Sialic Acids

but much less in those that have reached the olfactory bulb. We show that two of the naturally occurring Notch1 activators, Jagged1 and Delta1, are also expressed in the SVZ and within the RMS in the adult mouse brain. Finally, using a model of cortical stab wound, we show that the astrogliogenic response of the SVZ to injury is accompanied by activation of the Notch pathway.

Reported elevations of total serum sialic acid may be a result of shed tumour-related membrane sialylglycoprotein and/or concurrent elevation of nonspecific, acute-phase reactant sialylglycoprotein. To clarify further the specificity and sensitivity of serum sialic acid monitoring, analyses of sialic acid by the thiobarbituric acid method and acute-phase reactants by radial immunodiffusion were made using the same malignant melanoma patients' sera. Preliminary studies of IgG, IgA, IgM, ceruloplasmin and C-reactive protein suggested that these would not be valuable monitors of tumour burden. Single serum samples from 59 melanoma patients and age-and sex-matched controls were further examined for sialic acid, a -acid glycoprotein, a -antitrypsin, haptoglobin, and a2-macroglobulin. Patients were grouped according to tumour burden. In pairwise statistical tests, differences between groups tended to be greater for sialic acid than for acute-phase reactants. On discriminant analysis, sialic acid was clearly the most significant single discriminator between groups, with an F statistic of P < 0-00005. Although a, -acid glycoprotein was quite strongly correlated with sialic acid, it was not such a good discriminator and did not add significantly to the predictive power of sialic acid alone.

Over four decades ago, specific tumor characteristics were ascribed to the increased expression of sialic acid sugars on the surface of cancer cells, and this led to the definition of sialic acids as potential therapeutic targets. Recent advances in glycobiology and cancer research have defined the key processes underlying aberrant expression of sialic acids in cancer, and its consequences, more precisely. These consequences include effects on tumor growth, escape from apoptosis, metastasis formation, and resistance to therapy. Collectively, these novel insights provide further rationale for the design and development of therapeutic approaches that interfere with excessively high expression of sialic acids in cancer cells. Strategies to target aberrant sialylation in cancer, however, have evolved comparatively slowly. Here, we review recent findings that emphasize the detrimental effects of hypersialylation on multiple aspects of tumor growth and behavior. We also discuss novel therapeutic strategies.

Influenza virus remains among the most important pathogens infecting elderly people. Vaccination is the most cost-effective strategy to reduce morbidity and mortality due to influenza. For persons who are not vaccinated or for whom vaccines fail to prevent influenza, there are 2 classes of efficacious drugs for treatment or chemoprophylaxis: M2 channel inhibitors and neuraminidase inhibitors. Effective treatment, however, must commence within 48 h of the onset of symptoms, which can create problems for patients who wait to see whether their symptoms worsen or improve. Older adults who have relocated to the congregate housing environments of assisted living and long-term care facilities deserve special consideration, because influenza exposure risks are different for this group. Strategies for control of influenza must combine preventive approaches, such as vaccination, educational approaches, and the introduction of policies that allow health care professionals anticipate, identify, and efficiently respond to influenza outbreaks.

Novel 3,4-disubstituted-Neu5Ac2en derivatives have been synthesised to probe the open 150-loop conformation of influenza virus sialidases. Both equatorially and axially (epi) substituted C4 amino and guanidino 3-(p-tolyl)allyl-Neu5Ac2en derivatives were prepared, via the 4-epi-hydroxy derivative. The equatorially-substituted 4-amino derivative showed low micromolar inhibition of both group-1 (pdm09 H1N1) and group-2 (pdm57 H2N2) sialidases, and provides the first in vitro evidence that a group-2 sialidase may exhibit 150-loop flexibility.

Stem cell lines that provide a renewable and scaleable supply of central nervous system cell types would constitute an invaluable resource for basic and applied neurobiology. Here we describe the generation and long-term expansion of multiple human foetal neural stem (NS) cell lines in monolayer culture without genetic immortalization. Adherent human NS cells are propagated in the presence of epidermal growth factor (EGF) and fibroblast growth factor 2 (FGF2), under which conditions they stably express neural precursor markers and exhibit negligible differentiation into neurons or glia. However, they produce astrocytes, oligodendrocytes, and neurons upon exposure to appropriate differentiation factors. Single cell cloning demonstrates that human NS cells are tripotent. They retain a diploid karyotype and constant neurogenic capacity after over 100 generations. In contrast to human neurospheres, we observe no requirement for the cytokine leukaemia inhibitory factor (LIF) for continued expansion of adherent human NS cells. Human NS cells can be stably transfected to provide reporter lines and readily imaged in live monolayer cultures, creating the potential for high content genetic and chemical screens.

