Human Sialic Acid Research Articles

Swine Influenza Viruses Isolated from 2019 to 2022 in Shandong Province, China, Exemplify the Dominant Genotype.

Swine influenza viruses (SIVs) have been circulating in swine globally and are potential threats to human health. During the surveillance of SIVs in Shandong Province, China, from 2019 to 2022, 21 reassortant G4 genotype Eurasian avian-like (EA) H1N1 subtypes containing genes from the EA H1N1 (HA and NA), 2009 pandemic (pdm/09) H1N1 virus (PB2, PB1, PA, NP, and M), and classical swine (CS) H1N1 (NS) lineages were isolated. The analysis of the key functional amino acid sites in the isolated viruses showed that two mutation sites (190D and 225E) that preferentially bind to the human α2-6 sialic acid receptor were found in HA. In PB2, three mutation sites (271A, 590S, and 591R) that may increase mammalian fitness and a mutation site (431M) that increases pathogenicity in mice were found. A typical human signature marker that may promote infection in humans, 357K, was found in NP. The viruses could replicate efficiently in mouse lungs and turbinates, and one of the H1N1 isolates could replicate in mouse kidneys and brains without prior adaption, which indicates that the viruses potentially pose a threat to human health. Histopathological results showed that the isolated viruses caused typical bronchopneumonia and encephalitis in mice. The results indicate that G4 genotype H1N1 has potential transmissibility to humans, and surveillance should be enhanced, which could provide important information for assessing the pandemic potential of the viruses.

Human serum albumin and sialic acid co-modified liposome nanoreactors with tumor-specific activable cascade reactions for cooperative cancer therapy

Herein, a new type of human serum albumin (HSA) and sialic acid (SA) co-modified liposome nanoreactor was prepared to realize chemotherapy and chemodynamic therapy cooperative treatment of tumor. Firstly, Paclitaxel (PTX) acidic sensitive prodrug with a maleimide-terminated functional group and a sialic acid-modified reduced glutathione (GSH) sensitive paclitaxel prodrug have been synthesized. Then, a liposome nanoreactor was prepared by using PTX prodrugs, lecithin, and cholesterol as membrane materials, and simultaneously encapsulating water-soluble Fe3O4 and glucose oxidase. Finally, HSA was bound to the surface of liposomes through the Michael addition between maleimide-terminated functional group of PTX prodrug and the thiol group at the cysteine 34 residue of human serum albumin. The cumulative release rates of PTX in nanoreactor were 43.98 % at pH = 5.5 and 14.98 % at pH 7.4 after 96h respectively, exhibiting pH sensitive drug release behavior. Also 25.98 % and 17.52 % of PTX release were observed respectively in the medium with 10 mM DTT and without DTT after 96h, displaying redox responsive drug release behavior. The fluorescence intensity of tumor sites for the Dir loaded nanoreactor was higher than that of the free Dir group, displaying the targeting ability of HSA and SA. In tumor site, glucose oxidase can catalyze glucose to generate hydrogen peroxide, and hydroxyl radical is generated under an iron-based Fenton reaction between hydrogen peroxide and iron ion to kill tumor cells to realize chemodynamic therapy. After administration, the average tumor weight of the NS group was 2.516g, while nanoreactor group was 0.322g, displaying the synergistic therapy and excellent anticancer efficacy.

Detection of N-glycolyl-neuraminic acid-containing glycolipids in human skin.

Humans lack the enzyme that produces the sialic acid N-glycolyl neuraminic acid (Neu5Gc), but several lines of evidence have shown that Neu5Gc can be taken up by mammalian food sources and replace the common human sialic acid N-acetyl neuraminic acid (Neu5Ac) in glycans. Cancer tissue has been shown to have increased the presence of Neu5Gc and Neu5Gc-containing glycolipids such as the ganglioside GM3, which have been proposed as tumor-specific antigens for antibody treatment. Here, we show that a previously described antibody against Neu5Gc-GM3 is binding to Neu5GC-containing gangliosides and is strongly staining different cancer tissues. However, we also found a strong intracellular staining of keratinocytes of healthy skin. We confirmed this staining on freshly isolated keratinocytes by flow cytometry and detected Neu5Gc by mass spectrometry. This finding implicates that non-human Neu5Gc can be incorporated into gangliosides in human skin, and this should be taken into consideration when targeting Neu5Gc-containing gangliosides for cancer immunotherapy.

