varies. Their effector functions in turn are increased in the presence of anti-microbial peptides. This may point to a potentiating range of activities, which should serve the maintenance of health but may also cause disease. Because of the therapeutic implications, this review will consider closely studies dealing with complement activation and anti-microbial peptide activity in acute inflammation (e.g., dialysis-related peritonitis, appendicitis, and ischemia). PMID:25646095

  13. Mesenchymal stem cells are injured by complement after their contact with serum.

    PubMed

    Li, Yan; Lin, Feng

    2012-10-25

    Despite the potent immunosuppressive activity that mesenchymal stem cells (MSCs) display in vitro, recent clinical trial results are disappointing, suggesting that MSC viability and/or function are greatly reduced after infusion. In this report, we demonstrated that human MSCs activated complement of the innate immunity after their contact with serum. Although all 3 known intrinsic cell-surface complement regulators were present on MSCs, activated complement overwhelmed the protection of these regulators and resulted in MSCs cytotoxicity and dysfunction. In addition, autologous MSCs suffered less cellular injury than allogeneic MSCs after contacting serum. All 3 complement activation pathways were involved in generating the membrane attack complex to directly injure MSCs. Supplementing an exogenous complement inhibitor, or up-regulating MSC expression levels of CD55, one of the cell-surface complement regulators, helped to reduce the serum-induced MSC cytotoxicity. Finally, adoptively transferred MSCs in complement deficient mice or complement-depleted mice showed reduced cellular injury in vivo compared with those in wild type mice. These results indicate that complement is integrally involved in recognizing and injuring MSCs after their infusion, suggesting that autologous MSCs may have ad-vantages over allogeneic MSCs, and that inhibiting complement activation could be a novel strategy to improve existing MSC-based therapies. PMID:22966167

  14. In vivo biosynthetic studies of the Dolichos biflorus seed lectin

    SciTech Connect

    Quinn, J.M.; Etzler, M.E. )

    1989-12-01

    The in vivo biosynthesis of the Dolichos biflorus seed lectin was studied by pulse-chase labeling experiments using ({sup 35}S)methionine and ({sup 14}C)glucosamine. These studies demonstrate that each of the two mature lectin subunit types are derived by the processing of separate glycosylated precursors. The appearance of the precursor to subunit I before the precursor to subunit II supports the possibility raised by previous studies that both subunit types of this lectin may originate from a single gene product.

  15. Hydroxyl radical scavengers inhibit human lectin-dependent cellular cytotoxicity.

    PubMed Central

    Melinn, M; McLaughlin, H

    1986-01-01

    The role of oxygen-derived free radicals (ODFR) in lectin-dependent cellular cytotoxicity (LDCC) in humans was investigated. The hydroxyl radical traps thiourea, methanol, ethanol and phenol were effective in inhibiting LDCC, as was DABCO, a singlet oxygen quencher. The proposed pathway of hydroxyl radical production in living cells is either an iron catalysed Haber-Weiss reaction or a Fenton reaction. The effect of inhibitors of these pathways was investigated. The superoxide anion scavengers superoxide dismutase, ferricytochrome c and Tiron were without effect. It was shown that Tiron inhibits the lucigenin-amplified chemiluminescence produced by the action of xanthine oxidase, and also the lucigenin-amplified chemiluminescence produced by activated PMN, suggesting that this agent (Tiron) scavenges intracellular superoxide anion. Catalase gave slight inhibition of LDCC only. The ferric iron chelator desferrioxamine gave no protection of the target cells, while the ferrous chelator, 1,10-phenanthroline, inhibited LDCC and partially prevented the detection of hydroxyl radicals generated by the Fe2+-H2O2 system. Cibacron blue, an agent that inhibits NAD(P)H linked enzymes, also inhibited LDCC. The cyclo-oxygenase inhibitors indomethacin and salicylate were without effect, while the lipoxygenase inhibitor nordihydroguaiaretic acid (NDGA) inhibited cytolysis. None of the LDCC inhibitors was cytotoxic to the effector cells or to the target cells, neither did they inhibit lymphocyte-target binding. The findings would suggest that hydroxyl radicals have a role to play in human T-cell mediated cytolysis, either as the active lytic agent or as an epiphenomenon. PMID:3011654

  16. Complementation and Epistasis in Viral Coinfection Dynamics

    PubMed Central

    Gao, Hong; Feldman, Marcus W.

    2009-01-01

    Coinfection in RNA virus populations results in two important phenomena, complementation and recombination. Of the two, complementation has a strong effect on selection against deleterious mutations, as has been confirmed in earlier studies. As complementation delays the purging of less-fit mutations, coinfection may be detrimental to the evolution of a virus population. Here we employ both deterministic modeling and stochastic simulation to explore the mechanisms underlying the interactions between complementation and other evolutionary factors, namely, mutation, selection, and epistasis. We find that strong complementation reduces slightly the overall fitness of a virus population but substantially enhances its diversity and robustness, especially when interacting with selection and epistasis. PMID:19270273

  17. The enzymatic lectin of field bean (Dolichos lablab): salt assisted lectin-sugar interaction.

    PubMed

    Rao, Devavratha H; Vishweshwaraiah, Yashavanth L; Gowda, Lalitha R

    2012-11-01

    Field bean seed contains a Gal/GalNAc lectin (DLL-II) that exhibits associated polyphenol oxidase (PPO) activity and does not bind to its sugar specific affinity matrix. The molecular basis for this lack of binding is not known. The DLL-II gene was therefore cloned and its sequence analyzed. A conserved aromatic residue in the sugar binding site required for a stacking interaction with the apolar backbone of Gal is replaced by His in DLL-II, which explains the lack of binding. However, specific sugar binding is achieved by including (NH?)?SO? in the buffer. Interestingly two other salts of the Hofmeister series, K?HPO? and Na?SO? also assist binding to immobilized galactose. In the presence of (NH?)?SO? the surface hydrophobicity of DLL-II and dissociation constant for 8-anilino 1-naphthalene sulfonic acid were enhanced three fold. This increased surface hydrophobicity in the presence of salt is probably the cause for assisted sugar binding in legume lectins that lack aromatic stacking interactions. Accordingly, two other lectins which lack the conserved aromatic residue show similar salt assisted binding. The salt concentrations required for Gal/GalNAc binding are not physiologically relevant in vivo, suggesting that the role of DLL-II per se in the seed is primarily that of a PPO purportedly for plant defense. PMID:22959225

  18. Overview of Complement Activation and Regulation

    PubMed Central

    Noris, Marina; Remuzzi, Giuseppe

    2013-01-01

    Summary Complement is an important component of the innate immune system that is crucial for defense from microbial infections and for clearance of immune complexes and injured cells. In normal conditions complement is tightly controlled by a number of fluid-phase and cell surface proteins to avoid injury to autologous tissues. When complement is hyperactivated, as occurs in autoimmune diseases or in subjects with dysfunctional regulatory proteins, it drives a severe inflammatory response in numerous organs. The kidney appears to be particularly vulnerable to complement-mediated inflammatory injury. Injury may derive from deposition of circulating active complement fragments in glomeruli, but complement locally produced and activated in the kidney also may have a role. Many kidney disorders have been linked to abnormal complement activation, including immune-complexmediated glomerulonephritis and rare genetic kidney diseases, but also tubulointerstitial injury associated with progressive proteinuric diseases or ischemia-reperfusion. PMID:24161035

  19. The Semantics of Complementation in English: A Cognitive Semantic Account of Two English Complement Constructions

    ERIC Educational Resources Information Center

    Smith, Michael B.

    2009-01-01

    Studies on complementation in English and other languages have traditionally focused on syntactic issues, most notably on the constituent structures of different complement types. As a result, they have neglected the role of meaning in the choice of different complements. This paper investigates the semantics of complementation within the

  20. The Semantics of Complementation in English: A Cognitive Semantic Account of Two English Complement Constructions

    ERIC Educational Resources Information Center

    Smith, Michael B.

    2009-01-01

    Studies on complementation in English and other languages have traditionally focused on syntactic issues, most notably on the constituent structures of different complement types. As a result, they have neglected the role of meaning in the choice of different complements. This paper investigates the semantics of complementation within the…

  1. Correlation between proinflammatory role of a lectin from Typhonium giganteum Engl. and macrophage

    PubMed Central

    Pan, Yaozong; Yu, Hongli; Wu, Hao; Chen, Yeqing; Wang, Kuilong; Liu, Liping; Jin, Yangping; Zhang, Chengchao

    2015-01-01

    Purpose: To analyze the correlation between proinflammatory effects of a lectin from Typhonium giganteum Engl. and macrophage. Methods: T. giganteum lectin (TGL) was extracted from the tuber of T. giganteum and purified, and was then identified by using SDS-PAGE gel electrophoresis in combination with mass spectrometry. The morphologic changes of macrophage after being stimulated by TGL were observed with scanning electron microscopy. The influences of such stimulation on neutrophil migration were evaluated by establishing an in vitro macrophage-neutrophil co-culture migration model. By establishing a rat peritoneal macrophage in vitro cultured model, the effects of TGL stimulation on inflammatory factors TNF-α and IL-1β released by macrophage were analyzed. With p65 as the index, the expressions of the NF-κB signaling pathway in the cytoplasm and nucleus were detected before and after TGL stimulation respectively. Furthermore, we also investigated whether the inhibitor for NF-κB signaling pathway BAY11-7082 can block p65 nuclear translocation. Results: After being stimulated by TGL, macrophage had increased volume, number of pseudopodia and gradually cracked cell membrane, accompanied by evidently induced migration of neutrophils due to released inflammatory factors. As the concentration of TGL varied, NF-κB’s monomer p65 had different expression levels in the cytoplasm and nucleus, while BAY11-7082 can indeed block the nuclear translocation of p65. Conclusions: TGL-induced inflammation was closely related to macrophage mediation. PMID:26617695

  2. Alzheimer's disease and age-related macular degeneration have different genetic models for complement gene variation.

    PubMed

    Proitsi, Petroula; Lupton, Michelle K; Dudbridge, Frank; Tsolaki, Magda; Hamilton, Gillian; Daniilidou, Makrina; Pritchard, Megan; Lord, Kathryn; Martin, Belinda M; Johnson, Janet; Craig, David; Todd, Stephen; McGuinness, Bernadette; Hollingworth, Paul; Harold, Denise; Kloszewska, Iwona; Soininen, Hilkka; Mecocci, Patrizia; Velas, Bruno; Gill, Michael; Lawlor, Brian; Rubinsztein, David C; Brayne, Carol; Passmore, Peter A; Williams, Julie; Lovestone, Simon; Powell, John F

    2012-08-01

    Alzheimer's disease (AD) and age-related macular degeneration (AMD) are both neurodegenerative disorders which share common pathological and biochemical features of the complement pathway. The aim of this study was to investigate whether there is an association between well replicated AMD genetic risk factors and AD. A large cohort of AD (n = 3898) patients and controls were genotyped for single nucleotide polymorphisms (SNPs) in the complement factor H (CFH), the Age-related maculopathy susceptibility protein 2 (ARMS2) the complement component 2 (C2), the complement factor B (CFB), and the complement component 3 (C3) genes. While significant but modest associations were identified between the complement factor H, the age-related maculopathy susceptibility protein 2, and the complement component 3 single nucleotide polymorphisms and AD, these were different in direction or genetic model to that observed in AMD. In addition the multilocus genetic model that predicts around a half of the sibling risk for AMD does not predict risk for AD. Our study provides further support to the hypothesis that while activation of the alternative complement pathway is central to AMD pathogenesis, it is less involved in AD. PMID:22300950

  3. Complement Factor H Binds to Human Serum Apolipoprotein E and Mediates Complement Regulation on High Density Lipoprotein Particles.

    PubMed

    Haapasalo, Karita; van Kessel, Kok; Nissil, Eija; Metso, Jari; Johansson, Tiira; Miettinen, Sini; Varjosalo, Markku; Kirveskari, Juha; Kuusela, Pentti; Chroni, Angelika; Jauhiainen, Matti; van Strijp, Jos; Jokiranta, T Sakari

    2015-11-27

    The alternative pathway of complement is an important part of the innate immunity response against foreign particles invading the human body. To avoid damage to host cells, it needs to be efficiently down-regulated by plasma factor H (FH) as exemplified by various diseases caused by mutations in its domains 19-20 (FH19-20) and 5-7 (FH5-7). These regions are also the main interaction sites for microbial pathogens that bind host FH to evade complement attack. We previously showed that inhibition of FH binding by a recombinant FH5-7 construct impairs survival of FH binding pathogens in human blood. In this study we found that upon exposure to full blood, the addition of FH5-7 reduces survival of, surprisingly, also those microbes that are not able to bind FH. This effect was mediated by inhibition of complement regulation and subsequently enhanced neutrophil phagocytosis by FH5-7. We found that although FH5-7 does not reduce complement regulation in the actual fluid phase of plasma, it reduces regulation on HDL particles in plasma. Using affinity chromatography and mass spectrometry we revealed that FH interacts with serum apolipoprotein E (apoE) via FH5-7 domains. Furthermore, binding of FH5-7 to HDL was dependent on the concentration of apoE on the HDL particles. These findings explain why the addition of FH5-7 to plasma leads to excessive complement activation and phagocytosis of microbes in full anticoagulated blood. In conclusion, our data show how FH interacts with apoE molecules via domains 5-7 and regulates alternative pathway activation on plasma HDL particles. PMID:26468283

  4. Lectins but not antifungal proteins exhibit anti-nematode activity.

    PubMed

    Zhao, S; Guo, Y X; Liu, Q H; Wang, H X; Ng, T B

    2009-09-01

    A variety of lectins and antifungal proteins were tested for toxicity against the plant parasitic nematodes Ditylenchus dipsaci and Heterodera glycines. It was found that lectins from the mushrooms Xylaria hypoxylon, Agrocybe cylindracea and Tricholoma mongolicum (TML-1) were the most potent against D. dipsaci, with EC(50) being 4.7, 9, and 20mg/ml, respectively. Lectins from Pseudostellaria heterophylla, samta tomato, and the mushrooms T. mongolicum (TML-2), Ganoderma lucidum, and Boletus edulis, and antifungal proteins from Ginkgo biloba toward D. dipsaci and pumpkin Cucurbita moschata had much lower anti-nematode potencies and could be considered as inactive for practical purposes. All lectins except that from P.heterophylle were potent against H.glycines. PMID:21784014

  5. Lectins stain cells differentially in the coral, Montipora capitata

    USGS Publications Warehouse

    Work, Thierry M.; Farah, Yael

    2014-01-01

    A limitation in our understanding of coral disease pathology and cellular pathogenesis is a lack of reagents to characterize coral cells. We evaluated the utility of plant lectins to stain tissues of a dominant coral, Montipora capitata, from Hawaii. Of 22 lectins evaluated, nine of these stained structures in the upper or basal body wall of corals. Specific structures revealed by lectins that were not considered distinct or evident on routine hematoxylin and eosin sections of coral tissues included apical and basal granules in gastrodermis and epidermis, cnidoglandular tract and actinopharynx cell surface membranes, capsules of mature holotrichous isorhizas, and perivitelline and periseminal cells. Plant lectins could prove useful to further our understanding of coral physiology, anatomy, cell biology, and disease pathogenesis.

  6. An alternate high yielding purification method for Clitoria ternatea lectin.

    PubMed

    Naeem, Aabgeena; Ahmad, Ejaz; Khan, Rizwan Hasan

    2007-10-01

    In our previous publication we had reported the purification and characterization of Clitoria ternatea agglutinin from its seeds on fetuin CL agarose affinity column, designated CTA [A. Naeem, S. Haque, R.H. Khan. Protein J., 2007]. Since CTA binds beta-d-galactosides, this lectin can be used as valuable tool for glycobiology studies in biomedical and cancer research. So an attempt was made for a high yielding alternative purification method employing the use of asialofetuin CL agarose column for the above-mentioned lectin, designated CTL. The fetuin affinity purified agglutinin was found similar to asialofetuin affinity purified lectin in SDS pattern, HPLC and N-terminal sequence. The content of lectin was found to be 30mg/30g dry weight of pulse. The yield was 2.8% as compared to 0.3% obtained on fetuin column. The number of tryptophan and tyrosine estimated was four and six per subunit. PMID:17590430

  7. A multiplex lectin-channel monitoring method for human serum glycoproteins by quantitative mass spectrometry.

    PubMed

    Ahn, Yeong Hee; Ji, Eun Sun; Shin, Park Min; Kim, Kwang Hoe; Kim, Yong-Sam; Ko, Jeong Heon; Yoo, Jong Shin

    2012-02-01

    A mass profiling method and multiple reaction monitoring (MRM)-based quantitative approach were used to analyze multiple lectin-captured fractions of human serum using different lectins such as aleuria aurantia lectin (AAL), phytohemagglutinin-L(4) (L-PHA), concanavalin A (Con A), and Datura stramonium agglutinin (DSA) to quantitatively monitor protein glycosylation diversity. Each fraction, prepared by multiple lectin-fractionation and tryptic digestion, was analyzed by 1-D LC-MS/MS. Semi-quantitative profiling showed that the list of glycoproteins identified from each lectin-captured fraction is significantly different according to the used lectin. Thus, it was confirmed that the multiplex lectin-channel monitoring (LCM) using multiple lectins is useful for investigating protein glycosylation diversity in a proteome sample. Based on the semi-quantitative mass profiling, target proteins showing lectin-specificity among each lectin-captured fraction were selected and analyzed by the MRM-based method in triplicate using each lectin-captured fraction (average CV 7.9%). The MRM-based analysis for each lectin-captured fraction was similar to those obtained by the profiling experiments. The abundance of each target protein measured varied dramatically, based on the lectin-specificity. The multiplex LCM approach using MRM-based analyses is useful for quantitatively monitoring target protein glycoforms selectively fractionated by multiple lectins. Thus through multiplex LCM rather than single, we could inquire minutely into protein glycosylation states. PMID:22158852