A sTn double C-glycoside, sTn analogue 2, was synthesized using samarium chemistry developed in our laboratory. Complications in the oxidation reaction affording aldehyde acceptor were overcome by double protection of amide and the use of a room-temperature ionic liquid as solvent. Studies are underway to conjugate the sTn double C-glycoside hapten 2 to KLH carrier protein for biological evaluation as a vaccine.

The structural-reporter-group resonances in the 'H-n.m.r. spectra of a series of ganglioside-derived oligosaccharides are completely assigned. The three different types of sialic acid residues which may occur in these compounds could be adequately characterized. They are distinguishable on the basis of the set of ..chemical shifts of their H-3a and H-3e atoms. The mutual influence of GalNAc #I-(1+4) and NeuSAc (y-(2+3), both linked to the same Gal residue, on the 'H-n.m.r. parameters of each other, reflects a well-defined solution conformation, which is held to be responsible for the resistance of this "internal" sialic acid towards various sialidases.

Paramyxoviruses are the leading cause of respiratory disease in children. Several paramyxoviruses possess a surface glycoprotein, the hemagglutinin-neuraminidase (HN), that is involved in attachment to sialic acid receptors, promotion of fusion, and removal of sialic acid from infected cells and progeny virions. Previously we showed that Newcastle disease virus (NDV) HN contained a pliable sialic acid recognition site that could take two states, a binding state and a catalytic state. Here we present evidence for a second sialic acid binding site at the dimer interface of HN and present a model for its involvement in cell fusion. Three different crystal forms of NDV HN now reveal identical tetrameric arrangements of HN monomers, perhaps indicative of the tetramer association found on the viral surface.

Zanamivir (4-guanidino-Neu5Ac2en [4-GU-DANA]) inhibits not only the neuraminidase activity but also the receptor interaction of the human parainfluenza virus type 3 (HPIV3) hemagglutinin-neuraminidase (HN), blocking receptor binding and subsequent fusion promotion. All activities of the HPIV3 variant ZM1 HN (T193I/I567V) are less sensitive to 4-GU-DANA's effects. The T193I mutation in HN confers both increased receptor binding and increased neuraminidase activity, as well as reduced sensitivities of both activities to 4-GU-DANA inhibition, consistent with a single site on the HN molecule carrying out both catalysis and binding. We now provide evidence that the HPIV3 variant's resistance to receptor-binding inhibition by 4-GU-DANA is related to a reduced affinity of the HN receptor-binding site for this compound as well as to an increase in the avidity of HN for the receptor. Newcastle disease virus (NDV) HN and HPIV3 HN respond differently to inhibition in ways that suggest a fundamental distinction between them. NDV HN-receptor binding is less sensitive than HPIV3 HN-receptor binding to 4-GU-DANA, while its neuraminidase activity is highly sensitive. Both HPIV3 and NDV HNs are sensitive to receptor-binding inhibition by the smaller molecule DANA. However, for NDV HN, some receptor binding cannot be inhibited. These data are consistent with the presence in NDV HN of a second receptor-binding site that is devoid of enzyme activity and has a negligible, if any, affinity for 4-GU-DANA. Avidity for the receptor contributes to resistance by allowing the receptor to compete effectively with inhibitors for interaction with HN, while the further determinant of resistance is the reduced binding of the inhibitor molecule to the binding pocket on HN. Based upon our data and recent three-dimensional structural information on the HPIV3 and NDV HNs, we propose mechanisms for the observed sensitivity and resistance of HN to receptor-binding inhibition and discuss the implications of these mechanisms for the distribution of HN functions.

A number of polyamine (PA) derivatives of thiosemicarbazone of 1,3-dichloroacetone (TDA) have been prepared and their effect on growth in vivo of tumorigenic but not metastatic cell strain (LY-R) of mouse lymphoma L5178Y has been investigated. Polyamine derivatives of TDA (PDT) were injected i.p. every third day (4 times, 10 or 25 mg/kg per injection) into DBA/2 mice inoculated i.p. or s.c. with LY-R cells. It has been found that disubstituted putrescine (Put), spermidine (Spd) and spermine (Spin) derivatives TDA exhibit a prometastatic activity as indicated by the appearance of solid tumor foci in subcutaneous tissues, liver and spleen. This activity depends mainly on the structure of the PA fragment and the presence of TDA. An increase in lipid bound sialic acid content after treating LY-R cells in vitro and in vivo with a Spin derivative has been found. These findings suggest that disubstituted PA derivatives of TDA and LY-R cells may be a useful model for investigation of the final steps in formation of metastases by lymphoma cells.