  8. A latex agglutination test for lectin binding.

    PubMed

    Pongor, S; Riedl, Z

    1983-02-15

    Blood group A + H substance purified from hog gastric mucin was fixed to polystyrene particles by physical absorption to produce a lectin-agglutinable latex. Agglutination of this latex by soybean agglutinin, concanavalin A, and wheat germ agglutinin is specifically inhibited by the respective monosaccharide haptens and is not affected by known noninhibitory sugars. The sensitivity of the assay, a function of particle loading and latex concentration, approaches that of the hemagglutination test. NaCl, NaHCO3, NH4CO3, NH4-acetate, and Tris-HCl up to a concentration of 1 M do not interfere with the test. The latex suspension is stable for several months and can be stored in freeze-dried form. PMID:6859532

  9. Bacterial Isolation by Lectin-Modified Microengines

    PubMed Central

    Campuzano, Susana; Orozco, Jahir; Kagan, Daniel; Guix, Maria; Gao, Wei; Sattayasamitsathit, Sirilak; Claussen, Jonathan C.; Merkoi, Arben; Wang, Joseph

    2011-01-01

    New template-based self-propelled gold/nickel/polyaniline/platinum (Au/Ni/PANI/Pt) microtubular engines, functionalized with the Concanavalin A (ConA) lectin bioreceptor, are shown to be extremely useful for the rapid, real-time isolation of Escherichia coli (E. coli) bacteria from fuel-enhanced environmental, food and clinical samples. These multifunctional microtube engines combine the selective capture of E. coli with the uptake of polymeric drug-carrier particles to provide an attractive motion-based theranostics strategy. Triggered release of the captured bacteria is demonstrated by movement through a low-pH glycine-based dissociation solution. The smaller size of the new polymer-metal microengines offers convenient, direct and label-free optical visualization of the captured bacteria and discrimination against non-target cells. PMID:22136558

  10. Complement activation by PEGylated liposomes containing prednisolone.

    PubMed

    van den Hoven, Jolanda M; Nemes, Reka; Metselaar, Josbert M; Nuijen, Bastiaan; Beijnen, Jos H; Storm, Gert; Szebeni, Janos

    2013-05-13

    Infusion of PEGylated liposomes can give rise to hypersensitivity reactions (HSRs) in a relatively large number of patients. Previously it has been shown that these reactions can be caused by activation of the complement (C) system by a negative charge on the anchor molecule of PEG at the liposomal surface. In this study it is tested whether the activation of the C system by PEG-liposomes could be significantly reduced to values comparable to nonreactive liposomal formulations, by changing the PEGylation-profile on the liposomal surface. Therefore, the formation of C activation markers SC5b-9, C3a, C4d and Bb in normal human serum by both prednisolone loaded and empty liposomes with a variation of PEG chain length, PEG surface concentration, PEG anchor molecule and liposomal size was determined using in vitro assays. The tested liposomes caused no or only mild (30%) activation of C except for one formulation wherein the PEG2000 was anchored to cholesterol (CHOL-PEG2000). The latter liposomes caused paralleling rises in SC5b-9 and Bb levels, suggesting excess activation of the alternative pathway. While the relative safety of weak C activator liposomes remains to be confirmed in vivo, the unique, non-charge and non-antibody-mediated direct conversion of C3 by CHOL-PEG2000 liposomes (although argues against the clinical development of these vesicles) opens new opportunities to understand liposomal C activation at the molecular level. PMID:23528740

  11. A Lectin from Dioclea violacea Interacts with Midgut Surface of Lutzomyia migonei, Unlike Its Homologues, Cratylia floribunda Lectin and Canavalia gladiata Lectin

    PubMed Central

    Monteiro Tínel, Juliana Montezuma Barbosa; Benevides, Melina Fechine Costa; Frutuoso, Mércia Sindeaux; Rocha, Camila Farias; Arruda, Francisco Vassiliepe Sousa; Vasconcelos, Mayron Alves; Pereira-Junior, Francisco Nascimento; Cajazeiras, João Batista; do Nascimento, Kyria Santiago; Martins, Jorge Luiz; Teixeira, Edson Holanda; Cavada, Benildo Sousa; dos Santos, Ricardo Pires; Lima Pompeu, Margarida Maria

    2014-01-01

    Leishmaniasis is a vector-borne disease transmitted by phlebotomine sand fly. Susceptibility and refractoriness to Leishmania depend on the outcome of multiple interactions that take place within the sand fly gut. Promastigote attachment to sand fly midgut epithelium is essential to avoid being excreted together with the digested blood meal. Promastigote and gut sand fly surface glycans are important ligands in this attachment. The purpose of the present study was to evaluate the interaction of three lectins isolated from leguminous seeds (Diocleinae subtribe), D-glucose and D-mannose-binding, with glycans on Lutzomyia migonei midgut. To study this interaction the lectins were labeled with FITC and a fluorescence assay was performed. The results showed that only Dioclea violacea lectin (DVL) was able to interact with midgut glycans, unlike Cratylia floribunda lectin (CFL) and Canavalia gladiata lectin (CGL). Furthermore, when DVL was blocked with D-mannose the interaction was inhibited. Differences of spatial arrangement of residues and volume of carbohydrate recognition domain (CRD) may be the cause of the fine specificity of DVL for glycans in the surface on Lu. migonei midgut. The findings in this study showed the presence of glycans in the midgut with glucose/mannose residues in its composition and these residues may be important in interaction between Lu. migonei midgut and Leishmania. PMID:25431778

  12. A lectin from Dioclea violacea Interacts with midgut surface of Lutzomyia migonei, unlike its homologues, Cratylia floribunda lectin and Canavalia gladiata lectin.

    PubMed

    Monteiro Tínel, Juliana Montezuma Barbosa; Benevides, Melina Fechine Costa; Frutuoso, Mércia Sindeaux; Rocha, Camila Farias; Arruda, Francisco Vassiliepe Sousa; Vasconcelos, Mayron Alves; Pereira-Junior, Francisco Nascimento; Cajazeiras, João Batista; do Nascimento, Kyria Santiago; Martins, Jorge Luiz; Teixeira, Edson Holanda; Cavada, Benildo Sousa; dos Santos, Ricardo Pires; Lima Pompeu, Margarida Maria

    2014-01-01

    Leishmaniasis is a vector-borne disease transmitted by phlebotomine sand fly. Susceptibility and refractoriness to Leishmania depend on the outcome of multiple interactions that take place within the sand fly gut. Promastigote attachment to sand fly midgut epithelium is essential to avoid being excreted together with the digested blood meal. Promastigote and gut sand fly surface glycans are important ligands in this attachment. The purpose of the present study was to evaluate the interaction of three lectins isolated from leguminous seeds (Diocleinae subtribe), D-glucose and D-mannose-binding, with glycans on Lutzomyia migonei midgut. To study this interaction the lectins were labeled with FITC and a fluorescence assay was performed. The results showed that only Dioclea violacea lectin (DVL) was able to interact with midgut glycans, unlike Cratylia floribunda lectin (CFL) and Canavalia gladiata lectin (CGL). Furthermore, when DVL was blocked with D-mannose the interaction was inhibited. Differences of spatial arrangement of residues and volume of carbohydrate recognition domain (CRD) may be the cause of the fine specificity of DVL for glycans in the surface on Lu. migonei midgut. The findings in this study showed the presence of glycans in the midgut with glucose/mannose residues in its composition and these residues may be important in interaction between Lu. migonei midgut and Leishmania. PMID:25431778

  13. Complement Activation and Inhibition in Wound Healing

    PubMed Central

    Cazander, Gwendolyn; Jukema, Gerrolt N.; Nibbering, Peter H.

    2012-01-01

    Complement activation is needed to restore tissue injury; however, inappropriate activation of complement, as seen in chronic wounds can cause cell death and enhance inflammation, thus contributing to further injury and impaired wound healing. Therefore, attenuation of complement activation by specific inhibitors is considered as an innovative wound care strategy. Currently, the effects of several complement inhibitors, for example, the C3 inhibitor compstatin and several C1 and C5 inhibitors, are under investigation in patients with complement-mediated diseases. Although (pre)clinical research into the effects of these complement inhibitors on wound healing is limited, available data indicate that reduction of complement activation can improve wound healing. Moreover, medicine may take advantage of safe and effective agents that are produced by various microorganisms, symbionts, for example, medicinal maggots, and plants to attenuate complement activation. To conclude, for the development of new wound care strategies, (pre)clinical studies into the roles of complement and the effects of application of complement inhibitors in wound healing are required. PMID:23346185

  14. The Role of Complement in Tumor Growth

    PubMed Central

    Pio, Ruben; Corrales, Leticia; Lambris, John D.

    2015-01-01

    Complement is a central part of the immune system that has developed as a first defense against non-self cells. Neoplastic transformation is accompanied by an increased capacity of the malignant cells to activate complement. In fact, clinical data demonstrate complement activation in cancer patients. On the basis of the use of protective mechanisms by malignant cells, complement activation has traditionally been considered part of the body's immunosurveillance against cancer. Inhibitory mechanisms of complement activation allow cancer cells to escape from complement-mediated elimination and hamper the clinical efficacy of monoclonal antibody–based cancer immunotherapies. To overcome this limitation, many strategies have been developed with the goal of improving complement-mediated effector mechanisms. However, significant work in recent years has identified new and surprising roles for complement activation within the tumor microenvironment. Recent reports suggest that complement elements can promote tumor growth in the context of chronic inflammation. This chapter reviews the data describing the role of complement activation in cancer immunity, which offers insights that may aid the development of more effective therapeutic approaches to control cancer. PMID:24272362

  15. Annexin A2 Enhances Complement Activation by Inhibiting Factor H.

    PubMed

    Renner, Brandon; Tong, Hua Hua; Laskowski, Jennifer; Jonscher, Karen; Goetz, Lindsey; Woolaver, Rachel; Hannan, Jonathan; Li, Yong Xing; Hourcade, Dennis; Pickering, Matthew C; Holers, V Michael; Thurman, Joshua M

    2016-02-01

    Factor H is a circulating protein that regulates activation of the alternative pathway (AP) of complement. Mutations and genetic variations of factor H are associated with several AP-mediated diseases, highlighting the critical role of factor H in AP regulation. AP-mediated inflammation is typically triggered by illness or tissue injury, however, and tissue injury can trigger AP activation in individuals with fully functional factor H. This suggests that factor H function is affected by local conditions within tissues. We hypothesized that inducible proteins impair the ability of factor H to locally control the AP, thereby increasing AP activation. We used purified murine factor H to immunoprecipitate binding partners from mouse kidneys. Using immunoaffinity liquid chromatography-mass spectrometry, we identified annexin A2 as a factor H binding partner. Further experiments showed that annexin A2 reduces the binding of factor H to cell surfaces. Recombinant annexin A2 impaired complement regulation by factor H and increased complement activation on renal cell surfaces in vitro and in vivo. In a murine model of acute pneumococcal otitis media, the administration of annexin A2 increased AP-mediated bacterial opsonization and clearance. In conclusion, the local production of annexin A2 within tissues suppresses regulation of the AP by factor H. Annexin A2 can contribute to AP-mediated tissue inflammation by locally impairing factor H function, but it can also improve complement-mediated bacterial clearance. PMID:26729803

  16. A novel homodimeric lectin from Astragalus mongholicus with antifungal activity.

    PubMed

    Yan, Qiaojuan; Jiang, Zhengqiang; Yang, Shaoqing; Deng, Wei; Han, Lujia

    2005-10-01

    A novel lectin (AMML) was isolated from a Chinese herb, i.e., the roots of Astragalus mongholicus, using a combination of ammonium sulfate fraction and ion exchange chromatographies. The molecular mass of intact AMML was determined to be 66,396 Da by MALDI-TOF mass spectrometry and 61.8 kDa by gel filtration, respectively. AMML was a dimeric protein composed of two identical subunits each with a molecular mass of 29.6 kDa. The lectin was a glycoprotein with a neutral carbohydrate content of 19.6%. The purified lectin hemagglutinated both rabbit and human erythrocytes, and showed preference for blood types O (native) and AB (trypsin-treated). Among various carbohydrates tested, the lectin was best inhibited by D-galactose and its derivatives with pronounced preference for lactose (3.13 mM). N-terminal amino acid sequence of AMML was determined as ESGINLQGDATLANN. The optimal pH range for lectin activity was between pH 4.5 and 7.5, and the lectin was active up to 65 degrees C. It also exerted antifungal activity against Botrytis cincerea, Fusarium oxysporum, Colletorichum sp., and Drechslera turcia but not against Rhizoctonia solani and Mycosphaerella arachidicola. PMID:16140255

  17. Antifungal activity of lectins against yeast of vaginal secretion

    PubMed Central

    Gomes, Bruno Severo; Siqueira, Ana Beatriz Sotero; de Cssia Carvalho Maia, Rita; Giampaoli, Viviana; Teixeira, Edson Holanda; Arruda, Francisco Vassiliepe Sousa; do Nascimento, Kyria Santiago; de Lima, Adriana Nunes; Souza-Motta, Cristina Maria; Cavada, Benildo Sousa; Porto, Ana Lcia Figueiredo

    2012-01-01

    Lectins are carbohydrate-binding proteins of non-imune origin. This group of proteins is distributed widely in nature and they have been found in viruses, microorganisms, plants and animals. Lectins of plants have been isolated and characterized according to their chemical, physical-chemical, structural and biological properties. Among their biological activities, we can stress its fungicidal action. It has been previously described the effect of the lectins Dviol, DRL, ConBr and LSL obtained from the seeds of leguminous plants on the growth of yeasts isolated from vaginal secretions. In the present work the experiments were carried out in microtiter plates and the results interpreted by both methods: visual observations and a microplate reader at 530nm. The lectin concentrations varied from 0.5 to 256?g/mL, and the inoculum was established between 65-70% of trammitance. All yeast samples isolated from vaginal secretion were evaluated taxonomically, where were observed macroscopic and microscopic characteristics to each species. The LSL lectin did not demonstrate any antifungal activity to any isolate studied. The other lectins DRL, ConBr and DvioL, showed antifungal potential against yeast isolated from vaginal secretion. These findings offering offer a promising field of investigation to develop new therapeutic strategies against vaginal yeast infections, collaborating to improve women's health. PMID:24031889

  18. Assessment of Sauromatum guttatum lectin toxicity against Bactrocera cucurbitae.

    PubMed

    Kaur, Manpreet; Thakur, Kshema; Kamboj, Sukhdev Singh; Kaur, Satwinder; Kaur, Amritpal; Singh, Jatinder

    2015-11-01

    Lectins are proteins that bind specifically to foreign glycans. Due to this binding property, these molecules have potential application as bioinsecticidal tools replacing conventional chemical insecticides. The present study involved purification of phytolectin from the tubers of Sauromatum guttatum by affinity chromatography on asialofetuin-linked silica matrix. The purity of the sample was checked by SDS-PAGE at pH 8.3. Purified lectin was incorporated in the artificial diet of a Dipteran model, Bactrocera cucurbitae at different concentrations (10, 20, 40, 60 and 80 gml(-1)). The lectin significantly affected various developmental parameters that were studied. Percentage pupation and percentage emergence was reduced to 44 % and 7.9%, respectively, at 80 gml(-1) concentration as compared to control (100%). LC50 of Sauromatum guttatum lectin was calculated to be 19.42 gml(-1). Treatment of insect larvae with LC50 of Sauromatum guttatum lectin suppressed the activity of hydrolytic enzymes (esterases and acid phosphatases) and oxidative enzymes (superoxide dismutase and glutathione-S-transferase). Thus, with low LC50 and high mortality (approximately 92% at 80 gml(-1)) of the insect larvae, Sauromatum guttatum lectin offers a possibility to engineer crop plants for improved and safer agriculture. PMID:26688959

  19. Probing lectin-mucin interactions by isothermal titration microcalorimetry.

    PubMed

    Dam, Tarun K; Brewer, C Fred

    2015-01-01

    Isothermal titration microcalorimetry (ITC) can directly determine the thermodynamic binding parameters of biological molecules including affinity constant, binding stoichiometry, and heat of binding (enthalpy) and indirectly the entropy and free energy of binding. ITC has been extensively used to study the binding of lectins to mono- and oligosaccharides, but limited applications to lectin-glycoprotein interactions. Inherent experimental challenges to ITC include sample precipitation during the experiment and relative high amount of sample required, but careful design of experiments can minimize these problems and allow valuable information to be obtained. For example, the thermodynamics of binding of lectins to multivalent globular and linear glycoproteins (mucins) have been described. The results are consistent with a dynamic binding mechanism in which lectins bind and jump from carbohydrate to carbohydrate epitope in these molecules leading to increased affinity. Importantly, the mechanism of binding of lectins to mucins appears similar to that for a variety of protein ligands binding to DNA. Recent results also show that high affinity lectin-mucin cross-linking interactions are driven by favorable entropy of binding that is associated with the bind and jump mechanism. The results suggest that the binding of ligands to biopolymers, in general, may involve a common mechanism that involves enhanced entropic effects that facilitate binding interactions. PMID:25253134

  20. Membrane-assisted online renaturation for automated microfluidic lectin blotting.

    PubMed

    He, Mei; Novak, Jan; Julian, Bruce A; Herr, Amy E

    2011-12-14

    Aberrant glycosylation plays a pivotal role in a diverse set of diseases, including cancer. A microfluidic lectin blotting platform is introduced to enable and expedite the identification of protein glycosylation based on protein size and affinity for specific lectins. The integrated multistage assay eliminates manual intervention steps required for slab-gel lectin blotting, increases total assay throughput, limits reagent and sample consumption, and is completed using one instrument. The assay comprises non-reducing sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) followed by online post-sizing SDS filtration and lectin-based affinity blotting. Important functionality is conferred through both device and assay advances that enable integration of nanoporous membranes flanking a central microchamber to create sub-nanoliter volume compartments that trap SDS-protein complexes and allow electrophoretic SDS removal with buffer exchange. Recapitulation of protein binding for lectin was optimized through quantitative assessment of SDS-treated green fluorescent protein. Immunoglobulin A1 aberrantly glycosylated with galactose-deficient O-glycans was probed in ~6 min using ~3 ?L of sample. This new microfluidic lectin blotting platform provides a rapid and automated assay for the assessment of aberrant glycosylation. PMID:22070432

  1. Microbe-specific C3b deposition in the horseshoe crab complement system in a C2/factor B-dependent or -independent manner.