We report on 2 unrelated patients with Costello syndrome. The first is a 5-year-old girl with "coarse" face, nasal papillomata, redundant skin of feet and hands, hyperextensible hand and finger joints, curly hair, feeding problems due to oral motor apraxia, growth and psychomotor retardation. The second is a 3-year-old boy with "coarse" face, loose skin on hands and feet, curly hair, oral motor apraxia, severe growth and psychomotor retardation. In both patients urine sialic acid levels were found to be repeatedly high. The meaning of this biochemical abnormality is discussed.

Glycans attached to the cell surface via proteins or lipids or exposed in the extracellular matrix affect many cellular processes, including neuritogenesis, cell survival and migration, as well as synaptic activity and plasticity. These functions make glycans attractive molecules for stimulating repair of the injured nervous system. Yet, glycans are often difficult to synthesize or isolate and have the disadvantage to be unstable in a complex tissue environment. To circumvent these issues, we have screened a library of small organic compounds to search for structural and functional mimetics of the neurostimulatory glycan polysialic acid (PSA) and identified the 5-HT 4 receptor agonist tegaserod as a PSA mimetic. The PSA mimicking activity of tegaserod was shown in cultures of central and peripheral nervous system cells of the mouse and found to be independent of its described function as a serotonin (5-HT 4) receptor agonist. In an in vivo model for peripheral nerve regeneration, mice receiving tegaserod at the site of injury showed enhanced recovery compared to control mice receiving vehicle control as evidenced by functional measurements and histology. These data indicate that tegaserod could be repurposed for treatment of nervous system injuries and underscores the potential of using small molecules as mimetics of neurostimulatory glycans. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Taipoxin (taipan toxin), purified from the venom of the Australian taipan (Oxyuranus s. scutellutus) by gel filtration on Sephadex G-75 followed by column zone electrophoresis, is the most lethal neurotoxin yet isolated from any snake venom. The LDso is 2 pg/kg in the mouse. The main physiological effect is a gradual reduction to complete stop of evoked and spontaneous release of acetylcholine from motor nerve terminals. Intoxicated animals die of asphyxia caused by neuromuscular blockage of the respiratory muscles.

In 2007, Martin et al. developed a method for the analysis of sialic acids by HPLC following 1,2-diamino-4,5-methylenedioxybenzene (DMB) derivatisation (Martín et al., Anal Bioanal Chem 387:2943-2949. Within the article, the authors noted that lactose interfered with the analysis, giving erroneously high results when lactosecontaining products were analysed. Such an observation is important when analysing milk-based products, yet was contradictory to the observations of Nakamura et al. (Chem Pharm Bull 35(2):687-692, 1987) who demonstrated that DMB was specific for α-keto acids and did not react with simple sugars such as glucose or lactose. In order to clarify the situation, this phenomenon was investigated and it was confirmed that lactose does not interfere with the analysis. However, it was found that most commercial preparations of lactose do contain small amounts of sialic acids, either as the free monosaccharide or bound to lactose in the form of 3′and 6′-sialyllactose.

The specificity anti-Gd of human cold autoagglutinins is characterized using untreated and enzyme-treated human red blood cells. Gd determinants of human RBC are resistant to proteases, but are inactivated by neuraminidase (RDE). In contrast, I/i determinants are not inactivated by proteases or RDE, while Prl-3 determinants are inactivated by proteases and RDE, and Pra determinants are resistant to RDE, but are inactivated by proteases.

A ceramide hexasaccharide was purified from the gangliosides of human kidney. Its structure was studied by methylation analysis, neuraminidase treatment, weak acid hydrolysis and chromium trioxide oxidation. The structure is suggested to be a disialosyl derivative of paragloboside: formula: (see text) Data indicating the possible existence of a corresponding trisialosyl derivative are also presented.