    PubMed

    Tagawa, Keisuke; Yoshihara, Toyoki; Shibata, Toshio; Kitazaki, Kazuki; Endo, Yuichi; Fujita, Teizo; Koshiba, Takumi; Kawabata, Shun-ichiro

    2012-01-01

    Complement C3 plays an essential role in the opsonization of pathogens in the mammalian complement system, whereas the molecular mechanism underlying C3 activation in invertebrates remains unknown. To understand the molecular mechanism of C3b deposition on microbes, we characterized two types of C2/factor B homologs (designated TtC2/Bf-1 and TtC2/Bf-2) identified from the horseshoe crab Tachypleus tridentatus. Although the domain architectures of TtC2/Bf-1 and TtC2/Bf-2 were identical to those of mammalian homologs, they contained five-repeated and seven-repeated complement control protein domains at their N-terminal regions, respectively. TtC2/Bf-1 and TtC2/Bf-2 were synthesized and glycosylated in hemocytes and secreted to hemolymph plasma, which existed in a complex with C3 (TtC3), and their activation by microbes was absolutely Mg(2+)-dependent. Flow cytometric analysis revealed that TtC3b deposition was Mg(2+)-dependent on Gram-positive bacteria or fungi, but not on Gram-negative bacteria. Moreover, this analysis demonstrated that Ca(2+)-dependent lectins (C-reactive protein-1 and tachylectin-5A) were required for TtC3b deposition on Gram-positive bacteria, and that a Ca(2+)-independent lectin (Tachypleus plasma lectin-1) was definitely indispensable for TtC3b deposition on fungi. In contrast, a horseshoe crab lipopolysaccharide-sensitive protease factor C was necessary and sufficient to deposit TtC3b on Gram-negative bacteria. We conclude that plasma lectins and factor C play key roles in microbe-specific TtC3b deposition in a C2/factor B-dependent or -independent manner. PMID:22611464

  2. Complement Interaction with Trypanosomatid Promastigotes in Normal Human Serum

    PubMed Central

    Domínguez, Mercedes; Moreno, Inmaculada; López-Trascasa, Margarita; Toraño, Alfredo

    2002-01-01

    In normal human serum (NHS), axenic promastigotes of Crithidia, Phytomonas, and Leishmania trigger complement activation, and from 1.2 to 1.8 × 105 C3 molecules are deposited per promastigote within 2.5 min. In Leishmania, promastigote C3 binding capacity remains constant during in vitro metacyclogenesis. C3 deposition on promastigotes activated through the classical complement pathway reaches a 50% maximum after ∼50 s, and represents >85% of total C3 bound. In C1q- and C2-deficient human sera, promastigotes cannot activate the classical pathway (CP) unless purified C1q or C2 factors, respectively, are supplemented, demonstrating a requirement for CP factor in promastigote C3 opsonization. NHS depleted of natural anti-Leishmania antibodies cannot trigger promastigote CP activation, but IgM addition restores C3 binding. Furthermore, Leishmania binds natural antibodies in ethylenediaminetetracetic acid (EDTA)-treated NHS; after EDTA removal, promastigote-bound IgM triggers C3 deposition in natural antibody-depleted NHS. Serum collectins and pentraxins thus do not participate significantly in NHS promastigote C3 opsonization. Real-time kinetic analysis of promastigote CP-mediated lysis indicates that between 85–95% of parasites are killed within 2.5 min of serum contact. These data indicate that successful Leishmania infection in man must immediately follow promastigote transmission, and that Leishmania evasion strategies are shaped by the selective pressure exerted by complement. PMID:11854358

  3. The role of complement activation in atherogenesis: the first 40years.

    PubMed

    Vlaicu, Sonia I; Tatomir, Alexandru; Rus, Violeta; Mekala, Armugam P; Mircea, Petru A; Niculescu, Florin; Rus, Horea

    2016-02-01

    The pathogenesis of atherosclerotic inflammation is a multi-step process defined by the interweaving of excess modified lipid particles, monocyte-macrophages populations, and innate immune and adaptive immunity effectors. A part of innate immunity, the complement system, is an important player in the induction and progression of atherosclerosis. The accumulation of either oxidized or enzymatically modified LDL-bound to C-reactive protein or not-prompts complement activation leading to the assembly of the terminal complement C5b-9 complex in the atherosclerotic lesion. The sublytic C5b-9 assembly leads to the activation and proliferation of smooth muscle and endothelial cells, accompanied by the release of various chemotactic, pro-adhesion, and procoagulant cytokines from these cells. Response gene to complement (RGC)-32, an essential effector of the terminal complement complex C5b-9, also affects atherogenesis, propelling vascular smooth muscle cell proliferation and migration, stimulating endothelial proliferation, and promoting vascular lesion formation. A substantial amount of experimental work has suggested a role for the complement system activation during atherosclerotic plaque formation, with the proximal classical complement pathway seemingly having a protective effect and terminal complement contributing to accelerated atherogenesis. All these data suggest that complement plays an important role in atherogenesis. PMID:26091721

  4. Regulation of complement and modulation of its activity in monoclonal antibody therapy of cancer

    PubMed Central

    Meyer, Saskia; Leusen, Jeanette HW; Boross, Peter

    2014-01-01

    The complement system is a powerful tool of the innate immune system to eradicate pathogens. Both in vitro and in vivo evidence indicates that therapeutic anti-tumor monoclonal antibodies (mAbs) can activate the complement system by the classical pathway. However, the contribution of complement to the efficacy of mAbs is still debated, mainly due to the lack of convincing data in patients. A beneficial role for complement during mAb therapy is supported by the fact that cancer cells often upregulate complement-regulatory proteins (CRPs). Polymorphisms in various CRPs were previously associated with complement-mediated disorders. In this review the role of complement in anti-tumor mAb therapy will be discussed with special emphasis on strategies aiming at modifying complement activity. In the future, clinical efficacy of mAbs with enhanced effector functions together with comprehensive analysis of polymorphisms in CRPs in mAb-treated patients will further clarify the role of complement in mAb therapy. PMID:25517299

  5. Role of Complement in Autoimmune Hemolytic Anemia

    PubMed Central

    Berentsen, Sigbjørn

    2015-01-01

    Summary The classification of autoimmune hemolytic anemias and the complement system are reviewed. In autoimmune hemolytic anemia of the warm antibody type, complement-mediated cell lysis is clinically relevant in a proportion of the patients but is hardly essential for hemolysis in most patients. Cold antibody-mediated autoimmune hemolytic anemias (primary cold agglutinin disease, secondary cold agglutinin syndrome and paroxysmal cold hemoglobinuria) are entirely complement-mediated disorders. In cold agglutinin disease, efficient therapies have been developed in order to target the pathogenic B-cell clone, but complement modulation remains promising in some clinical situations. No established therapy exists for secondary cold agglutinin syndrome and paroxysmal cold hemoglobinuria, and the possibility of therapeutic complement inhibition is interesting. Currently, complement modulation is not clinically documented in any autoimmune hemolytic anemia. The most relevant candidate drugs and possible target levels of action are discussed. PMID:26696798

  6. Shiga toxin activates complement and binds factor H: evidence for an active role of complement in hemolytic uremic syndrome.

    PubMed

    Orth, Dorothea; Khan, Abdul Basit; Naim, Asma; Grif, Katharina; Brockmeyer, Jens; Karch, Helge; Joannidis, Michael; Clark, Simon J; Day, Anthony J; Fidanzi, Sonja; Stoiber, Heribert; Dierich, Manfred P; Zimmerhackl, Lothar B; Wrzner, Reinhard

    2009-05-15

    Infections with enterohemorrhagic Escherichia coli (EHEC) are a major cause of hemolytic uremic syndrome (HUS). Shiga toxins (Stxs), especially Stx2, are believed to represent major virulence factors of EHEC, contributing to HUS pathogenesis. Beside EHEC-associated HUS, there are hereditary atypical forms of HUS, which are mostly caused by mutations of complement regulators. The aim of the present study was to investigate whether or not complement is also involved in the pathogenesis of EHEC-induced typical HUS, by being activated either directly or indirectly by involvement of its inhibitors. Purified Stx2 markedly activated complement via the alternative pathway and was found to bind to factor H (FH), however, only when it was active. No apparent cleavage or destruction of FH was visible, and cofactor activity in fluid phase was unaffected, but clearly delayed for surface-attached FH, where it is essential for host cell protection. Binding studies using FH constructs revealed that Stx2 binds to short consensus repeats (SCRs) 6-8 and SCRs18-20, but not to SCRs16-17, i.e., to regions involved in the surface recognition function of FH. In conclusion, complement, and in particular FH, not only plays an important role in atypical HUS, but most probably also in EHEC-induced HUS. PMID:19414792

  7. Pasteurella pneumotropica Evades the Human Complement System by Acquisition of the Complement Regulators Factor H and C4BP

    PubMed Central

    Sahagún-Ruiz, Alfredo; Granados Martinez, Adriana Patricia; Breda, Leandro Carvalho Dantas; Fraga, Tatiana Rodrigues; Castiblanco Valencia, Mónica Marcela; Barbosa, Angela Silva; Isaac, Lourdes

    2014-01-01

    Pasteurella pneumotropica is an opportunist Gram negative bacterium responsible for rodent pasteurellosis that affects upper respiratory, reproductive and digestive tracts of mammals. In animal care facilities the presence of P. pneumotropica causes severe to lethal infection in immunodeficient mice, being also a potential source for human contamination. Indeed, occupational exposure is one of the main causes of human infection by P. pneumotropica. The clinical presentation of the disease includes subcutaneous abscesses, respiratory tract colonization and systemic infections. Given the ability of P. pneumotropica to fully disseminate in the organism, it is quite relevant to study the role of the complement system to control the infection as well as the possible evasion mechanisms involved in bacterial survival. Here, we show for the first time that P. pneumotropica is able to survive the bactericidal activity of the human complement system. We observed that host regulatory complement C4BP and Factor H bind to the surface of P. pneumotropica, controlling the activation pathways regulating the formation and maintenance of C3-convertases. These results show that P. pneumotropica has evolved mechanisms to evade the human complement system that may increase the efficiency by which this pathogen is able to gain access to and colonize inner tissues where it may cause severe infections. PMID:25347183

  8. Complement Factors and their G Protein Coupled Receptors in Neuroprotection and Neurodegeneration

    PubMed Central

    Yanamadala, Vijay; Friedlander, Robert M.

    2010-01-01

    Acute neurodegeneration is associated with high morbidity and mortality, and there are few effective treatments. Inflammation is central to the process of neuronal death, and within this field, the roles of the complement cascade have proven to be complex. The complement cascade is involved in triggering cell death and recruiting inflammatory cells to sites of inflammation, including the brain. However, complement might also have important neuroprotective roles that are only now coming to light. Recent evidence suggests that targeted activation of complement may be a potential avenue for treatment of stroke and other acute neurodegenerative diseases. Herein, we describe these novel neuroprotective roles of the complement cascade, focusing on signaling pathways that may be therapeutically targetable in acute neuronal injury. PMID:20116331

  9. Purification and characterization of two mannan-binding lectins from mouse serum.

    PubMed

    Hansen, S; Thiel, S; Willis, A; Holmskov, U; Jensenius, J C

    2000-03-01

    Mannan-binding lectin (MBL) is a serum protein that activates the complement system after binding to glycoconjugates found on the surface of microorganisms. By molecular cloning two forms of MBL have been identified in the mouse (mMBL-A and mMBL-C), but only mMBL-A has been purified and characterized at the protein level. MBL-C has been termed the liver form of MBL. The present report describes the purification and characterization of mMBL-A and mMBL-C from serum. The two forms of mMBL could be separated both by ion-exchange and carbohydrate-affinity chromatography. The initial identification by immunochemical technique was confirmed by N-terminal amino-acid sequencing. Both proteins give bands corresponding to polypeptide chains of 28 kDa on SDS-PAGE in the reduced state, but mMBL-A migrated more rapidly than mMBL-C in acid/urea-PAGE, in accordance with the calculated pIs. Both forms mediated activation of complement component C4 in mannan-coated microtiter wells. MBL-A showed a higher affinity for d -glucose and alpha-methyl-d -glucose then did MBL-C. Serum concentrations of mMBL-A in laboratory strains and wild mice were found to vary from 5 to 80 microg/ml, with wild mice tending to show higher levels than laboratory strains. PMID:10679100

  10. Complement Factor H and Simian Virus 40 bind the GM1 ganglioside in distinct conformations.

    PubMed

    Blaum, Bärbel S; Frank, Martin; Walker, Ross C; Neu, Ursula; Stehle, Thilo

    2016-05-01

    Mammalian cell surfaces are decorated with a variety of glycan chains that orchestrate development and defense and are exploited by pathogens for cellular attachment and entry. While glycosidic linkages are, in principle, flexible, the conformational space that a given glycan can sample is subject to spatial and electrostatic restrictions imposed by its overall chemical structure. Here, we show how the glycan moiety of the GM1 ganglioside, a branched, monosialylated pentasaccharide that serves as a ligand for various proteins, undergoes differential conformational selection in its interactions with different lectins. Using STD NMR and X-ray crystallography, we found that the innate immune regulator complement Factor H (FH) binds a previously not reported GM1 conformation that is not compatible with the GM1-binding sites of other structurally characterized GM1-binding lectins such as the Simian Virus 40 (SV40) capsid. Molecular dynamics simulations of the free glycan in explicit solvent on the 10 μs timescale reveal that the FH-bound conformation nevertheless corresponds to a minimum in the Gibbs free energy plot. In contrast to the GM1 conformation recognized by SV40, the FH-bound GM1 conformation is associated with poor NOE restraints, explaining how it escaped(1)H-(1)H NOE-restrained modeling in the past and highlighting the necessity for ensemble representations of glycan structures. PMID:26715202

  11. Systematic assessment of the influence of complement gene polymorphisms on kidney transplant outcome.

    PubMed

    Ermini, Luca; Weale, Michael E; Brown, Katherine M; Mesa, Irene Rebollo; Howell, W Martin; Vaughan, Robert; Chowdhury, Paramit; Sacks, Steven H; Sheerin, Neil S

    2016-04-01

    The importance of the innate immune system, including complement, in causing transplant injury and augmenting adaptive immune responses is increasingly recognized. Therefore variability in graft outcome may in part be due to genetic polymorphism in genes encoding proteins of the immune system. This study assessed the relationship between single nucleotide polymorphisms (SNPs) in complement genes and outcome after transplantation. Analysis was performed on two patient cohorts of 650 and 520 transplant recipients. 505 tagged SNPs in 47 genes were typed in both donor and recipient. The relationships between SNPs and graft survival, serum creatinine, delayed graft function and acute rejection were analyzed. One recipient SNP in the gene encoding mannose binding lectin was associated with graft outcome after correction for analysis of multiple SNPs (p=6.41×10(-5)). When further correction was applied to account for analysis of the effect of SNPs in both donor and recipient this lost significance. Despite association p values of <0.001 no SNP was significantly associated with clinical phenotypes after Bonferroni correction. In conclusion, the variability seen in transplant outcome in this patient cohort cannot be explained by variation in complement genes. If causal genetic effects exist in these genes, they are too small to be detected by this study. PMID:26797657

  12. The Pivotal Role of the Complement System in Aging and Age-related Macular Degeneration: Hypothesis Re-visited

    PubMed Central

    Anderson, Don H.; Radeke, Monte J.; Gallo, Natasha B.; Chapin, Ethan A.; Johnson, Patrick T.; Curletti, Christy R.; Hancox, Lisa S.; Hu, Jane; Ebright, Jessica N.; Malek, Goldis; Hauser, Michael A.; Rickman, Catherine Bowes; Bok, Dean; Hageman, Gregory S.; Johnson, Lincoln V.

    2009-01-01

    During the past ten years, dramatic advances have been made in unraveling the biological bases of age-related macular degeneration (AMD), the most common cause of irreversible blindness in western populations. In that timeframe, two distinct lines of evidence emerged which implicated chronic local inflammation and activation of the complement cascade in AMD pathogenesis. First, a number of complement system proteins, complement activators, and complement regulatory proteins were identified as molecular constituents of drusen, the hallmark extracellular deposits associated with early AMD. Subsequently, genetic studies revealed highly significant statistical associations between AMD and variants of several complement pathway-associated genes including: Complement factor H (CFH), complement factor H-related 1 and 3 (CFHR1 and CFHR3), complement factor B (CFB), complement component 2 (C2), and complement component 3 (C3). In this article, we revisit our original hypothesis that chronic local inflammatory and immune-mediated events at the level of Bruchs membrane play critical roles in drusen biogenesis and, by extension, in the pathobiology of AMD. Secondly, we report the results of a new screening for additional AMD-associated polymorphisms in a battery of 63 complement-related genes. Third, we identify and characterize the local complement system in the RPE-choroid complex -- thus adding a new dimension of biological complexity to the role of the complement system in ocular aging and AMD. Finally, we evaluate the most salient, recent evidence that bears directly on the role of complement in AMD pathogenesis and progression. Collectively, these recent findings strongly re-affirm the importance of the complement system in AMD. They lay the groundwork for further studies that may lead to the identification of a transcriptional disease signature of AMD, and hasten the development of new therapeutic approaches that will restore the complement-modulating activity that appears to be compromised in genetically susceptible individuals. PMID:19961953

  13. Complement C3 and C5 Deficiency Affects Fracture Healing

    PubMed Central

    Ehrnthaller, Christian; Huber-Lang, Markus; Nilsson, Per; Bindl, Ronny; Redeker, Simon; Recknagel, Stefan; Rapp, Anna; Mollnes, Tom; Amling, Michael; Gebhard, Florian; Ignatius, Anita

    2013-01-01

    There is increasing evidence that complement may play a role in bone development. Our previous studies demonstrated that the key complement receptor C5aR was strongly expressed in the fracture callus not only by immune cells but also by bone cells and chondroblasts, indicating a function in bone repair. To further elucidate the role of complement in bone healing, this study investigated fracture healing in mice in the absence of the key complement molecules C3 and C5. C3-/- and C5-/- as well as the corresponding wildtype mice received a standardized femur osteotomy, which was stabilized using an external fixator. Fracture healing was investigated after 7 and 21 days using histological, micro-computed tomography and biomechanical measurements. In the early phase of fracture healing, reduced callus area (C3-/-: -25%, p=0.02; C5-/-: -20% p=0.052) and newly formed bone (C3-/-: -38%, p=0.01; C5-/-: -52%, p=0.009) was found in both C3- and C5-deficient mice. After 21 days, healing was successful in the absence of C3, whereas in C5-deficient mice fracture repair was significantly reduced, which was confirmed by a reduced bending stiffness (-45%; p=0.029) and a smaller callus volume (-17%; p=0.039). We further demonstrated that C5a was activated in C3-/- mice, suggesting cleavage via extrinsic pathways. Our results suggest that the activation of the terminal complement cascade in particular may be crucial for successful fracture healing. PMID:24260573

  14. Lectin-like molecules in transcriptome of Littorina littorea hemocytes.

    PubMed

    Gorbushin, Alexander M; Borisova, Elena A

    2015-01-01

    The common periwinkle Littorina littorea was introduced in the list of models for comparative immunobiology as a representative of phylogenetically important taxon Caenogastropoda. Using Illumina sequencing technology, we de novo assembled the transcriptome of Littorina littorea hemocytes from 182 million mRNA-Seq pair-end 100?bp reads into a total of 15,526 contigs clustered in 4472 unigenes. The transcriptome profile was analyzed for presence of carbohydrate-binding molecules in a variety of architectural contexts. Hemocytes' repertoire of lectin-like proteins bearing conserved carbohydrate-recognition domains (CRDs) is highly diversified, including 11 of 15 lectin families earlier described in animals, as well as the novel members of lectin family found for the first time in mollusc species. The new molluscan lineage-specific domain combinations were confirmed by cloning and sequencing, including the fuco-lectin related molecules (FLReMs) composed of N-terminal region with no sequence homology to any known protein, a middle Fucolectin Tachylectin-4 Pentaxrin (FTP) domain, and a C-terminal epidermal growth factor (EGF) repeat region. The repertoire of lectin-like molecules is discussed in terms of their potential participation in the receptor phase of immune response. In total, immune-associated functions may be attributed to 70 transcripts belonging to 6 lectin families. These lectin-like genes show low overlap between species of invertebrates, suggesting relatively rapid evolution of immune-associated genes in the group. The repertoire provides valuable candidates for further characterization of the gene functions in mollusc immunity. PMID:25451301

  15. Studies on Galalpha3-binding proteins: comparison of the glycosphingolipid binding specificities of Marasmius oreades lectin and Euonymus europaeus lectin.

    PubMed

    Teneberg, Susann; Alsn, Bjorn; Angstrm, Jonas; Winter, Harry C; Goldstein, Irwin J

    2003-06-01

    The carbohydrate binding preferences of the Galalpha3Galbeta4 GlcNAc-binding lectins from Marasmius oreades and Euonymus europaeus were examined by binding to glycosphingolipids on thin-layer chromatograms and in microtiter wells. The M. oreades lectin bound to Galalpha3-terminated glycosphingolipids with a preference for type 2 chains. The B6 type 2 glycosphingolipid (Galalpha3[Fucalpha2]Galbeta4GlcNAcbeta3Galbeta4Glcbeta1Cer) was preferred over the B5 glycosphingolipid (Galalpha3Galbeta4GlcNAcbeta3Galbeta4Glcbeta1Cer), suggesting that the alpha2-linked Fuc is accommodated in the carbohydrate binding site, providing additional interactions. The lectin from E. europaeus had broader binding specificity. The B6 type 2 glycosphingolipid was the best ligand also for this lectin, but binding to the B6 type 1 glycosphingolipid (Galalpha3[Fucalpha2]Galbeta3GlcNAcbeta3Galbeta4Glcbeta1Cer) was also obtained. Furthermore, the H5 type 2 glycosphingolipid (Fucalpha2Galbeta4GlcNAcbeta3Galbeta4Glcbeta1Cer), devoid of a terminal alpha3-linked Gal, was preferred over the the B5 glycosphingolipid, demonstrating a significant contribution to the binding affinity by the alpha2-linked Fuc. The more tolerant nature of the lectin from E. europaeus was also demonstrated by the binding of this lectin, but not the M. oreades lectin, to the x2 glycosphingolipid (GalNAcbeta3Galbeta4GlcNAcbeta3Galbeta4Glcbeta1Cer) and GlcNAcbeta3Galbeta4GlcNAcbeta3Galbeta4Glcbeta1Cer. The A6 type 2 glycosphingolipid (GalNAcalpha3[Fucalpha2]Galbeta4GlcNAcbeta3Galbeta4Glcbeta1Cer) and GalNAcalpha3Galbeta4GlcNAcbeta3Galbeta4Glcbeta1-Cer were not recognized by the lectins despite the interaction with B6 type 2 glycosphingolipid and the B5 glycosphingolipid. These observations are explained by the absolute requirement of a free hydroxyl in the 2-position of Galalpha3 and that the E. europaea lectin can accommodate a GlcNAc acetamido moiety close to this position by reorienting the terminal sugar, whereas the M. oreades lectin cannot. PMID:12626398

  16. Learnings from over 25 years of PNH experience: the era of targeted complement inhibition.

    PubMed

    Heitlinger, Ellen

    2013-12-01

    Paroxysmal nocturnal haemoglobinuria (PNH) is a progressive and life-threatening disease that causes thrombosis, end organ damage and impaired quality of life. Chronic uncontrolled complement activation leads to chronic haemolysis, causing progressive morbidities and early mortality. Hence, early diagnosis is essential for improved patient management and prognosis. Eculizumab (SOLIRIS) specifically inhibits chronic, uncontrolled complement activation and is the first-in-class, humanised, monoclonal antibody targeting C5 within the terminal complement pathway. Eculizumab is the first and only approved treatment for PNH in adults and children. PMID:24331206

  17. Anti-complement activity of the Ixodes scapularis salivary protein Salp20.

    PubMed

    Hourcade, Dennis E; Akk, Antonina M; Mitchell, Lynne M; Zhou, Hui-Fang; Hauhart, Richard; Pham, Christine T N

    2016-01-01

    Complement, a major component of innate immunity, presents a rapid and robust defense of the intravascular space. While regulatory proteins protect host cells from complement attack, when these measures fail, unrestrained complement activation may trigger self-tissue injury, leading to pathologic conditions. Of the three complement activation pathways, the alternative pathway (AP) in particular has been implicated in numerous disease and injury states. Consequently, the AP components represent attractive targets for therapeutic intervention. The common hard-bodied ticks from the family Ixodidae derive nourishment from the blood of their mammalian hosts. During its blood meal the tick is exposed to host immune effectors, including the complement system. In defense, the tick produces salivary proteins that can inhibit host immune functions. The Salp20 salivary protein of Ixodes scapularis inhibits the host AP pathway by binding properdin and dissociating C3bBbP, the active C3 convertase. In these studies we examined Salp20 activity in various complement-mediated pathologies. Our results indicate that Salp20 can inhibit AP-dependent pathogenesis in the mouse. Its efficacy may be part in due to synergic effects it provides with the endogenous AP regulator, factor H. While Salp20 itself would be expected to be highly immunogenic and therefore inappropriate for therapeutic use, its emergence speaks for the potential development of a non-immunogenic Salp20 mimic that replicates its anti-properdin activity. PMID:26675068

  18. Complement receptor of the immunoglobulin superfamily reduces murine lupus nephritis and cutaneous disease.

    PubMed

    Lieberman, Linda A; Mizui, Masayuki; Nalbandian, Angle; Boss, Robin; Crispn, Jos C; Tsokos, George C

    2015-10-01

    Complement activation takes place in autoimmune diseases and accounts for tissue inflammation. Previously, complement inhibition has been considered for the treatment of SLE. Complement receptor of the immunoglobulin superfamily (CRIg) is a selective inhibitor of the alternative pathway of complement and a soluble form reverses established inflammation and bone destruction in experimental autoimmune arthritis. We asked whether specific inhibition of the alternative pathway could inhibit autoimmunity and/or organ damage in lupus-prone mice. Accordingly, we treated lupus-prone MRL/lpr mice with a soluble form of CRIg (CRIg-Fc) and we found that it significantly diminished skin lesions, proteinuria and pyuria, and kidney pathology. Interestingly, serum levels of anti-DNA antibodies were not affected despite the fact that serum complement 3 (C3) levels increased significantly. Immunofluorescent staining of kidney tissues revealed a reduction in staining intensity for C3, IgG, and the macrophage marker Mac-2. Thus our data show that inhibition of the alternative pathway of complement controls skin and kidney inflammation even in the absence of an effect on the production of autoantibodies. We propose that CRIg should be considered for clinical trials in patients with systemic lupus erythematosus. PMID:25988858

  19. Conglutinin exhibits a complement-dependent enhancement of the respiratory burst of phagocytes stimulated by E. coli.

    PubMed Central

    Friis, P; Svehag, S E; Andersen, O; Gahrn-Hansen, B; Leslie, R G

    1991-01-01

    Conglutinin is a mammalian C-type lectin which shows anti-bacterial activity when tested in vivo and in vitro. This study concerns the effect of conglutinin on the respiratory burst of murine spleen cells, using a chemiluminescence assay for measurement of generated reactive oxygen metabolites. Conglutinin enhances, in a dose-dependent manner, the respiratory burst of spleen cells stimulated with serum-opsonized Escherichia coli. The enhancement was only demonstrable in the presence of a functional complement system. The conglutinin-mediated enhancement of the respiratory burst was inhibited in the presence of a N-acetyl-D-glucosamine, D-mannose and N-acetyl-D-mannosamine, monosaccharides reported to inhibit conglutinin-binding to zymosan and the complement factor iC3b. On the other hand, N-acetyl-D-galactosamine was non-inhibitory. PMID:1783427

  20. Tissue binding pattern of plant lectins in benign and malignant lesions of thyroid.

    PubMed

    Vijayakumar, T; Augustine, J; Mathew, L; Aleykutty, M A; Nair, M B; Remani, P; Nair, M K

    1992-01-01

    N-acetyl D-galactosamine specific lectins were isolated from the seeds of Jack Fruit (Artocarpus integrifolia) and Winged bean (Psophocarpus tetragonolobus) and D-galactose specific lectin was isolated from peanut (Arachis hypogaea). These lectins were conjugated to Horse Radish Peroxidase (HRP) and were used to study the lectin binding properties of benign and malignant lesions of the thyroid. For comparison of the results 10 normal fresh autopsy specimens were included in the study. The Peanut lectin (PNL) and Jack fruit lectin (JFL) conjugates showed positive binding with the cells in different lesions, while Winged Bean Lectin (WBL), despite its having a common inhibitory sugar, showed no binding even after neuraminidase treatment. These lectins revealed difference in the composition of glycoconjugates of benign and malignant thyroid cells. The HRP conjugated JFL and PNL may be of use in distinguishing carcinomatous tissues from benign tissues which makes them potential tools in the differential diagnosis of thyroid lesions. PMID:1625034

  1. Dissecting complement blockade for clinic use.

    PubMed

    Risitano, Antonio M

    2015-01-29

    In this issue of Blood, Peffault de Latour et al describe ex vivo measurements of complement activity in paroxysmal nocturnal hemoglobinuria (PNH) patients on eculizumab treatment. This is the first systematic pharmacodynamic (PD) study of eculizumab in PNH patients which shows that CH50 is a promising biomarker of therapeutic complement blockade. PMID:25634611

  2. Progress and Trends in Complement Therapeutics

    PubMed Central

    Ricklin, Daniel; Lambris, John D.

    2012-01-01

    The past few years have proven to be a highly successful and exciting period for the field of complement-directed drug discovery and development. Driven by promising experiences with the first marketed complement drugs, increased knowledge about the involvement of complement in health and disease, and improvements in structural and analytical techniques as well as animal models of disease, the field has seen a surge in creative approaches to therapeutically intervene at various stages of the cascade. An impressive panel of compounds that show promise in clinical trials is meanwhile being lined up in the pipelines of both small biotechnology and big pharmaceutical companies. Yet with this new focus on complement-targeted therapeutics, important questions concerning target selection, point and length of intervention, safety, and drug delivery emerge. In view of the diversity of the clinical disorders involving abnormal complement activity or regulation, which include both acute and chronic diseases and affect a wide range of organs, diverse yet specifically tailored therapeutic approaches may be needed to shift complement back into balance. This chapter highlights the key changes in the field that shape our current perception of complement-targeted drugs and provides a brief overview of recent strategies and emerging trends. Selected examples of complement-related diseases and inhibitor classes are highlighted to illustrate the diversity and creativity in field. PMID:22990692

  3. White kidney bean lectin exerts anti-proliferative and apoptotic effects on cancer cells.

    PubMed

    Chan, Yau Sang; Xia, Lixin; Ng, Tzi Bun

    2016-04-01

    A 60-kDa glucosamine binding lectin, white kidney bean lectin (WKBL), was purified from Phaseolus vulgaris cv. white kidney beans, by application of anion exchange chromatography on Q-Sepharose, affinity chromatography on Affi-gel blue gel, and FPLC-size exclusion on Superdex 75. The anti-proliferative activity of WKBL on HONE1 cells and HepG2 cells was stronger than the activity on MCF7 cells and WRL68 cells (IC50 values for a 48-h treatment with WKBL on HONE1 cells: 18.8?M; HepG2 cells: 19.7?M; MCF7 cells: 26.9?M; and WRL68 cells: >80?M). The activity could be reduced by addition of glucosamine, which occupies the binding sites of WKBL, indicating that carbohydrate binding is crucial for the activity. Annexin V-FITC and PI staining, JC-1 staining and Hoechst 33342 staining revealed that apoptosis was induced on WKBL-treated HONE1 cells and HepG2 cells, but not as obviously on MCF7 cells. Cell cycle analysis also showed a slight cell cycle arrest on HONE1 cells after WKBL treatment. Western blotting suggested that WKBL induced apoptosis of HONE1 cells occurred through the extrinsic apoptosis pathway, with detection of increased level of active caspase 3, 8 and 9. PMID:26769089

  4. Characterization of the Grp94/OS-9 chaperone-lectin complex

    PubMed Central

    Seidler, Paul M.; Shinsky, Stephen A.; Hong, Feng; Li, Zihai; Cosgrove, Michael S.; Gewirth, Daniel T.

    2014-01-01

    Grp94 is a macromolecular chaperone belonging to the hsp90 family and is the most abundant glycoprotein in the endoplasmic reticulum of mammals. In addition to its essential role in protein folding, Grp94 was proposed to participate in the ER associated degradation (ERAD) quality control pathway by interacting with the lectin OS-9, a sensor for terminally misfolded proteins (TMPs). To understand how OS-9 interacts with ER chaperone proteins, we mapped its interaction with Grp94. Glycosylation of the full length Grp94 protein was essential for OS-9 binding, although deletion of the Grp94 N-terminal domain relieved this requirement suggesting that the effect was allosteric rather than direct. Although yeast OS-9 is composed of a well-established N-terminal MRH lectin domain and a C-terminal dimerization domain, we find that the C-terminal domain of OS-9 in higher eukaryotes contains ‘mammalian-specific insets’ that are specifically recognized by the middle and C-terminal domains of Grp94. Additionally, the Grp94 binding domain in OS-9 was found to be intrinsically disordered. The biochemical analysis of the interacting regions provides insight into the manner by which the two associate, and additionally hints at a plausible biological role for the Grp94/OS-9 complex. PMID:25193139

  5. Lectin uptake and incorporation into the calcitic spicule of sea urchin embryos.

    PubMed

    Mozingo, Nancy M

    2015-06-01

    Primary mesenchyme cells (PMCs) are skeletogenenic cells that produce a calcareous endoskeleton in developing sea urchin larvae. The PMCs fuse to form a cavity in which spicule matrix proteins and calcium are secreted forming the mineralized spicule. In this study, living sea urchin embryos were stained with fluorescently conjugated wheat germ agglutinin, a lectin that preferentially binds to PMCs, and the redistribution of this fluorescent tag was examined during sea urchin development. Initially, fluorescence was associated primarily with the surface of PMCs. Subsequently, the fluorescent label redistributed to intracellular vesicles in the PMCs. As the larval skeleton developed, intracellular granular staining diminished and fluorescence appeared in the spicules. Spicules that were cleaned to remove membranous material associated with the surface exhibited bright fluorescence, which indicated that fluorescently labelled lectin had been incorporated into the spicule matrix. The results provide evidence for a cellular pathway in which material is taken up at the cell surface, sequestered in intracellular vesicles and then incorporated into the developing spicule. PMID:24735584

  6. Purification and properties of the basic lectins from winged bean seed [Psophocarpus tetragonolobus (L.) DC].

    PubMed

    Kortt, A A

    1984-02-01

    The seeds of winged bean, Psophocarpus tetragonolobus (L.) DC, contain a group of acidic lectins (pI approximately equal to 5.5) and a group of basic lectins (pI greater than 9.5) characterised by different erythrocyte hemagglutinating specificities. Three basic lectins were separated and purified to apparent homogeneity by chromatography on Ultrogel AcA44 and SP-Sephadex C-25. These lectins are glycoproteins with relative molecular mass of 58 000. The total carbohydrate content of the major lectins, B2 and B3, was 5% and was comprised of mannose, N-acetylglucosamine, fucose and xylose in amounts corresponding to 9, 8, 8 and 3 mol/58 000 g, respectively, for both lectins. Electrophoresis in dodecyl sulfate gave a single subunit of relative molecular mass 29 000 for each lectin. Isoelectric focusing in 8 M urea revealed two differently charged subunits in each of the isolated lectins; a total of three different subunits in the three lectins was observed. The basic lectins have essentially the same amino acid composition and contain no half-cystine. Two of the lectins (B2 and B3) showed identical amino-terminal sequences and the sequence of lectin B3 to residue 40 revealed extensive homology with other legume lectins such as soybean lectin. The basic lectins agglutinated trypsinized rabbit and trypsinized human (type A and B) erythrocytes but not trypsinized human (type O) erythrocytes. The lectins were inhibited by N-acetylgalactosamine, D-galactose and D-galactose containing disaccharides and trisaccharides. The specificity appears to be directed to glycosides having non-reducing (terminal) alpha-D-galactopyranosyl groups. beta-D-Galactosides, such as lactose, are poor inhibitors. PMID:6692833

  7. Complement activation in malaria: friend or foe?

    PubMed

    Biryukov, Sergei; Stoute, Jos A

    2014-05-01

    Complement is activated during malaria infection, but there is little evidence that it benefits the host. On the contrary, growing evidence points to the central role of complement activation in the pathogenesis of complicated malaria. Recent evidence suggests a critical role for C5a and the membrane attack complex in the pathogenesis of cerebral malaria, and for C5a in the pathogenesis of placental malaria. In addition, erythrocytes of children with severe malarial anemia have increased deposition of C3b and decreased capacity to regulate complement activation, that probably increase their susceptibility to destruction by liver and splenic macrophages. These observations justify further investigation of the role of complement in malaria and the testing of complement inhibitors as adjunctive treatment for severe malaria. PMID:24508275

  8. Merging carbohydrate chemistry with lectin histochemistry to study inhibition of lectin binding by glycoclusters in the natural tissue context.