We report here that human T cells give much stronger proliferative responses to specific activation via the T cell receptor (TCR) than those from chimpanzees, our closest evolutionary relatives. Nonspecific activation using phytohemagglutinin was robust in chimpanzee T cells, indicating that the much lower response to TCR simulation is not due to any intrinsic inability to respond to an activating stimulus. CD33-related Siglecs are inhibitory signaling molecules expressed on most immune cells and are thought to down-regulate cellular activation pathways via cytosolic immunoreceptor tyrosine-based inhibitory motifs. Among human immune cells, T lymphocytes are a striking exception, expressing little to none of these molecules. In stark contrast, we find that T lymphocytes from chimpanzees as well as the other closely related "great apes" (bonobos, gorillas, and orangutans) express several CD33-related Siglecs on their surfaces. Thus, human-specific loss of T cell Siglec expression occurred after our last common ancestor with great apes, potentially resulting in an evolutionary difference with regard to inhibitory signaling. We confirmed this by studying Siglec-5, which is prominently expressed on chimpanzee lymphocytes, including CD4 T cells. Ab-mediated clearance of Siglec-5 from chimpanzee T cells enhanced TCR-mediated activation. Conversely, primary human T cells and Jurkat cells transfected with Siglec-5 become less responsive; i.e., they behave more like chimpanzee T cells. This human-specific loss of T cell Siglec expression associated with T cell hyperactivity may help explain the strikingly disparate prevalence and severity of T cell-mediated diseases such as AIDS and chronic active hepatitis between humans and chimpanzees. chimpanzees | HIV/AIDS | T cells

A method for the assay of neuraminidase in human cultured fibroblasts has been worked out. The substrates, all naturally occurring, were: sialyloligosaccharides (a(2 ~ 3)sialyUactose, a(2-~ 6)sialyllactose, disialyllactose), sialylglycolipids (disialogangliosides GDla and GDlb), sialylglycoproteins and sialylglycopeptides (ovine submaxillary glycoprotein and its pronase-glycopeptides). The method was based on the determination of the enzymically liberated N-acetylneuraminic acid (NeuAc) by a chromatographic-colorimetric microprocedure.

Dimeric derivatives (compounds 7 to 9) of the influenza virus neuraminidase inhibitor zanamivir (compound 2), which have linking groups of 14 to 18 atoms in length, are approximately 100-fold more potent inhibitors of influenza virus replication in vitro and in vivo than zanamivir. The observed optimum linker length of 18 to 22 Å, together with observations that the dimers cause aggregation of isolated neuraminidase tetramers and whole virus, indicate that the dimers benefit from multivalent binding via intertetramer and intervirion linkages. The outstanding long-lasting protective activities shown by compounds 8 and 9 in mouse influenza infectivity experiments and the extremely long residence times observed in the lungs of rats suggest that a single low dose of a dimer would provide effective treatment and prophylaxis for influenza virus infections.

The O-specificity of cell wall lipopolysaccharides (LPS) is due to their polysaccharide moiety'. Whereas the O-specific polysaccharides are neutral in most genera, such as Klebsiella or Salmonella, they are negatively charged in a number of Escherichia coli strains ',* Mostly, the negative charge in these O-specific .

Sialic acids, or the more broad term nonulosonic acids, comprise a family of nine-carbon keto-sugars ubiquitous on mammalian mucous membranes as terminal modifications of mucin glycoproteins. Sialic acids have a limited distribution among bacteria, and the ability to catabolize sialic acids is mainly confined to pathogenic and commensal species. This ability to utilize sialic acid as a carbon source is correlated with bacterial virulence, especially, in the sialic acid rich environment of the oral cavity, respiratory, intestinal, and urogenital tracts. This chapter discusses the distribution of sialic acid catabolizers among the sequenced bacterial genomes and examines the studies that have linked sialic acid catabolism with increased in vivo fitness in a number of species using several animal models. This chapter presents the most recent findings in sialobiology with a focus on sialic acid catabolism, which demonstrates an important relationship between the catabolism of sialic acid and bacterial pathogenesis.

Peptide and protein drugs are a growing class of therapeutics. However, their effective application in the clinic is compromised by problems, for instance proteolysis in the circulating blood, premature clearance through the kidneys, and immunogenicity. A number of approaches have been used to circumvent such shortcomings including changes in the primary peptide structure, entrapment into nanoparticles (e.g. liposomes) and conjugation to polymers. Polysialylation, namely, conjugation of peptides and proteins to the naturally occurring, biodegradable alpha-(2--&gt;8) linked polysialic acid is a recent development, which promises to be at least as effective as PEGylation but without its potential toxicity. Polysialylation of a range of peptide and protein therapeutics has led to markedly reduced proteolysis, retention of their activity in vivo, prolongation of their half-life in the circulation and reduction in immunogenicity and antigenicity. It is anticipated that polysialylation wi...