    PubMed

    André, Sabine; Kaltner, Herbert; Kayser, Klaus; Murphy, Paul V; Gabius, Hans-Joachim

    2016-02-01

    Recognition of glycans by lectins leads to cell adhesion and growth regulation. The specificity and selectivity of this process are determined by carbohydrate structure (sequence and shape) and topology of its presentation. The synthesis of (neo)glycoconjugates with bi- to oligo-valency (glycoclusters) affords tools to delineate structure-activity relationships by blocking lectin binding to an artificial matrix, often a glycoprotein, or cultured cell lines. The drawback of these assays is that glycan presentation is different from that in tissues. In order to approach the natural context, we here introduce lectin histochemistry on fixed tissue sections to determine the susceptibility of binding of two plant lectins, i.e., GSA-II and WGA, to a series of 10 glycoclusters. Besides valency, this panel covers changes in the anomeric position (α/β) and the atom at the glycosidic linkage (O/S). Flanked by cell and solid-phase assays with human tumor lines and two mucins, respectively, staining (intensity and profile) was analyzed in sections of murine jejunum, stomach and epididymis as a function of glycocluster presence. The marked and differential sensitivity of signal generation to structural aspects of the glycoclusters proves the applicability of this method. This enables comparisons between data sets obtained by using (neo)glycoconjugates, cells and the tissue context as platforms. The special advantage of processing tissue sections is the monitoring of interference with lectin association at sites that are relevant for functionality. Testing glycoclusters in lectin histochemistry will especially be attractive in cases of multi-target recognition (glycans, proteins and lipids) by a tissue lectin. PMID:26553286

  9. Probing the cons and pros of lectin-induced immunomodulation: case studies for the mistletoe lectin and galectin-1.

    PubMed

    Gabius, H J

    2001-07-01

    When imagining to monitor animal cells through a microscope with resolution at the molecular level, a salient attribute of their surfaces will be the abundance of glycan chains. They present galactosides at their termini widely extending like tentacles into the extracellular space. Their spatial accessibility and their potential for structural variability endow especially these glycan parts with capacity to act as docking points for molecular sensors (sugar receptors such as lectins). Binding and ligand clustering account for transmission of post-binding signals into the cell interior. The range of triggered activities has turned plant lectins into popular tools in cell biology and immunology. Potential for clinical application has been investigated rigorously only in recent years. As documented in vitro and in vivo for the galactoside-specific mistletoe lectin, its apparent immunomodulatory capacity reflected in upregulation of production of proinflammatory cytokines will not necessarily be clinically favorable but a double-edged sword. In fact, lectin application has been shown to stimulate tumor growth in cell lines, histocultures of human tumors and in two animal models using chemical carcinogenesis or tumor transplantation. When testing immunological effects of the endogenous lectin galectin-1, protection against disorders mediated by activated T cells came up for consideration. Elimination of these cells via CD7-dependent induction of apoptosis, and a shift to the Th2 response by the galectin, are factors to ameliorate disease states. This result encourages further efforts with other galectins. Functional redundancy, synergism, diversity or antagonism among galectins are being explored to understand the actual role of this class of endogenous lectins in inflammation. Regardless of the results of further preclinical testing for galectin-1, these two case studies break new ground in our understanding how glycans as ligands for lectins convey reactivity to immune cells, with impact on the course of a tumor or autoimmune disease. PMID:11522395

  10. Structure Predictions of Two Bauhinia variegata Lectins Reveal Patterns of C-Terminal Properties in Single Chain Legume Lectins

    PubMed Central

    Moreira, Gustavo M. S. G.; Conceio, Fabricio R.; McBride, Alan J. A.; Pinto, Luciano da S.

    2013-01-01

    Bauhinia variegata lectins (BVL-I and BVL-II) are single chain lectins isolated from the plant Bauhinia variegata. Single chain lectins undergo post-translational processing on its N-terminal and C-terminal regions, which determines their physiological targeting, carbohydrate binding activity and pattern of quaternary association. These two lectins are isoforms, BVL-I being highly glycosylated, and thus far, it has not been possible to determine their structures. The present study used prediction and validation algorithms to elucidate the likely structures of BVL-I and -II. The program Bhageerath-H was chosen from among three different structure prediction programs due to its better overall reliability. In order to predict the C-terminal region cleavage sites, other lectins known to have this modification were analysed and three rules were created: (1) the first amino acid of the excised peptide is small or hydrophobic; (2) the cleavage occurs after an acid, polar, or hydrophobic residue, but not after a basic one; and (3) the cleavage spot is located 5-8 residues after a conserved Leu amino acid. These rules predicted that BVL-I and II would have fifteen C-terminal residues cleaved, and this was confirmed experimentally by Edman degradation sequencing of BVL-I. Furthermore, the C-terminal analyses predicted that only BVL-II underwent ?-helical folding in this region, similar to that seen in SBA and DBL. Conversely, BVL-I and -II contained four conserved regions of a GS-I association, providing evidence of a previously undescribed X4+unusual oligomerisation between the truncated BVL-I and the intact BVL-II. This is the first report on the structural analysis of lectins from Bauhinia spp. and therefore is important for the characterisation C-terminal cleavage and patterns of quaternary association of single chain lectins. PMID:24260572

  11. Mannan-Binding Lectin in Cardiovascular Disease

    PubMed Central

    Cedzy?ski, Maciej

    2014-01-01

    Cardiovascular disease remains the leading cause of mortality and morbidity worldwide so research continues into underlying mechanisms. Since innate immunity and its potent component mannan-binding lectin have been proven to play an important role in the inflammatory response during infection and ischaemia-reperfusion injury, attention has been paid to its role in the development of cardiovascular complications as well. This review provides a general outline of the structure and genetic polymorphism of MBL and its role in inflammation/tissue injury with emphasis on associations with cardiovascular disease. MBL appears to be involved in the pathogenesis of atherosclerosis and, in consequence, coronary artery disease and also inflammation and tissue injury after myocardial infarction and heart transplantation. The relationship between MBL and disease is rather complex and depends on different genetic and environmental factors. That could be why the data obtained from animal and clinical studies are sometimes contradictory proving not for the first time that innate immunity is a double-edge sword, sometimes beneficial and, at other times disastrous for the host. PMID:24877121

  12. Lectin-like activity from Persea americana.

    PubMed

    Meade, N A; Staat, R H; Langley, S D; Doyle, R J

    1980-01-15

    An extract from the seeds of Persea americana possessed an erythro-agglutinating activity. The agglutinin was devoid of specificity for carbohydrates, but interacted readily with basic proteins or basic polyamino acids. The interaction between the agglutinin and egg-white lysozyme was not inhibited by chaotropic salts, but was sensitive to relatively low concentrations of urea. An affinity chromatographic procedure was developed in an effort to purify the agglutinin. Products from the chromatographic procedure were found not to contain higher specific agglutinating activities than the crude extract. Amino acid acid analyses of the extract showed the presence of relatively high proportions of glutamic and aspartic acids. In addition, the extract contained phosphorus and a visible chromophore. The agglutinin was resistant to detergents and denaturants, and proteases, nucleases, and other enzymes. The results suggest that, as opposed to other plant agglutinins, the active component from Persea is not a protein. Similarly, in contrast to many lectins, the agglutinin from Persea was not mitogenic for mouse lymphocytes. The agglutinin partially inhibited the mitogenesis of lymphocytes when the cells were treated with concanavalin A, or with bacterial lipopolysaccharide. PMID:7353211

  13. Complement System Part II: Role in Immunity

    PubMed Central

    Merle, Nicolas S.; Noe, Remi; Halbwachs-Mecarelli, Lise; Fremeaux-Bacchi, Veronique; Roumenina, Lubka T.

    2015-01-01

    The complement system has been considered for a long time as a simple lytic cascade, aimed to kill bacteria infecting the host organism. Nowadays, this vision has changed and it is well accepted that complement is a complex innate immune surveillance system, playing a key role in host homeostasis, inflammation, and in the defense against pathogens. This review discusses recent advances in the understanding of the role of complement in physiology and pathology. It starts with a description of complement contribution to the normal physiology (homeostasis) of a healthy organism, including the silent clearance of apoptotic cells and maintenance of cell survival. In pathology, complement can be a friend or a foe. It acts as a friend in the defense against pathogens, by inducing opsonization and a direct killing by C5b–9 membrane attack complex and by triggering inflammatory responses with the anaphylatoxins C3a and C5a. Opsonization plays also a major role in the mounting of an adaptive immune response, involving antigen presenting cells, T-, and B-lymphocytes. Nevertheless, it can be also an enemy, when pathogens hijack complement regulators to protect themselves from the immune system. Inadequate complement activation becomes a disease cause, as in atypical hemolytic uremic syndrome, C3 glomerulopathies, and systemic lupus erythematosus. Age-related macular degeneration and cancer will be described as examples showing that complement contributes to a large variety of conditions, far exceeding the classical examples of diseases associated with complement deficiencies. Finally, we discuss complement as a therapeutic target. PMID:26074922

  14. Molecular biology of seed storage proteins and lectins

    SciTech Connect

    Shannon, L.M.; Chrispeels, M.J.

    1986-01-01

    The seeds of many plants contain abundant storage protein and lectin which have been the subject of biochemical investigations for over a hundred years. Because these proteins represent abundant gene products translated from abundant messages, they were among the first plant proteins to which the techniques of molecular biology were applied. Many of the proteins have now been purified and characterized and their amino acid sequences have been determined; some have been crystallized and their 3-dimensional structure is known. Studies of their biosynthesis, transport and accumulation in protein bodies have lead to a greater understanding of the dynamics of plant cell organelles. Seed storage proteins and lectins are encoded by small gene families whose members show considerable homology and appear to have been conserved in evolution. The expression of these genes is highly regulated in time (development) and space (tissue). The recent discovery that other plant organs synthesize lectins, or lectin-like proteins which are closely related to the seed lectins, has lent additional support to the search for a function for these intriguing proteins; this finding also indicates that different members of the gene family are expressed in different tissues. Individual papers were processed separately for the data base.

  15. Prevalence of the F-type lectin domain.

    PubMed

    Bishnoi, Ritika; Khatri, Indu; Subramanian, Srikrishna; Ramya, T N C

    2015-08-01

    F-type lectins are fucolectins with characteristic fucose and calcium-binding sequence motifs and a unique lectin fold (the "F-type" fold). F-type lectins are phylogenetically widespread with selective distribution. Several eukaryotic F-type lectins have been biochemically and structurally characterized, and the F-type lectin domain (FLD) has also been studied in the bacterial proteins, Streptococcus mitis lectinolysin and Streptococcus pneumoniae SP2159. However, there is little knowledge about the extent of occurrence of FLDs and their domain organization, especially, in bacteria. We have now mined the extensive genomic sequence information available in the public databases with sensitive sequence search techniques in order to exhaustively survey prokaryotic and eukaryotic FLDs. We report 437 FLD sequence clusters (clustered at 80% sequence identity) from eukaryotic, eubacterial and viral proteins. Domain architectures are diverse but mostly conserved in closely related organisms, and domain organizations of bacterial FLD-containing proteins are very different from their eukaryotic counterparts, suggesting unique specialization of FLDs to suit different requirements. Several atypical phylogenetic associations hint at lateral transfer. Among eukaryotes, we observe an expansion of FLDs in terms of occurrence and domain organization diversity in the taxa Mollusca, Hemichordata and Branchiostomi, perhaps coinciding with greater emphasis on innate immune strategies in these organisms. The naturally occurring FLDs with diverse domain organizations that we have identified here will be useful for future studies aimed at creating designer molecular platforms for directing desired biological activities to fucosylated glycoconjugates in target niches. PMID:25943580

  16. Characterization of a new lectin of soybean vegetative tissues.

    PubMed Central

    Spilatro, S R; Cochran, G R; Walker, R E; Cablish, K L; Bittner, C C

    1996-01-01

    Lectins are carbohydrate-binding proteins that occur widely among plants. Lectins of plant vegetative tissues are less well characterized than those of seeds. Previously, a protein of soybean (Glycine max [L.] Merr.) leaves was shown to possess properties similar to the seed lectin. Here we show that the N-terminal amino acid sequence of this protein shares 63% identity with the seed lectin. Immunoblot analysis indicated that the protein occurs in leaves, petioles, stems, and cotyledons of seedlings but not in seeds. These observations prompted designation of the protein as a soybean vegetative lectin (SVL). Immunohistochemical localization in leaves indicated that SVL was localized to the vacuoles of bundle-sheath and paraveinal mesophyll cells. Removal of sink tissues or exposure to atmospheric methyl jasmonate caused increased levels of SVL in leaves and cotyledons. Co-precipitation of SVL and the soybean vegetative storage protein (VSP) during purification suggested an interaction between these proteins. SVL-horseradish peroxidase conjugate bound to dot blots of VSP or SVL, and binding was inhibited by porcine stomach mucin and heparin but not simple carbohydrates. Binding between SVL and VSP and similarities in localization and regulation support a possible in vivo interaction between these proteins. PMID:8819869

  17. The specificity of frutalin lectin using biomembrane models.

    PubMed

    Nobre, Thatyane M; Pavinatto, Felippe J; Cominetti, Mrcia R; Selistre de-Arajo, Helosa S; Zaniquelli, Maria E D; Beltramini, Leila M

    2010-08-01

    Frutalin is a homotetrameric alpha-d-galactose (d-Gal)-binding lectin that activates natural killer cells in vitro and promotes leukocyte migration in vivo. Because lectins are potent lymphocyte stimulators, understanding the interactions that occur between them and cell surfaces can help to the action mechanisms involved in this process. In this paper, we present a detailed investigation of the interactions of frutalin with phospho- and glycolipids using Langmuir monolayers as biomembrane models. The results confirm the specificity of frutalin for d-Gal attached to a biomembrane. Adsorption of frutalin was more efficient for the galactose polar head lipids, in contrast to the one for sulfated galactose, in which a lag time is observed, indicating a rearrangement of the monolayer to incorporate the protein. Regarding ganglioside GM1 monolayers, lower quantities of the protein were adsorbed, probably due to the farther apart position of d-galactose from the interface. Binary mixtures containing galactocerebroside revealed small domains formed at high lipid packing in the presence of frutalin, suggesting that lectin induces the clusterization and the forming of domains in vitro, which may be a form of receptor internalization. This is the first experimental evidence of such lectin effect, and it may be useful to understand the mechanism of action of lectins at the molecular level. PMID:20353752

  18. Lectin Activity in Gut Extract of Culex pipiens

    PubMed Central

    Koosha, Mona; Abai, Mohammad Reza; Abolhasani, Mandan; Charedar, Soroor; Basseri, Hamid Reza

    2013-01-01

    Background: The role of lectins is important in interaction between pathogens and mosquito vectors. This study was performed to identify agglutinin activities of protein molecules on the midgut of Culex pipiens. Methods: Culex pipiens was reared in insectray condition and the midguts of males and females (blood fed and unfed) were dissected separately in Tris-HCl buffer. The extracts of midguts were applied for hemagglutinin assay against red blood cells of rabbit, mouse, rat, dog, horse, sheep, guinea pig, cow, human (A, B, AB, O groups). Then, the RBCs with relatively high agglutinin activity were chosen for carbohydrate inhibition assay. D (+) glucose, D (+) galactose, D (+) mannose, D (?) fructose, D (?) arabinose, L (?) fucose, lactose, N-acetyl-D-glucosamine, N-acetyl-D-galactosamine, sialic acid were used to specify carbohydrate binding lectin. Results: The highest agglutinin activities were found against sheep and rabbits RBCs. Sexual diversity of agglutinin activities was observed among midgut extraction of males and females. In addition, variation in agglutinin activity of blood fed and unfed female mosquitoes were detected. The lectin activity was inhibited highly with glucose, galactose, fucose and fructose but less inhibitor activities was observed by arabinose, N-acetyl-D-galactosamine, n-acetyl-d-glucosamine, lactose and mannose. Conclusion: The secretion of hemagglutinins (lectins or lectin-like molecules) in the digestive system depends on the type of food in the gut. This suggests that emptying of the gut in preparation for protein rich food probably starts the secretion of hemagglutinins. PMID:23785692

  19. Complement modulation of T cell immune responses during homeostasis and disease

    PubMed Central

    Clarke, Elizabeth V.; Tenner, Andrea J.

    2014-01-01

    The complement system is an ancient and critical effector mechanism of the innate immune system as it senses, kills, and clears infectious and/or dangerous particles and alerts the immune system to the presence of the infection and/or danger. Interestingly, an increasing number of reports have demonstrated a clear role for complement in the adaptive immune system as well. Of note, a number of recent studies have identified previously unknown roles for complement proteins, receptors, and regulators in T cell function. Here, we will review recent data demonstrating the influence of complement proteins C1q, C3b/iC3b, C3a (and C3aR), and C5a (and C5aR) and complement regulators DAF (CD55) and CD46 (MCP) on T cell function during homeostasis and disease. Although new concepts are beginning to emerge in the field of complement regulation of T cell function, future experiments should focus on whether complement is interacting directly with the T cell or is having an indirect effect on T cell function via APCs, the cytokine milieu, or downstream complement activation products. Importantly, the identification of the pivotal molecular pathways in the human systems will be beneficial in the translation of concepts derived from model systems to therapeutic targeting for treatment of human disorders. PMID:25210145

  20. Complement in action: an analysis of patent trends from 1976 through 2011.

    PubMed

    Yang, Kun; DeAngelis, Robert A; Reed, Janet E; Ricklin, Daniel; Lambris, John D

    2013-01-01

    Complement is an essential part of the innate immune response. It interacts with diverse endogenous pathways and contributes to the maintenance of homeostasis, the modulation of adaptive immune responses, and the development of various pathologies. The potential usefulness, in both research and clinical settings, of compounds that detect or modulate complement activity has resulted in thousands of publications on complement-related innovations in fields such as drug discovery, disease diagnosis and treatment, and immunoassays, among others. This study highlights the distribution and publication trends of patents related to the complement system that were granted by the United States Patent and Trademark Office from 1976 to the present day. A comparison to complement-related documents published by the World Intellectual Property Organization is also included. Statistical analyses revealed increasing diversity in complement-related research interests over time. More than half of the patents were found to focus on the discovery of inhibitors; interest in various inhibitor classes exhibited a remarkable transformation from chemical compounds early on to proteins and antibodies in more recent years. Among clinical applications, complement proteins and their modulators have been extensively patented for the diagnosis and treatment of eye diseases (especially age-related macular degeneration), graft rejection, cancer, sepsis, and a variety of other inflammatory and immune diseases. All of the patents discussed in this chapter, as well as those from other databases, are available from our newly constructed complement patent database: www.innateimmunity.us/patent. PMID:23402036

  1. Complement in Action: An Analysis of Patent Trends from 1976 Through 2011.

    PubMed

    Yang, Kun; Deangelis, Robert A; Reed, Janet E; Ricklin, Daniel; Lambris, John D

    2013-01-01

    Complement is an essential part of the innate immune response. It interacts with diverse endogenous pathways and contributes to the maintenance of homeostasis, the modulation of adaptive immune responses, and the development of various pathologies. The potential usefulness, in both research and clinical settings, of compounds that detect or modulate complement activity has resulted in thousands of publications on complement-related innovations in fields such as drug discovery, disease diagnosis and treatment, and immunoassays, among others. This study highlights the distribution and publication trends of patents related to the complement system that were granted by the United States Patent and Trademark Office from 1976 to the present day. A comparison to complement-related documents published by the World Intellectual Property Organization is also included. Statistical analyses revealed increasing diversity in complement-related research interests over time. More than half of the patents were found to focus on the discovery of inhibitors; interest in various inhibitor classes exhibited a remarkable transformation from chemical compounds early on to proteins and antibodies in more recent years. Among clinical applications, complement proteins and their modulators have been extensively patented for the diagnosis and treatment of eye diseases (especially age-related macular degeneration), graft rejection, cancer, sepsis, and a variety of other inflammatory and immune diseases. All of the patents discussed in this chapter, as well as those from other databases, are available from our newly constructed complement patent database: www.innateimmunity.us/patent . PMID:22990712

  2. Human genetic deficiencies reveal the roles of complement in the inflammatory network: Lessons from nature

    PubMed Central

    Lappegrd, Knut Tore; Christiansen, Dorte; Pharo, Anne; Thorgersen, Ebbe Billmann; Hellerud, Bernt Christian; Lindstad, Julie; Nielsen, Erik Waage; Bergseth, Grethe; Fadnes, Dag; Abrahamsen, Tore G.; Hiby, E. Arne; Schejbel, Lone; Garred, Peter; Lambris, John D.; Harboe, Morten; Mollnes, Tom Eirik

    2009-01-01

    Complement component C5 is crucial for experimental animal inflammatory tissue damage; however, its involvement in human inflammation is incompletely understood. The responses to Gram-negative bacteria were here studied taking advantage of human genetic complement-deficienciesnature's own knockoutsincluding a previously undescribed C5 defect. Such deficiencies provide a unique tool for investigating the biological role of proteins. The experimental conditions allowed cross-talk between the different inflammatory pathways using a whole blood model based on the anticoagulant lepirudin, which does not interfere with the complement system. Expression of tissue factor, cell adhesion molecules, and oxidative burst depended highly on C5, mediated through the activation product C5a, whereas granulocyte enzyme release relied mainly on C3 and was C5a-independent. Release of cytokines and chemokines was mediated to varying degrees by complement and CD14; for example, interleukin (IL)-1? and IL-8 were more dependent on complement than IFN-? and IL-6, which were highly dependent on CD14. IL-1 receptor antagonist (IL-1ra) and IFN-? inducible protein 10 (IP-10) were fully dependent on CD14 and inversely regulated by complement, that is, complement deficiency and complement inhibition enhanced their release. Granulocyte responses were mainly complement-dependent, whereas monocyte responses were more dependent on CD14. Notably, all responses were abolished by combined neutralization of complement and CD14. The present study provides important insight into the comprehensive role of complement in human inflammatory responses to Gram-negative bacteria. PMID:19717455

  3. A sialic acid-binding lectin from the mushroom Hericium erinaceum.

    PubMed

    Kawagishi, H; Mori, H; Uno, A; Kimura, A; Chiba, S

    1994-02-28

    A lectin was isolated from the mushroom Hericium erinaceum. This lectin is composed of two different subunits of 15 and 16 kDa and the molecular mass of the intact lectin was estimated to be 54 kDa by gel filtration. It exhibits specificity towards sialic acids, especially N-glycolylneuraminic acid. PMID:8119408

  4. Histological and lectin histochemical studies on the olfactory and respiratory mucosae of the sheep.

    PubMed

    Ibrahim, Dalia; Nakamuta, Nobuaki; Taniguchi, Kazumi; Yamamoto, Yoshio; Taniguchi, Kazuyuki

    2014-03-01

    The olfactory and respiratory mucosae of the Corriedale sheep were examined using lectin histochemistry in order to clarify the histochemical and glycohistochemical differences between these two tissues. The olfactory epithelium was stained with 13 lectins out of 21 lectins examined, while the respiratory epithelium was positive to 16 lectins. The free border of both of the olfactory and respiratory epithelia was stained with 12 lectins: Wheat germ agglutinin (WGA), succinylated-wheat germ agglutinin (s-WGA), Lycopersicon esculentum lectin (LEL), Solanum tuberosum lectin (STL), Datura stramonium lectin (DSL), Soybean agglutinin (SBA), Bandeiraea simplicifolia lectin-I (BSL-I), Ricinus communis agglutinin-I (RCA-120), Erythrina cristagalli lectin (ECL), Concanavalin A (Con A), Phaseolus vulgaris agglutinin-E (PHA-E) and Phaseolus vulgaris agglutinin-L (PHA-L). The associated glands of the olfactory mucosa, Bowman's glands, were stained with 13 lectins. While both the goblet cells and mucous nasal glands were stained with 8 lectins; five of them (WGA, s-WGA, STL, Vicia villosa agglutinin (VVA) and ECL) were mutually positive among the Bowman's glands, mucous nasal glands and the goblet cells. These findings indicate that the glycohistochemical characteristics of the free borders of both olfactory and respiratory epithelia are similar to each other, suggesting that secretions from the Bowman's glands and those of the goblet cells and mucous nasal glands are partially exchanged between the surface of two epithelia to contribute the functions of the respiratory epithelium and the olfactory receptor cells, respectively. PMID:24200894

  5. Further screening of Aspergillus species for occurrence of lectins and their partial characterization.

    PubMed

    Singh, Ram Sarup; Bhari, Ranjeeta; Rai, Jyoti

    2010-02-01

    Fifteen species of Aspergillus were screened for occurrence of lectins. Nine of them (A. sydowii, A. candidus, A. allahabadi, A. terricola, A. ficuum, A. sparsus, A. carneus, A. pulvinus and A. aculeatus) were found to possess lectin activity. None of the species elaborated lectin in culture supernatant. All the lectins agglutinated rat, pig and rabbit erythrocytes. A. sydowii, A. candidus, A. allahabadi, A. terricola, A. ficuum, A. sparsus, A. carneus and A. aculeatus lectins agglutinated all human type erythrocytes equally, while A. pulvinus lectin specifically agglutinated human type A and O erythrocytes. Neuraminidase and protease treatment to erythrocytes substantially augmented lectin titres manyfold. Lectins showed specificity to mucin and asialofetuin and all of them were specific to L-arabinose except that of A. carneus. Lectins from A. sydowii, A. ficuum, A. sparsus and A. carneus displayed remarkable specificities to D-xylose. Maximum lectin activity was expressed by 11 day old cultures of A. sydowii (titre 32), A. ficuum (titre 64) and A. sparsus (titre 1024). Lectins from A. aculeatus, A. candidus and A. terricola were expressed by 7-10 days, 6-9 days and 5-11 days old cultures, respectively. A. allahabadi cultures exhibited maximum lectin activity (titre 32) after 8-10 days of cultivation. A. carneus and A. pulvinus expressed optimal titres of 32 and 8, respectively on the 9th day. PMID:20175125

  6. EspP, a serine protease of enterohemorrhagic Escherichia coli, impairs complement activation by cleaving complement factors C3/C3b and C5.

    PubMed

    Orth, Dorothea; Ehrlenbach, Silvia; Brockmeyer, Jens; Khan, Abdul Basit; Huber, Georg; Karch, Helge; Sarg, Bettina; Lindner, Herbert; Wrzner, Reinhard

    2010-10-01

    Hemolytic-uremic syndrome (HUS) is a life-threatening disorder characterized by hemolytic anemia, thrombocytopenia, and renal insufficiency. It is caused mainly by infections with enterohemorrhagic Escherichia coli (EHEC). Recently, Shiga toxin 2, the best-studied virulence factor of EHEC, was reported to interact with complement, implying that complement may be involved in the pathogenesis of EHEC-induced HUS. The aim of the present study was to investigate whether or not the serine protease EspP, an important virulence factor of EHEC, interacts with complement proteins. EspP did not have any effect on the integrity of factor H or factor I. However, EspP was shown to cleave purified C3/C3b and C5. Cleavage of the respective complement proteins also occurred in normal human serum (NHS) as a source of C3/C3b or C5 or when purified complement proteins were added to the supernatant of an EspP-producing wild-type strain. Edman degradation allowed unequivocal mapping of all three main C3b fragments but not of the three main C5 fragments. Complement activation was significantly downregulated in all three pathways for C5-depleted serum to which C5, preincubated with EspP, was added (whereas C5 preincubated with an EspP mutant was able to fully reconstitute complement activation). This indicates that EspP markedly destroyed the functional activity, as measured by a commercial total complement enzyme-linked immunosorbent assay (Wieslab). Downregulation of complement by EspP in vivo may influence the colonization of EHEC bacteria in the gut or the disease severity of HUS. PMID:20643852

  7. EspP, a Serine Protease of Enterohemorrhagic Escherichia coli, Impairs Complement Activation by Cleaving Complement Factors C3/C3b and C5?

    PubMed Central

    Orth, Dorothea; Ehrlenbach, Silvia; Brockmeyer, Jens; Khan, Abdul Basit; Huber, Georg; Karch, Helge; Sarg, Bettina; Lindner, Herbert; Wrzner, Reinhard

    2010-01-01

    Hemolytic-uremic syndrome (HUS) is a life-threatening disorder characterized by hemolytic anemia, thrombocytopenia, and renal insufficiency. It is caused mainly by infections with enterohemorrhagic Escherichia coli (EHEC). Recently, Shiga toxin 2, the best-studied virulence factor of EHEC, was reported to interact with complement, implying that complement may be involved in the pathogenesis of EHEC-induced HUS. The aim of the present study was to investigate whether or not the serine protease EspP, an important virulence factor of EHEC, interacts with complement proteins. EspP did not have any effect on the integrity of factor H or factor I. However, EspP was shown to cleave purified C3/C3b and C5. Cleavage of the respective complement proteins also occurred in normal human serum (NHS) as a source of C3/C3b or C5 or when purified complement proteins were added to the supernatant of an EspP-producing wild-type strain. Edman degradation allowed unequivocal mapping of all three main C3b fragments but not of the three main C5 fragments. Complement activation was significantly downregulated in all three pathways for C5-depleted serum to which C5, preincubated with EspP, was added (whereas C5 preincubated with an EspP mutant was able to fully reconstitute complement activation). This indicates that EspP markedly destroyed the functional activity, as measured by a commercial total complement enzyme-linked immunosorbent assay (Wieslab). Downregulation of complement by EspP in vivo may influence the colonization of EHEC bacteria in the gut or the disease severity of HUS. PMID:20643852

  8. Human and rodent decay-accelerating factors (CD55) are not species restricted in their complement-inhibiting activities

    PubMed Central

    Harris, C L; Spiller, O B; Morgan, B P

    2000-01-01

    Homologous complement activation is restricted on cells by the complement regulators, decay-accelerating factor (DAF), membrane cofactor protein (MCP) and CD59. These proteins act in concert with other membrane structures to protect cells from homologous complement attack. In contrast, cells are usually sensitive to heterologous complement attack. It has been suggested that species-specific restriction of complement activation can be attributed to the inability of regulators to inhibit across species. We have investigated the capacities of human, rat and mouse analogues of DAF to regulate homologous and heterologous complement. Cells transfected with cDNA encoding these analogues were protected from heterologous complement attack. C3b-deposition experiments indicated that whilst cells were best protected by DAF from the same species, all three analogues inhibited human, rat and mouse complement. Comparable results were obtained in haemolysis assays using soluble, recombinant forms of the proteins. Inhibition of the classical pathway (CP) was best achieved with homologous DAF, although human DAF also inhibited rat complement, rat DAF also inhibited human complement and mouse DAF inhibited complement from all species. Human DAF was the best inhibitor of alternative pathway (AP)-mediated attack, inhibiting complement from all species. Mouse DAF inhibited mouse and rat AP, whilst rat DAF inhibited only rat AP. These data indicate that human and rodent analogues of DAF are not species restricted and highlights interesting differences in the capacity to regulate AP and CP. This has implications in broader fields of research, such as xenotransplantation, where cross-species regulation of complement is of paramount importance. PMID:10929073

  9. Crystal structure of Dioclea violacea lectin and a comparative study of vasorelaxant properties with Dioclea rostrata lectin.

    PubMed

    Bezerra, Maria Júlia Barbosa; Rodrigues, Natália Velloso Fontenelle Camelo; Pires, Alana de Freitas; Bezerra, Gustavo Arruda; Nobre, Camila Bezerra; Alencar, Kássia Lys de Lima; Soares, Pedro Marcos Gomes; do Nascimento, Kyria Santiago; Nagano, Celso Shiniti; Martins, Jorge Luiz; Gruber, Karl; Sampaio, Alexandre Holanda; Delatorre, Plinio; Rocha, Bruno Anderson Matias; Assreuy, Ana Maria Sampaio; Cavada, Benildo Sousa

    2013-04-01

    Lectins from Diocleinae subtribe belong to the family of legume lectins and are characterized by high identity between their amino acids sequences. It has been shown that punctual differences in amino acid sequences, such as one single amino acid or an alternative conformation, represent changes in biological activities caused by these lectins. Therefore, a more detailed understanding of three-dimensional structures of these proteins is essential for accurate analyzing the relationship between structure and function. In this study lectins purified from the seeds of Dioclea violacea (DVL) and Dioclea rostrata (DRL) were compared with regard to crystal structure and vasorelaxant properties. Differences in structure of lectins were found to be reflected in differences in vasorelaxant effects based on their high specificity and selectivity for cell glycans. Binding activity was related to the position of specific residues in the carbohydrate recognition domain (CRD). DVL complexed structure was solved by X-ray crystallography and was compared to native DVL and DRL. Therefore, DVL was co-crystallized with X-Man, and a molecular modeling with X-Man complexed with DVL was done to compare the complexed and native forms adjusted fit. The relatively narrow and deep CRD in DVL promotes little interaction with carbohydrates; in contrast, the wider and shallower CRD in DRL favors interaction. This seems to explain differences in the level of relaxation induced by DVL (43%) and DRL (96%) in rat aortic rings. PMID:23353644

  10. Complement regulation: physiology and disease relevance

    PubMed Central

    2015-01-01

    The complement system is part of the innate immune response and as such defends against invading pathogens, removes immune complexes and damaged self-cells, aids organ regeneration, confers neuroprotection, and engages with the adaptive immune response via T and B cells. Complement activation can either benefit or harm the host organism; thus, the complement system must maintain a balance between activation on foreign or modified self surfaces and inhibition on intact host cells. Complement regulators are essential for maintaining this balance and are classified as soluble regulators, such as factor H, and membrane-bound regulators. Defective complement regulators can damage the host cell and result in the accumulation of immunological debris. Moreover, defective regulators are associated with several autoimmune diseases such as atypical hemolytic uremic syndrome, dense deposit disease, age-related macular degeneration, and systemic lupus erythematosus. Therefore, understanding the molecular mechanisms by which the complement system is regulated is important for the development of novel therapies for complement-associated diseases. PMID:26300937

  11. A lectin with antifungal activity from the mussel Crenomytilus grayanus.

    PubMed

    Chikalovets, Irina V; Chernikov, Oleg V; Pivkin, Mikhail V; Molchanova, Valentina I; Litovchenko, Alina P; Li, Wei; Lukyanov, Pavel A

    2015-02-01

    Lectins (carbohydrate-binding proteins) are well known to actively participate in the defense functions of vertebrates and invertebrates where they play an important role in the recognition of foreign particles. In this study, we investigated of in vitro antifungal activity of lectin from the mussel Crenomytilus grayanus (CGL). Enzyme-linked immunosorbent assay indicated that CGL was predominantly detectable in tissues of mantle and to a lesser degree in the tissues of muscle, hepatopancreas, gill and hemocytes. After challenged by Pichia pastoris the level of CGL was upregulated and reached the maximum level at 12 h post challenge and recovered to the original level at 24 h. The lectin was capable of inhibiting the germination of spores and hyphal growth in the fungi. All these results indicated that CGL is involved in the innate immune response in mollusc animals. PMID:25482060