The neural cell adhesion molecule (NCAM) plays important roles during development, plasticity, and regeneration in the adult nervous system. Its function is strongly influenced by attachment of the unusual α2-8-linked polysialic acid (PSA). Here we analyzed the N-glycosylation pattern of polysialylated NCAM from brains of newborn calves. Purified PSA-NCAM glycoprotein was digested with trypsin, and PSA-glycopeptides were separated by immunoaffinity chromatography. For determining the N-glycosylation sites, PNGase F-treated glycopeptides were analyzed by Edman degradation and matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). They were found to be exclusively linked to the fifth (Asn 439) and sixth (Asn 468) N-glycosylation sites in the fifth immunoglobulin-like domain of NCAM. The chain length of PSA consisted of at least 30 sialic acid residues, as shown by anion exchange chromatography. For analysis of the core structures, endoneuraminidase N-treated PSA-NCAM was separated by SDS-PAGE and digested with PNGase F. The core structures of polysialylated glycans were characterized by MALDI-MS combined with exoglycosidase digestions and chromatographic fractionation. They include hybrid, di-, tri-, and small amounts of tetraantennary carbohydrates, which were all fucosylated at the innermost N-acetylglucosamine. For the triantennary glycans, the "2,6" arm was preferred in polysialylated structures. High levels of sulfated groups were found on polysialylated structures and to a lower extent also on nonpolysialylated glycans. In addition, high-mannose-type glycans could be detected on PSA-NCAM glycoforms ranging from (GlcNAc) 2 (Man) 5 up to (GlcNAc) 2 (Man) 9 . In conclusion, we observed a structural variability and high regional selectivity for the PSA-glycans attached to the NCAM molecule that are most likely influencing its biological functions.

We used a novel approach to study the role of the Asn-linked oligosaccharides for human thyrotropin (hTSH) activity. Mutagenesis of Asn (N) within individual glycosylation recognition sequences to Gln (Q) was combined with expression of wild type and mutant hTSH in cell lines with different glycosylation patterns. The in vitro activity of hTSH lacking the Asn ␣52 oligosaccharide (␣Q52/TSH␤) expressed in CHO-K1 cells (sialylated oligosaccharides) was increased 6-fold compared with wild type, whereas the activities of ␣Q78/TSH␤ and ␣/TSH␤Q23 were increased 2-3-fold. Deletion of the Asn ␣52 oligosaccharide also increased the thyrotropic activity of human chorionic gonadotropin, in contrast to previous findings at its native receptor. The in vitro activity of wild type hTSH expressed in CHO-LEC2 cells (sialic acid-deficient oligosaccharides), CHO-LEC1 cells (Man 5 GlcNAc 2 intermediates), and 293 cells (sulfated oligosaccharides) was 5-8-fold higher than of wild type from CHO-K1 cells. In contrast to CHO-K1 cells, there was no difference in the activity between wild type and selectively deglycosylated mutants expressed in these cell lines. Thus, in hTSH, the oligosaccharide at Asn ␣52 and, specifically, its terminal sialic acid residues attenuate in vitro activity, in contrast to the previously reported stimulatory role of this chain for human chorionic gonadotropin and human follitropin activity. The increased thyrotropic activity of ␣Q52/CG␤ suggests that receptor-related mechanisms may be responsible for these differences among the glycoprotein hormones. Despite their increased in vitro activity, ␣Q52/TSH␤, and ␣Q78/TSH␤ from CHO-K1 cells had a faster serum disappearance rate and decreased effect on T 4 production in mice. These findings highlight the importance of individual oligosaccharides in maintaining circulatory half-life and hence in vivo activity of hTSH.

The inhibition of sialidase activity from influenza viruses A and B, parainfluenza 2 virus, Vibrio cholerae, Arthrobacter ureafaciens, Clostridium perfringens, and sheep liver by a range of 2-deoxy-2,3-didehydro-Nacetylneuraminic acid analogues modified at the C-4 position has been studied. All substitutions tested resulted in a decrease in the degree of inhibition of the bacterial and mammalian sialidases. For sialidases from influenza viruses A and B, on the other hand, most of the substitutions tested either had no significant effect on binding or, in the case of the basic amino and guanidino substituents, resulted in significantly stronger inhibition. The results for parainfluenza 2 virus sialidase were mostly intermediate, in that inhibition was neither significantly increased nor decreased by most of the modifications. We conclude that only the influenza A and B sialidase active sites possess acid groups correctly positioned to participate in charge-charge interactions in the region of C-4 of bound substrate, and that the C-4 binding pockets of the bacterial and mammalian sialidases examined are considerably smaller than is observed for either the influenza virus or parainfluenza virus sialidases.