  12. Cell surface lectin-binding glycoconjugates on marine planktonic protists.

    PubMed

    Roberts, Emily C; Zubkov, Mikhail V; Martin-Cereceda, Mercedes; Novarino, Gianfranco; Wootton, Emma C

    2006-12-01

    Carbohydrate-protein interactions appear to play an important role in the phagocytosis of microbial prey by free-living protozoa. The present study utilizes FITC-labelled plant lectins to investigate the presence and localization of cell surface glycoconjugates on live and fixed planktonic protists (Dunaliella primolecta, Oxyrrhis marina, Goniomonas amphinema, Paraphysomonas vestita and Euplotes vannus). With live flagellate preparations, lectins primarily bound to external cell surfaces, with minimal internal staining observed. In contrast, cell fixation permeabilized cell membranes, allowing lectins to bind to internal structures, such as nuclear membranes and food vacuoles, interfering with the characterization of cell surface glycoconjugates. The method developed to label cell surface sugar moieties of live planktonic protists successfully overcomes the problems associated with fixation, and thus provides a useful protocol for future studies on protistan cell surface carbohydrate characterization. PMID:17147765

  13. Marine sponge lectins: actual status on properties and biological activities.

    PubMed

    Gomes Filho, Sandro Mascena; Cardoso, Jusclio Donizete; Anaya, Katya; Silva do Nascimento, Edilza; de Lacerda, Jos Thalles Jucelino Gomes; Mioso, Roberto; Santi Gadelha, Tatiane; de Almeida Gadelha, Carlos Alberto

    2015-01-01

    Marine sponges are primitive metazoans that produce a wide variety of molecules that protect them against predators. In studies that search for bioactive molecules, these marine invertebrates stand out as promising sources of new biologically-active molecules, many of which are still unknown or little studied; thus being an unexplored biotechnological resource of high added value. Among these molecules, lectins are proteins that reversibly bind to carbohydrates without modifying them. In this review, various structural features and biological activities of lectins derived from marine sponges so far described in the scientific literature are discussed. From the results found in the literature, it could be concluded that lectins derived from marine sponges are structurally diverse proteins with great potential for application in the production of biopharmaceuticals, especially as antibacterial and antitumor agents. PMID:25549059

  14. Complementation of a red-light-indifferent cyanobacterial mutant.

    PubMed Central

    Chiang, G G; Schaefer, M R; Grossman, A R

    1992-01-01

    Many cyanobacteria alter their phycobilisome composition in response to changes in light wavelength in a process termed complementary chromatic adaptation. Mutant strains FdR1 and FdR2 of the filamentous cyanobacterium Fremyella diplosiphon are characterized by aberrant chromatic adaptation. Instead of adjusting to different wavelengths of light, FdR1 and FdR2 behave as if they are always in green light; they do not respond to red light. We have previously reported complementation of FdR1 by conjugal transfer of a wild-type genomic library. The complementing DNA has now been localized by genetic analysis to a region on the rescued genomic subclone that contains a gene designated rcaC. This region of DNA is also able to complement FdR2. Southern blot analysis of genomic DNA from FdR1 and FdR2 indicates that these strains harbor DNA insertions within the rcaC sequence that may have resulted from the activity of transposable genetic elements. The predicted amino acid sequence of RcaC shares strong identity to response regulators of bacterial two-component regulatory systems. This relationship is discussed in the context of the signal-transduction pathway mediating regulation of genes encoding phycobilisome polypeptides during chromatic adaptation. Images PMID:1409650

  15. Emerging and Novel Functions of Complement Protein C1q

    PubMed Central

    Kouser, Lubna; Madhukaran, Shanmuga Priyaa; Shastri, Abhishek; Saraon, Anuvinder; Ferluga, Janez; Al-Mozaini, Maha; Kishore, Uday

    2015-01-01

    Complement protein C1q, the recognition molecule of the classical pathway, performs a diverse range of complement and non-complement functions. It can bind various ligands derived from self, non-self, and altered self and modulate the functions of immune and non-immune cells including dendritic cells and microglia. C1q involvement in the clearance of apoptotic cells and subsequent B cell tolerance is more established now. Recent evidence appears to suggest that C1q plays an important role in pregnancy where its deficiency and dysregulation can have adverse effects, leading to preeclampsia, missed abortion, miscarriage or spontaneous loss, and various infections. C1q is also produced locally in the central nervous system, and has a protective role against pathogens and possible inflammatory functions while interacting with aggregated proteins leading to neurodegenerative diseases. C1q role in synaptic pruning, and thus CNS development, its anti-cancer effects as an immune surveillance molecule, and possibly in aging are currently areas of extensive research. PMID:26175731

  16. Lectin histochemical studies on the vomeronasal organ of the sheep.

    PubMed

    Ibrahim, Dalia; Nakamuta, Nobuaki; Taniguchi, Kazumi; Taniguchi, Kazuyuki

    2013-01-01

    The vomeronasal organ of sheep was examined using lectin histochemistry in order to compare the types and amounts of the glycoconjugates among various components of the vomeronasal sensory and non-sensory epithelia. In the vomeronasal sensory epithelium, Dolichos biflorus agglutinin (DBA) stained particular cells, located at the same level as the vomeronasal receptor cells, while the distribution, shape and number of the stained cells did not correspond to those of the vomeronasal receptor cells. Datura stramonium lectin (DSL), Concanavalin A (Con A), Phaseolus vulgaris agglutinin-E (PHA-E) and Phaseolus vulgaris agglutinin-L (PHA-L) labeled the basal cells of both vomeronasal sensory and non-sensory epithelia. While, Wheat germ agglutinin (WGA), Succinylated-wheat germ agglutinin (s-WGA), Lycopersicon esculentum lectin (LEL), Solanum tuberosum lectin (STL) and Ricinus communis agglutinin-I (RCA-120) labeled the basal cells of the sensory epithelium, and Bandeiraea simplicifolia lectin-I (BSL-I) stained the basal cells of the non-sensory epithelium, respectively. Seventeen lectins labeled the free border of both vomeronasal sensory and non-sensory epithelia, while Sophora japonica agglutinin (SJA), Jacalin and Pisum sativum agglutinin (PSA) labeled neither free border of the sensory nor that of non-sensory epithelia. The expression pattern of glycoconjugate was similar, but not identical, in the free border between the sensory and non-sensory epithelia. These results indicate that there are dissimilar features in the type and amount of glycoconjugates between the vomeronasal sensory and non-sensory epithelia, and at the same time, among the various cell types either in the vomeronasal sensory or non-sensory epithelium. PMID:23595118

  17. Involvement of ER stress in apoptosis induced by sialic acid-binding lectin (leczyme) from bullfrog eggs

    PubMed Central

    TATSUTA, TAKEO; HOSONO, MASAHIRO; MIURA, YUKI; SUGAWARA, SHIGEKI; KARIYA, YUKIKO; HAKOMORI, SENITIROH; NITTA, KAZUO

    2013-01-01

    Sialic-acid binding lectin (SBL) isolated from bullfrog (Rana catesbeiana) oocytes is a multifunctional protein which has lectin activity, ribonuclease activity and cancer-selective antitumor activity. It has been reported that SBL induces apoptosis accompanied by rigid mitochondrial perturbation, which indicates mediation of the intrinsic pathway. However, the mechanism of the antitumor effect of SBL has not been fully elucidated. We report, here, that ER stress is evoked in SBL-treated cells. We show that caspase-4, an initiator caspase of ER stress-mediated apoptosis was activated, and inhibition of caspase-4 resulted in significant attenuation of apoptosis induced by SBL. We analyzed the precise mechanism of activation of the caspase cascade induced by SBL, and found that caspase-9 and -4 are activated upstream of activation of caspase-8. Further study revealed that SBL induces the mitochondrial and ER stress-mediated pathways independently. It is noteworthy that SBL can induce cancer-selective apoptosis by multiple apoptotic signaling pathways, and it can serve as a candidate molecule for anticancer drugs in a novel field. PMID:24100413

  18. Ferromagnetic levan composite: an affinity matrix to purify lectin.

    PubMed

    Angeli, Renata; da Paz, Nathalia V N; Maciel, Jackeline C; Arajo, Flvia F B; Paiva, Patrcia M G; Calazans, Glcia M T; Valente, Ana Paula; Almeida, Fbio C L; Coelho, Luana C B B; Carvalho, Luiz B; Silva, Maria da Paz C; dos Santos Correia, Maria Tereza

    2009-01-01

    A simple and inexpensive procedure used magnetite and levan to synthesize a composite recovered by a magnetic field. Lectins from Canavalia ensiformis (Con A) and Cratylia mollis (Cramoll 1 and Cramoll 1, 4) did bind specifically to composite. The magnetic property of derivative favored washing out contaminating proteins and recovery of pure lectins with glucose elution. Cramoll 1 was purified by this affinity binding procedure in two steps instead of a previous three-step protocol with ammonium sulfate fractionation, affinity chromatography on Sephadex G-75, and ion exchange chromatography through a CM-cellulose column. PMID:19547713

  19. The role of complement in C3 glomerulopathy.

    PubMed

    Zipfel, Peter F; Skerka, Christine; Chen, Qian; Wiech, Thorsten; Goodship, Tim; Johnson, Sally; Fremeaux-Bacchi, Veronique; Nester, Clara; de Crdoba, Santiago Rodrguez; Noris, Marina; Pickering, Matthew; Smith, Richard

    2015-09-01

    C3 glomerulopathy describes a spectrum of disorders with glomerular pathology associated with C3 cleavage product deposition and with defective complement action and regulation (Fakhouri et al., 2010; Sethi et al., 2012b). Kidney biopsies from these patients show glomerular accumulation or deposition of C3 cleavage fragments, but no or minor deposition of immunoglobulins (Appel et al., 2005; D'Agati and Bomback, 2012; Servais et al., 2007; Sethi and Fervenza, 2011). At present the current situation asks for a better definition of the underlining disease mechanisms, for precise biomarkers, and for a treatment for this disease. The complement system is a self activating and propelling enzymatic cascade type system in which inactive, soluble plasma components are activated spontaneously and lead into an amplification loop (Zipfel and Skerka, 2009). Activation of the alternative pathway is spontaneous, occurs by default, and cascade progression leads to amplification by complement activators. The system however is self-controlled by multiple regulators and inhibitors, like Factor H that control cascade progression in fluid phase and on surfaces. The activated complement system generates a series of potent effector components and activation products, which damage foreign-, as well as modified self cells, recruit innate immune cells to the site of action, coordinate inflammation and the response of the adaptive immune system in form of B cells and T lymphocytes (Kohl, 2006; Medzhitov and Janeway, 2002; Ogden and Elkon, 2006; Carroll, 2004; Kemper and Atkinson, 2007; Morgan, 1999; Muller-Eberhard, 1986; Ricklin et al., 2010). Complement controls homeostasis and multiple reactions in the vertebrate organism including defense against microbial infections (Diaz-Guillen et al., 1999; Mastellos and Lambris, 2002; Nordahl et al., 2004; Ricklin et al., 2010). In consequence defective control of the spontaneous self amplifying cascade or regulation is associated with numerous human disorders (Ricklin and Lambris, 2007; Skerka and Zipfel, 2008; Zipfel et al., 2006). Understanding the exact action and regulation of this sophisticated homeotic cascade system is relevant to understand disease pathology of various complement associated human disorders. Furthermore this knowledge is relevant for a better diagnosis and appropriate therapy. At present diagnosis of C3 glomerulopathy is primarily based on the kidney biopsy, and histological, immmunohistological and electron microscopical evaluation (D'Agati and Bomback, 2012; Fakhouri et al., 2010; Medjeral-Thomas et al., 2014a,b; Sethi et al., 2012b). The challenge is to define the actual cause of the diverse glomerular changes or damages, to define how C3 deposition results in the reported glomerular changes, the location of the cell damage and the formation of deposits. PMID:25929733

  20. Effect of Pleurotus florida lectin (PFL) on arsenic-induced activities of splenocytes in rat.

    PubMed

    Rana, Tanmoy; Bera, Asit Kumar; Bhattacharya, Debasis; Das, Subhashree; Pan, Diganta; Das, Subrata Kumar

    2014-05-01

    The present study was undertaken to investigate the protective effect of Pleurotus florida lectin (PFL) against arsenic-induced cytotoxicity and oxidative damages in freshly isolated splenocytes of rodents. Our finding indicated that arsenic caused reduction in cell adhesion, morphological alterations, cell proliferation, nitro blue tetrazolium (NBT) index, superoxide dismutase (SOD) activity, catalase (CAT) activity and relative mRNA expression of SOD2 in relation to housekeeping gene glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and increased production of nitric oxide (NO), protein carbonyl (PC) and lipid peroxidation levels (LPO) assembled to play key factors for cytotoxicity and oxidative stress. PFL normalized cellular damages and enhanced SOD production pathway relating to gene expression. Further studies are needed to address effective phytochemicals of the edible mushroom and their mechanism. PMID:22914258

  1. Signaling by myeloid C-type lectin receptors in immunity and homeostasis

    PubMed Central

    Sancho, David; Reis e Sousa, Caetano

    2015-01-01

    Myeloid cells are key drivers of physiological responses to pathogen invasion or tissue damage. Members of the C-type lectin receptor (CLR) family stand out among the specialized receptors utilized by myeloid cells to orchestrate these responses. CLR ligands include carbohydrate, protein and lipid components of both pathogens and self, which variably trigger endocytic, phagocytic, pro-inflammatory or anti-inflammatory reactions. These varied outcomes rely on a versatile system for CLR signaling that includes tyrosine based motifs that recruit kinases, phosphatases or endocytic adaptors, as well as non-tyrosine based signals that modulate the activation of other pathways or couple to the uptake machinery. Here, we review the signaling properties of myeloid CLRs and how they impact the role of myeloid cells in innate and adaptive immunity. PMID:22224766

  2. Differential mechanisms of complement-mediated neutralization of the closely related paramyxoviruses simian virus 5 and mumps virus

    SciTech Connect

    Johnson, John B.; Capraro, Gerald A.; Parks, Griffith D.

    2008-06-20

    The complement system is an important component of the innate immune response to virus infection. The role of human complement pathways in the in vitro neutralization of three closely related paramyxoviruses, Simian Virus 5 (SV5), Mumps virus (MuV) and Human Parainfluenza virus type 2 (HPIV2) was investigated. Sera from ten donors showed high levels of neutralization against HPIV2 that was largely complement-independent, whereas nine of ten donor sera were found to neutralize SV5 and MuV only in the presence of active complement pathways. SV5 and MuV neutralization proceeded through the alternative pathway of the complement cascade. Electron microscopy studies and biochemical analyses showed that treatment of purified SV5 with human serum resulted in C3 deposition on virions and the formation of massive aggregates, but there was relatively little evidence of virion lysis. Treatment of MuV with human serum also resulted in C3 deposition on virions, however in contrast to SV5, MuV particles were lysed by serum complement and there was relatively little aggregation. Assays using serum depleted of complement factors showed that SV5 and MuV neutralization in vitro was absolutely dependent on complement factor C3, but was not dependent on downstream complement factors C5 or C8. Our results indicate that even though antibodies exist that recognize both SV5 and MuV, they are mostly non-neutralizing and viral inactivation in vitro occurs through the alternative pathway of complement. The implications of our work for development of paramyxovirus vectors and vaccines are discussed.

  3. Complement fixation test to C. burnetii

    MedlinePLUS

    ... a laboratory where it is examined for Coxiella antibodies using a laboratory method called complement fixation. This ... checks if the body has produced substances called antibodies to a specific foreign substance (antigen), in this ...

  4. Efficacy of Targeted Complement Inhibition in Experimental C3 Glomerulopathy.

    PubMed

    Ruseva, Marieta M; Peng, Tao; Lasaro, Melissa A; Bouchard, Keith; Liu-Chen, Susan; Sun, Fang; Yu, Zhao-Xue; Marozsan, Andre; Wang, Yi; Pickering, Matthew C

    2016-02-01

    C3 glomerulopathy refers to renal disorders characterized by abnormal accumulation of C3 within the kidney, commonly along the glomerular basement membrane (GBM). C3 glomerulopathy is associated with complement alternative pathway dysregulation, which includes functional defects in complement regulator factor H (FH). There is no effective treatment for C3 glomerulopathy. We investigated the efficacy of a recombinant mouse protein composed of domains from complement receptor 2 (CR2) and FH (CR2-FH) in two models of C3 glomerulopathy with either preexisting or triggered C3 deposition along the GBM. FH-deficient mice spontaneously develop renal pathology associated with abnormal C3 accumulation along the GBM and secondary plasma C3 deficiency. CR2-FH partially restored plasma C3 levels in FH-deficient mice 2 hours after intravenous injection. CR2-FH specifically targeted glomerular C3 deposits, reduced the linear C3 reactivity assessed with anti-C3 and anti-C3b/iC3b/C3c antibodies, and prevented further spontaneous accumulation of C3 fragments along the GBM. Reduction in glomerular C3d and C9/C5b-9 reactivity was observed after daily administration of CR2-FH for 1 week. In a second mouse model with combined deficiency of FH and complement factor I, CR2-FH prevented de novo C3 deposition along the GBM. These data show that CR2-FH protects the GBM from both spontaneous and triggered C3 deposition in vivo and indicate that this approach should be tested in C3 glomerulopathy. PMID:26047789

  5. Polyanion-Induced Self Association of Complement Factor H1

    PubMed Central

    Pangburn, Michael K.; Rawal, Nenoo; Cortes, Claudio; Alam, M. Nurul; Ferreira, Viviana P.; Atkinson, Mark A. L.

    2008-01-01

    Factor H is the primary soluble regulator of activation of the alternative pathway of complement. It prevents activation of complement on host cells and tissues upon association with C3b and surface polyanions such as sialic acids, heparin and other glycosaminoglycans. Here we show that interaction with polyanions causes self-association forming tetramers of the 155,000 Da glycosylated protein. Monomeric human factor H is an extended flexible protein that exhibits an apparent size of 330,000 Da, relative to globular standards, during gel filtration chromatography in the absence of polyanions. In the presence of dextran sulfate (5,000 Da) or heparin an intermediate species of apparent m.w. 700,000 and a limit species of m.w. 1,400,000 were observed by gel filtration. Sedimentation equilibrium analysis by analytical ultracentrifugation indicated a monomer Mr of 163,000 in the absence of polyanions and a Mr of 607,000, corresponding to a tetramer, in the presence of less than a 2-fold molar excess of dextran sulfate. Increasing concentrations of dextran sulfate increased binding of factor H to zymosan-C3b 4.5-fold. This was accompanied by an increase in both the decay accelerating and cofactor activity of factor H on these cells. An expressed fragment encompassing the C-terminal polyanion binding site (complement control protein domains 1820) also exhibited polyanion-induced self association, suggesting that the C-terminal ends of factor H mediate self-association. The results suggest that recognition of polyanionic markers on host cells and tissues by factor H, and the resulting regulation of complement activation, may involve formation of dimers and tetramers of factor H. PMID:19124749

  6. Altered expression of sialylated glycoproteins in ovarian cancer sera using lectin-based ELISA assay and quantitative glycoproteomics analysis.

    PubMed

    Wu, Jing; Xie, Xiaolei; Nie, Song; Buckanovich, Ronald J; Lubman, David M

    2013-07-01

    Herein, we identify and confirm differentially expressed sialoglycoproteins in the serum of patients with ovarian cancer. On the basis of Sambucus nigra (SNA) lectin enrichment and on an isobaric chemical labeling quantitative strategy, clusterin (CLUS), leucine-rich alpha-2-glycoprotein (LRG1), hemopexin (HEMO), vitamin D-binding protein (VDB), and complement factor H (CFH) were found to be differentially expressed in the serum of patients with ovarian cancer compared to benign diseases. The abnormal sialylation levels of CLUS, CFH, and HEMO in serum of ovarian cancer patients were verified by a lectin-based ELISA assay. ELISA assays were further applied to measure total protein level changes of these glycoproteins. Protein levels of CLUS were found to be down-regulated in the serum of ovarian cancer patients, while protein levels of LRG1 were increased. The combination of CLUS and LRG1 (AUC = 0.837) showed improved performance for distinguishing stage III ovarian cancer from benign diseases compared to CA125 alone (AUC = 0.811). In differentiating early stage ovarian cancer from benign diseases or healthy controls, LRG1 showed comparable performance to CA125. An independent sample set was further used to confirm the ability of these candidate markers to detect patients with ovarian cancer. Our study provides a comprehensive strategy for the identification of candidate biomarkers that show the potential for diagnosis of ovarian cancer. Further studies using a large number of samples are necessary to validate the utility of this panel of proteins. PMID:23731285

  7. Evolution of the complement system in protostomes revealed by de novo transcriptome analysis of six species of Arthropoda.

    PubMed

    Sekiguchi, Reo; Nonaka, Masaru

    2015-05-01

    To elucidate the evolutionary history of the complement system in Arthropoda, de novo transcriptome analysis was performed with six species among the Chelicerata, Myriapoda, and Crustacea, and complement genes were identified based on their characteristic domain structures. Complement C3 and factor B (FB) were identified from a sea spider, a jumping spider, and a centipede, but not from a sea firefly or two millipede species. No additional complement components identifiable by their characteristic domain structures were found from any of these six species. These results together with genome sequence information for several species of the Hexapoda suggest that the common ancestor of the Arthropoda possessed a simple complement system comprising C3 and FB, and thus resembled the alternative pathway of the mammalian complement system. It was lost at least twice independently during the evolution of Arthropoda in the millipede lineage and in the common ancestor of Crustacea and Hexapoda. PMID:25530095

  8. Retinal Pigment Epithelial Cell Death by the Alternative Complement Cascade: Role of Membrane Regulatory Proteins, Calcium, PKC, and Oxidative Stress

    PubMed Central

    Yang, Ping; Baciu, Peter; Kerrigan, Brittany C. Parker; Etheridge, Menna; Sung, Eric; Toimil, Brett A.; Berchuck, Jacob E.; Jaffe, Glenn J.

    2014-01-01

    Purpose. Retinal pigment epithelial (RPE) cell death is an important feature of the advanced forms of AMD. Complement alternative pathway (AP) activation is associated with RPE cell death in AMD. In this study, we developed a new model to initiate AP activation on RPE cells and investigated the cellular mechanisms modulating AP activationmediated RPE cell death. Methods. An anti-RPE antibody was developed. A spontaneously arising human RPE cell line (ARPE-19) and donor RPE cells were primed with this antibody followed by stimulation with 6% C1q-depleted human serum (C1q-Dep) to activate AP. Complement activation was evaluated by flow cytometry and immunofluorescent staining. Cellular response to complement activation was examined by measurement of intracellular calcium and adenosine triphosphate (ATP) release. Cell viability was assessed by Sytox orange, tetrazolium salt, and lactate dehydrogenase release assays. Results. Alternative pathway complementmediated RPE cell death was associated with membrane attack complex formation and a rapid rise in intracellular calcium followed by release of ATP. Downregulation of membrane complement regulatory proteins and protein kinase C (PKC) inhibition increased cell susceptibility to complement attack. Pretreatment of RPE cells with either hydrogen peroxide or hydroquinone enhanced cell death. Chronic repetitive treatment of RPE cells with low levels of oxidants also enhanced complement-mediated cell death. Conclusions. Activation of complement through the alternative pathway induces sublytic and lytic phases of complement attack on RPE cells, leading to cell death modulated by extracellular calcium, membrane complement regulatory proteins, and intracellular signaling mechanisms. Single-dose oxidant exposure and low-dose repetitive oxidant exposure rendered RPE cells more susceptible to complement-mediated death. PMID:24677108

  9. Lectin-binding sites in epithelial cells of the mouse prostate gland.

    PubMed

    Sakuda, Kentaro; Yoshida, Ayaka; Muragishi, Ryoki; Yoshinaga, Kazuya

    2014-01-01

    The prostate is an exocrine gland in the male reproductive tract that secretes seminal fluids. To gain insight into the cytochemical properties of prostatic epithelial cells, the characteristics of glycoconjugates in mouse prostate sections were examined by lectin histochemistry and immunohistochemistry. Characteristic staining patterns were observed, depending on the type of lectins present in the epithelia. Luminal cells reacted specifically with mannose-binding lectins (Galanthus nivalis lectin, Hippeastrum hybrid lectin, Narcissus pseudonarcissus lectin) and Maclura pomifera lectin in all lobes of the prostate. Luminal cells also expressed galactose, N-acetyl-D-galactosamine (GalNAc), N-acetyl-D-glucosamine (GlcNAc), and fucose residues in the lateral and ventral lobes. Basal cells expressed GlcNAc and fucose, and reacted with Datura stramonium lectin and Aleuria aurantia lectin in all lobes. These results indicate that in the mouse prostate, the selectivity of lectin-binding sites for distinct cell types and lobe-dependent staining may relate to cellular and regional differences in function. Furthermore, some lectins selectively bound to prostatic epithelial cells, indicating their potential use as markers for the histopathological evaluation of prostatic diseases, cancer diagnosis, or male infertility. PMID:26004072

  10. A simple strategy for the creation of a recombinant lectin microarray.

    PubMed

    Hsu, Ku-Lung; Gildersleeve, Jeffrey C; Mahal, Lara K

    2008-06-01

    Glycomics, i.e. the high-throughput analysis of carbohydrates, has yet to reach the level of ease and import of its counterparts, genomics and proteomics, due to the difficulties inherent in carbohydrate analysis. The advent of lectin microarray technology addresses many of these problems, providing a straightforward approach for glycomic analysis. However, current microarrays are limited to the available lectin set, which consists mainly of plant lectins isolated from natural sources. These lectins have inherent problems including inconsistent activity and availability. Also, many plant lectins are glycosylated, complicating glycomic evaluation of complex samples, which may contain carbohydrate-binding proteins. The creation of a recombinant, well-defined lectin set would resolve many of these issues. Herein, we describe an efficient strategy for the systematic creation of recombinant lectins for use in microarray technology. We present a small panel of simple-to-purify bacterially-derived lectins that show reliable activity and define their binding specificities by both carbohydrate microarray and ELISA. We utilize this panel to create a recombinant lectin microarray that is able to distinguish glycopatterns for both proteins and cell samples. This work opens the door to the establishment of a vast set of defined lectins via high-throughout approaches, advancing lectin microarray technology for glycomic analysis. PMID:18493664

  11. The use of lectin microarray for assessing glycosylation of therapeutic proteins.

    PubMed

    Zhang, Lei; Luo, Shen; Zhang, Baolin

    2016-04-01

    Glycans or carbohydrates attached to therapeutic glycoproteins can directly affect product quality, safety and efficacy, and therefore must be adequately analyzed and controlled throughout product life cycles. However, the complexity of protein glycosylation poses a daunting analytical challenge. In this study, we evaluated the utility of a lectin microarray for assessing protein glycans. Using commercial lectin chips, which contain 45 lectins toward distinct glycan structures, we were able to determine the lectin binding patterns of a panel of 15 therapeutic proteins, including 8 monoclonal antibodies. Lectin binding signals were analyzed to generate glycan profiles that were generally consistent with the known glycan patterns for these glycoproteins. In particular, the lectin-based microarray was found to be highly sensitive to variations in the terminal carbohydrate structures such as galactose versus sialic acid epitopes. These data suggest that lectin microarray could be used for screening glycan patterns of therapeutic glycoproteins. PMID:26918373

  12. A comparison of the carbohydrate binding properties of two Dolichos biflorus lectins.

    PubMed

    Etzler, M E

    1994-10-01

    The carbohydrate binding properties of the Dolichos biflorus seed lectin and DB58, a vegetative tissue lectin from this plant, were compared using two types of solid phase assays. Both lectins bind to hog blood group A + H substance covalently coupled to Sepharose 4B and this binding can be inhibited with free blood group A + H substance. However, the binding of the seed lectin is inhibited by D-GalNAc whereas DB58 binding was not inhibited by any monosaccharide tested, thus suggesting that its carbohydrate combining site may be more extensive than that of the seed lectin. The activities of these two lectins also differ from one another in ability to recognize blood group A + H substance adsorbed on to plastic and in the effects of salt and urea on their carbohydrate binding activities. Neither lectin showed glycosidase activity with p-nitrophenyl alpha-D-GalNAc or p-nitrophenyl beta-D-GalNAc. PMID:7696843

  13. Purification and characterization of mannose/glucose-specific lectin from seeds of Trigonella foenumgraecum.

    PubMed

    Naeem, Aabgeena; Ahmad, Ejaz; Ashraf, Mohd Tashfeen; Khan, Rizwan Hasan

    2007-01-01

    A lectin present in seeds of Trigonella foenumgraecum was isolated and purified by acid precipitation, salt fractionation, and affinity chromatography on mannan cross-linked agarose. SDS-PAGE revealed a single band corresponding to a molecular weight of 27,350 daltons. The lectin agglutinated trypsin-treated rat erythrocytes. Sugar specificity as determined by hemagglutination inhibition assay indicated that the lectin belongs to a glucose/mannose-specific group. The reaction of the lectin with glycoprotein was affected by pH changes. The carbohydrate binding specificity of the lectin was investigated by turbidity and activity measurements. As the lectin belongs to the Leguminoceae family, the specificity of the lectin for glucose/mannose renders it a valuable tool for Rhizobium-legume symbiosis. PMID:17309436

  14. Distribution of Lectins in Tissues, Derived Callus, and Roots of Psophocarpus tetragonolobus (Winged Bean).

    PubMed

    Meimeth, T; Van, K T; Marcotte, J L; Trinn, T H; Clarke, A E

    1982-08-01

    The distribution of lectin in parental tissues, roots formed de novo from parental stem tissue, and derived callus cells of Psophocarpus tetragonolobus has been measured by hemagglutinating activity and radioimmunoassay. The antisera used for the radioimmunoassay was raised in rabbits to lectin isolated from seeds by affinity chromatography using insolubilized hog gastric mucin. The distribution of lectin in buffer extracts of the tissues (or cells) and the extracellular medium favors the tissues for in vitro grown roots, regardless of the culture conditions used. The lectin content of the extracellular medium is more significant for callus, regardless of its conditions of culture. The lectin activity of extracts of in vitro grown roots was higher than that of mature roots from whole plants. Differences in relative levels of lectin activity measured by hemagglutination and by radioimmunoassay, and differences in saccharide inhibition of hemagglutination, suggest the presence of multiple lectins in extracts of different tissues. PMID:16662537

  15. Structures and binding specificity of galactose- and mannose-binding lectins from champedak: differences from jackfruit lectins.

    PubMed

    Gabrielsen, Mads; Abdul-Rahman, Puteri Shafinaz; Othman, Shatrah; Hashim, Onn H; Cogdell, Richard J

    2014-06-01

    Galactose-binding and mannose-binding lectins from the champedak fruit, which is native to South-east Asia, exhibit useful potential clinical applications. The specificity of the two lectins for their respective ligands allows the detection of potential cancer biomarkers and monitoring of the glycosylated state of proteins in human serum and/or urine. To fully understand and expand the use of these natural proteins, their complete sequences and crystal structures are presented here, together with details of sugar binding. PMID:24915077

  16. Carbohydrate specificity and quaternary association in basic winged bean lectin: X-ray analysis of the lectin at 2.5 A resolution.

    PubMed

    Prabu, M M; Sankaranarayanan, R; Puri, K D; Sharma, V; Surolia, A; Vijayan, M; Suguna, K

    1998-03-01

    The structure of basic Winged Bean Agglutinin (WBAI) with two dimeric molecules complexed with methyl-alpha-D-galactopyranoside in the asymmetric unit, has been determined by the molecular replacement method and refined with 2.5 A X-ray intensity data. The polypeptide chain of each monomer has the characteristic legume lectin tertiary fold. The structure clearly defines the lectin-carbohydrate interactions. It reveals how the unusually long variable loop in the binding region endows the lectin with its characteristic sugar specificity. The lectin forms non-canonical dimers of the type found in Erythrina corallodendron lectin (EcorL) even though glycosylation, unlike in EcorL, does not prevent the formation of canonical dimers. The structure thus further demonstrates that the mode of dimerisation of legume lectins is not necessarily determined by the covalently bound carbohydrate but is governed by features intrinsic to the protein. The present analysis and our earlier work on peanut lectin (PNA), show that legume lectins are a family of proteins in which small alterations in essentially the same tertiary structure lead to wide variations in quaternary association. A relationship among the non-canonical modes of dimeric association in legume lectins is presented. PMID:9500920

  17. Inhibition of aberrant complement activation by a dimer of acetylsalicylic acid.

    PubMed

    Lee, Moonhee; Wathier, Matthew; Love, Jennifer A; McGeer, Edith; McGeer, Patrick L

    2015-10-01

    We here report synthesis for the first time of the acetyl salicylic acid dimer 5,5'-methylenebis(2-acetoxybenzoic acid) (DAS). DAS inhibits aberrant complement activation by selectively blocking factor D of the alternative complement pathway and C9 of the membrane attack complex. We have previously identified aurin tricarboxylic and its oligomers as promising agents in this regard. DAS is much more potent, inhibiting erythrocyte hemolysis by complement-activated serum with an IC50 in the 100-170 nanomolar range. There are numerous conditions where self-damage from the complement system has been implicated in the pathology, including such chronic degenerative diseases of aging as Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, and age-related macular degeneration. Consequently, there is a high priority for the discovery and development of agents that can successfully treat such conditions. DAS holds considerable promise for being such an agent. PMID:26248865

  18. Recruitment of Factor H as a Novel Complement Evasion Strategy for Blood-Stage Plasmodium falciparum Infection.

    PubMed

    Kennedy, Alexander T; Schmidt, Christoph Q; Thompson, Jennifer K; Weiss, Greta E; Taechalertpaisarn, Tana; Gilson, Paul R; Barlow, Paul N; Crabb, Brendan S; Cowman, Alan F; Tham, Wai-Hong

    2016-02-01

    The human complement system is the frontline defense mechanism against invading pathogens. The coexistence of humans and microbes throughout evolution has produced ingenious molecular mechanisms by which microorganisms escape complement attack. A common evasion strategy used by diverse pathogens is the hijacking of soluble human complement regulators to their surfaces to afford protection from complement activation. One such host regulator is factor H (FH), which acts as a negative regulator of complement to protect host tissues from aberrant complement activation. In this report, we show that Plasmodium falciparum merozoites, the invasive form of the malaria parasites, actively recruit FH and its alternative spliced form FH-like protein 1 when exposed to human serum. We have mapped the binding site in FH that recognizes merozoites and identified Pf92, a member of the six-cysteine family of Plasmodium surface proteins, as its direct interaction partner. When bound to merozoites, FH retains cofactor activity, a key function that allows it to downregulate the alternative pathway of complement. In P. falciparum parasites that lack Pf92, we observed changes in the pattern of C3b cleavage that are consistent with decreased regulation of complement activation. These results also show that recruitment of FH affords P. falciparum merozoites protection from complement-mediated lysis. Our study provides new insights on mechanisms of immune evasion of malaria parasites and highlights the important function of surface coat proteins in the interplay between complement regulation and successful infection of the host. PMID:26700768

  19. Factor H in Porcine Seminal Plasma Protects Sperm against Complement Attack in Genital Tracts*

    PubMed Central

    Sakaue, Tomohisa; Takeuchi, Keisuke; Maeda, Toshinaga; Yamamoto, Yoshio; Nishi, Katsuji; Ohkubo, Iwao

    2010-01-01

    We found that factor H (FH) exists in porcine seminal plasma. Purified FH strongly inhibited serum alternative pathway complement activation against lipopolysaccharide. The molecular weight, pI, and heparin-binding activity of the purified protein were different from those of purified FH from porcine serum. The complement regulatory activity of seminal plasma FH was ∼2-fold stronger than that of serum FH. Treatment of purified serum FH with sialidase and N-glycosidase F gave almost the same results as those of seminal plasma FH. The deletion of sialic acid from the carbohydrate chains of both FHs contributed to heparin-binding and complement regulatory activities. Results of reverse transcriptase-PCR, Western blot analysis, and immunohistochemistry showed that seminal plasma FH is mainly secreted from epithelial cells of the seminal vesicle in male genital tracts. FH was also detected in the outer acrosomal region of ejaculated sperm by immunofl