Site-specific glycoprofiling of N-linked glycopeptides using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is an emerging technique, but its quantitative accuracy lacks documentation. Thus, a systematic study of widely different glycopeptides was performed to determine the relationship between the relative abundances of the individual glycoforms and the MALDI-TOF MS signal strength. Glycopeptides derived from glycoproteins containing neutral glycans (ribonuclease B, IgG, and ovalbumin) were initially profiled and yielded excellent and reproducible quantitation (correlation coefficient r ) 0.9958, n ) 5) when evaluated against a normal phase HPLC 2-AB glycan profile. Similarly, precise quantitation was observed for various forms of N-glycans (free, permethylated, and fluorescence-labeled) using MS. In addition, three different sialoglycopeptides from fetuin were site-specifically profiled, and good correlation between peak intensities and relative abundances was found with only a minor loss of sialic acids (r ) 0.9664, n ) 5). For glycopeptide purification, a range of hydrophilic and graphite materials packed in microcolumn format proved capable of performing desalting without loss of quantitative information, but highlighted the column capacity as a critical parameter. In conclusion, MALDI-TOF MS signal strength of glycopeptides has been found to accurately reflect the relative quantities of glycoforms, providing that certain technical issues are considered, i.e., nonbiased sample handling, matrix choice, and instrumental settings. This enables rapid and sensitive site-specific glycoprofiling of N-glycan populations to promote biomarker discovery and elucidation of glycan structure/function relationships.

Sialic acids are structurally diverse molecules that have important roles in the physiological reactions and characteristics of prokaryotes and eukaryotes. These include the ability to mask epitopes on underlying glycan chains and to repulse negatively charged moieties. Here, we describe the metabolism and immunological relevance of sialic acids and outline how their properties have been exploited by the pharmaceutical industry to enhance the therapeutic properties of proteins such as asparaginase and darbepoetin a.

Polysialic acid (PSA) is a major regulator of cell-cell interactions in the developing nervous system and in neural plasticity in the adult. As a polyanionic molecule with high water-binding capacity, PSA increases the intercellular space generating permissive conditions for cell motility. PSA enhances stem cell migration and axon path finding and promotes repair in the lesioned peripheral and central nervous systems, thus contributing to regeneration. As a next step in developing an improved PSA-based approach to treat nervous system injuries, we searched for small organic compounds that mimic PSA and identified as a PSA mimetic 5-nonyloxytryptamine oxalate, described as a selective 5hydroxytryptamine receptor 1B (5-HT 1B) agonist. Similar to PSA, 5-nonyloxytryptamine binds to the PSA-specific monoclonal antibody 735, enhances neurite outgrowth of cultured primary neurons and process formation of Schwann cells, protects neurons from oxidative stress, reduces migration of astrocytes and enhances myelination in vitro. Furthermore, nonyloxytryptamine treatment enhances expression of the neural cell adhesion molecule (NCAM) and its polysialylated form PSA-NCAM and reduces expression of the microtubuleassociated protein MAP2 in cultured neuroblastoma cells. These results demonstrate that 5-nonyloxytryptamine mimics

Despite the importance of protein glycosylation in all physiological and pathological processes and their potential as diagnostic markers and drug targets, the glycome of children is still unexplored. We analyzed N-linked plasma and IgG glycomes in 170 children and adolescents between 6 and 18 years of age. The results showed large biological variability at the population level as well as a large number of associations between different glycans and age. The plasma N-glycome of younger children was found to contain a larger proportion of large complex glycan structures (r = −0.71 for tetrasialylated glycans; r = −0.41 for trisialylated glycans) as well as an increase in disialylated biantennary structures (r = 0.55) with age. Core fucosylation and the level of agalactosylated plasma and IgG glycans decreased while digalactosylated glycans increased with age. This pattern of age-dependent changes in children differs from changes reported in adult population in both, direction and the intensity of changes. Also, sex differences are much smaller in children than in adults and are present mainly during puberty. These important observations should be accounted for when glycan-based diagnostic tests or therapeutics are being developed or evaluated.

Complex carbohydrates can frequently be separated using hydrophilic-interaction chromatography (HILIC). The mechanism was investigated using small oligosaccharides and a new column, PolyGLYCOPLEX. Some carbohydrates exhibited anomer separation, which made it possible to determine the orientation of the reducing end relative to the stationary phase. Amide sugars were consistently good contact regions. Relative to amide sugars, sialic acids and neutral hexoses were better contact regions at lower levels of organic solvents than at higher levels. HILIC readily resolved carbohydrates differing in residue composition and position of linkage. Complex carbohydrate mixtures could be resolved using volatile mobile phases. This was evaluated with native glycans and with glycans derivatized with 2-aminopyridine or a nitrobenzene derivative. Both asialo-and sialylated glycans could be resolved using the same set of conditions. With derivatized carbohydrates, detection was possible at the picomole level by UV detection or on-line electrospray mass spectrometry. Selectivity compared favorably with that of other modes of HPLC. HILIC is promising for a variety of analytical and preparative applications.

4-GU-DANA (zanamivir) (as well as DANA and 4-AM-DANA) was found to inhibit the neuraminidase activity of human parainfluenza virus type 3 (HPF3). The viral neuraminidase activity is attributable to hemagglutinin-neuraminidase (HN), an envelope protein essential for viral attachment and for fusion mediated by the other envelope protein, F. While there is no evidence that HN's neuraminidase activity is essential for receptor binding and syncytium formation, we found that 4

In a previous paper, we determined the effect of either ethanol or pentazocine administered separately on the ganglioside content of rat liver. In the present paper we have investigated the effect of pentazocine injection on the liver ganglioside contents of chronic alcoholic rats. The effect of high carbohydrate ingestion was also studied. Thirty male Wistar rats were divided into three experimental groups that received ethanol and pentazocine, a carbohydrate supplemented diet or a laboratory diet and water. Liver ganglioside contents were slightly increased in the ethanol plus pentazocine group as compared to the control and high carbohydrate diet groups. No differences were found between the two latter groups. The percentage distribution of individual gangliosides (ganglioside pattern) was also modified. A decrease in gangliosides belonging to the b-series (GD3, GD1b, GT1b and GQ1b) in parallel with an increase in that of the a-series (GM2, GM1 and GD1a) were found for both the ethanol plus pentazocine and the highcarbohydrate fed rats. The results suggest that ethanol or high carbohydrate ingestion diminishes the activity of GD3 synthase, a key enzyme in the metabolism of gangliosides, which determines the proportion of gangliosides, belonging to the a- and b-series.

The lipid composition of plasma membranes isolated from bovine adrenal chromaffin cells has been determined. Choline and ethanolamine phosphatides were predominant; the level of lyso compounds was very low. The amount of cholesterol and the cholesterol/phospholipid molar ratio was low compared to those of the other subcellular fractions of chromaffin cells. A complex pattern of neutral glycolipids was observed in contrast to that of gangliosides.

The following structure of the O-specific polysaccharide of Proteus mirabilis O5 was established by 1 H and 13 C NMR spectroscopy at 500 MHz, including two-dimensional COSY, TOCSY, NOESY, and H-detected 1 H, 13 C heteronuclear multiple-quantum coherence (HMQC) experiments:

How high doses of intravenous IgG (IVIG) suppress autoimmune diseases remains unresolved. We have recently shown that the antiinfl ammatory activity of IVIG can be attributed to a minor species of IgGs that is modifi ed with terminal sialic acids on their Fc-linked glycans. Here we propose that these Fc-sialylated IgGs engage a unique receptor on macrophages that, in turn, leads to the upregulation of an inhibitory Fcγ receptor (FcγR), thereby protecting against autoantibody-mediated pathology. . Two-cell model for the mechanism of IVIG activity. Sialic acid-rich antibodies (red) in the IVIG preparation bind to an as yet unknown IVIG receptor on CSF-1-dependent regulatory macrophages. This induces the up-regulation of the inhibitory FcγRIIB on effector macrophages, thereby increasing the threshold for cell activation in response to the binding of immune complexes and inhibiting the release of destructive infl ammatory and cytotoxic mediators.

Capsular polysaccharide was isolated by the phenol-water extraction of Acinetobacter baumannii ACICU cells and studied by sugar analysis, partial acid hydrolysis, and 1D and 2D 1 H and 13 C NMR spectroscopy. The polysaccharide was found to contain 5,7-diacetamido-3,5,7,9-tetradeoxy-L-glycero-L-manno-non-2ulosonic or di-N-acetylpseudaminic acid (Pse5Ac7Ac), and the following structure of the branched tetrasaccharide repeating unit was established:

A simple, sensitive, and rapid quantitative LC-MS/MS assay was designed for the simultaneous quantification of free and glycoprotein bound monosaccharides using a multiple reaction monitoring (MRM) approach. This study represents the first example of using LC-MS/MS methods to simultaneously quantify all common glycoprotein monosaccharides, including neutral and acidic monosaccharides. Sialic acids and reduced forms of neutral monosaccharides are efficiently separated

Bacterial superinfections are the most common cause of mortality during influenza epidemics. Depression of phagocyte functions by influenza A viruses (IAVs) is a likely contributory cause of such infections. We used an in vitro model of viral depression of neutrophil respiratory burst responses to FMLP and PMA to examine the mechanism of IAV-induced phagocyte deactivation. Respiratory burst responses or intracellular calcium mobilization were triggered by the virus itself, but these were not causally related to deactivation. By treating neutrophils with neuraminidase, and by use of purified IAV hemagglutinin (HA) preparations, cross-linking of sialic acid-bearing neutrophil surface components by the IAV HA was shown to be responsible for deactivation. IAV competed for binding to neutrophils with Abs directed against CD43, sialyl-Le&quot;, CD45, and gangliosides. Deactivation could be reproduced by treating neutrophils with anti-CD43 or-sialyl-Le &quot; Abs in the absence of IAV. How...

Studies were performed on two anti-N sera from patients who had formed anti-N antibodies during their period of haemodialysis. No specific inhibition of these antibodies was obtained by extracts prepared from a new and a used dialysis unit, and it was concluded that there was no evidence of a compound in the units which would stimulate production of the anti-N. The specificity of the anti-N's was shown to be against a precursor of MN antigens, since all activity in the sera could be removed by neuraminidase-treated M cells, which expressed MN precursor. Since the anti-N's would react with untreated N cells only at 4 O C , but with formaldehyde-treated cells at 4, 25 and 37 OC, it was concluded that the specificity of the anti-Ns was directed against an antigen developed by red cells after formaldehyde treatment. It is proposed that the production of the anti-N antibodies is stimulated by formaldehyde-treated red cells passing into the patient during re-use of a dialysis unit sterilized with formaldehyde.

In order to understand the biological importance of naturally occurring sialic acid variations on disialyl structures in nature, we developed an efficient two-step multienzyme approach for the synthesis of a series of GD3 ganglioside oligosaccharides and other disialyl glycans containing a terminal SiaR2-8Sia component with different natural and non-natural sialic acids. In the first step, R2-3-or R2-6-linked monosialylated oligosaccharides were obtained using a one-pot three-enzyme approach. These compounds were then used as acceptors for the R2-8-sialyltransferase activity of a recombinant truncated multifunctional Campylobacter jejuni sialyltransferase CstII mutant, CstII∆32 I53S , to produce disialyl oligosaccharides. The R2-8-sialyltransferase activity of CstII∆32 I53S has promiscuous donor substrate specificity and can tolerate various substitutions at C-5 or C-9 of the sialic acid in CMP-sialic acid, while its acceptor substrate specificity is relatively restricted. The terminal sialic acid residues in the acceptable monosialylated oligosaccharide acceptors are restricted to Neu5Ac, Neu5Gc, KDN, and some of their C-9-modified forms but not their C-5 derivatives. The disialyl oligosaccharides obtained are valuable probes for their biological studies.

New sialic acid-specific lectin has been isolated from culture supernatant of the protozoan Tritrichomonas mobilensis. It was purified by adsorption by erythrocytes or bovine submaxillary gland mucin (BSM)-Sepharose affinity chromatography. The T. mobilensis lectin (TML) does not require bivalent cations for activity and agglutinates all human erythrocytes. The lectin forms multimeric complexes with molecular mass 556 and 491 kDa as determined by size-exclusion chromatography. SDS/PAGE under reducing conditions disclosed a large band of 343 kDa and three bands of 246, 265 and 286 kDa which, after denaturation with urea, were split into three subunits of 56, 61 and 66 kDa; under non-reducing conditions there were two bands, of 360 and 260 kDa. Western blots performed with anti-TML monoclonal antibodies revealed bands identical with those in the silver-stained gels, suggesting homogeneity of the BSM -Sepharose-purified lectin. TML is a highly glycosylated protein with approx. 8% of N-...