Risk assessment of the newly emerged H7N9 avian influenza viruses

ABSTRACT Since the first human case in 2013, H7N9 avian influenza viruses (AIVs) have caused more than 1500 human infections with a mortality rate of approximately 40%. Despite large-scale poultry vaccination regimes across China, the H7N9 AIVs continue to persist and evolve rapidly in poultry. Recently, several strains of H7N9 AIVs have been isolated and shown the ability to escape vaccine-induced immunity. To assess the zoonotic risk of the recent H7N9 AIV isolates, we rescued viruses with hemagglutinin (HA) and neuraminidase (NA) from these H7N9 AIVs and six internal segments from PR8 virus and characterized their receptor binding, pH of fusion, thermal stability, plaque morphology and in ovo virus replication. We also assessed the cross-reactivity of the viruses with human monoclonal antibodies (mAbs) against H7N9 HA and ferret antisera against H7N9 AIV candidate vaccines. The H7N9 AIVs from the early epidemic waves had dual sialic acid receptor binding characteristics, whereas the more recent H7N9 AIVs completely lost or retained only weak human sialic acid receptor binding. Compared with the H7N9 AIVs from the first epidemic wave, the 2020/21 viruses formed larger plaques in Madin-Darby canine kidney (MDCK) cells and replicated to higher titres in ovo, demonstrating increased acid stability but reduced thermal stability. Further analysis showed that these recent H7N9 AIVs had poor cross-reactivity with the human mAbs and ferret antisera, highlighting the need to update the vaccine candidates. To conclude, the newly emerged H7N9 AIVs showed characteristics of typical AIVs, posing reduced zoonotic risk but a heightened threat for poultry.

Structures of the Inhibitory Receptor Siglec-8 in Complex with a High-Affinity Sialoside Analogue and a Therapeutic Antibody.

Human sialic acid binding immunoglobulin-like lectin-8 (Siglec-8) is an inhibitory receptor that triggers eosinophil apoptosis and can inhibit mast cell degranulation when engaged by specific monoclonal antibodies (mAbs) or sialylated ligands. Thus, Siglec-8 has emerged as a critical negative regulator of inflammatory responses in diverse diseases, such as allergic airway inflammation. Herein, we have deciphered the molecular recognition features of the interaction of Siglec-8 with the mAb lirentelimab (2C4, under clinical development) and with a sialoside mimetic with the potential to suppress mast cell degranulation. The three-dimensional structure of Siglec-8 and the fragment antigen binding (Fab) portion of the anti-Siglec-8 mAb 2C4, solved by X-ray crystallography, reveal that 2C4 binds close to the carbohydrate recognition domain (V-type Ig domain) on Siglec-8. We have also deduced the binding mode of a high-affinity analogue of its sialic acid ligand (9-N-napthylsufonimide-Neu5Ac, NSANeuAc) using a combination of NMR spectroscopy and X-ray crystallography. Our results show that the sialoside ring of NSANeuAc binds to the canonical sialyl binding pocket of the Siglec receptor family and that the high affinity arises from the accommodation of the NSA aromatic group in a nearby hydrophobic patch formed by the N-terminal tail and the unique G-G' loop. The results reveal the basis for the observed high affinity of this ligand and provide clues for the rational design of the next generation of Siglec-8 inhibitors. Additionally, the specific interactions between Siglec-8 and the N-linked glycans present on the high-affinity receptor FcεRIα have also been explored by NMR.

Concentration and distribution of sialic acid in human milk and its correlation with dietary intake.

PurposeThis study evaluates the content, distribution, and changing trend of sialic acid in human milk and the correlation between dietary intake of sialic acid and that in human milk.MethodsThe study included 33 mothers of full-term and exclusively breastfed infants. At least 2 ml of milk was collected on the 3rd, 8th, 30th, and 90th day after delivery, and 24-h diet recalls of the lactating mothers were obtained each time. The correlation of human milk sialic acid concentration with lactating women's dietary sialic acid intake during lactation was analyzed by statistical analysis software SPSS.ResultsThe average concentration of sialic acid in colostrum, transition, and 1 and 3 months were 1,670.74 ± 94.53, 1,272.19 ± 128.74, 541.64 ± 55.2, and 297.65 ± 20.78 mg/L, respectively. The total sialic acid concentration in colostrum was about 5.6 times higher than that at 3 months (P < 0.001). The average dietary sialic acid intake of lactating mothers on the 2nd, 7th, 30th, and 90th day after delivery were 106.06 ± 7.51, 127.64 ± 8.61, 120.34 ± 10.21, and 95.40 ± 6.34 mg/day, respectively. The intake of sialic acid was relatively high on the 7th day, and there was no significant difference in dietary intake of sialic acid on different days (P < 0.05). In addition, there was no correlation between the intake of dietary sialic acid and the content of total sialic acid and various forms of sialic acid in milk (P < 0.05).ConclusionDuring the lactation period, the distribution of sialic acid in breast milk is relatively stable and its content fluctuates greatly, which may not be affected by the mother's diet, but mainly depends on the self-regulation oft physiological needs.

Characterizing Receptor Flexibility to Predict Mutations That Lead to Human Adaptation of Influenza Hemagglutinin.

A key step in the emergence of human pandemic influenza strains has been a switch in binding preference of the viral glycoprotein hemagglutinin (HA) from avian to human sialic acid (SA) receptors. The conformation of the bound SA varies substantially with HA sequence, and crystallographic evidence suggests that the bound SA is flexible, making it difficult to predict which mutations are responsible for changing HA-binding preference. We performed molecular dynamics (MD) simulations of SA analogues binding to various HAs and observed a dynamic equilibrium among structurally diverse receptor conformations, including conformations that have not been experimentally observed. Using one such novel conformation, we predicted—and experimentally confirmed—a set of mutations that substantially increased an HA’s affinity for a human SA analogue. This prediction could not have been inferred from the existing crystal structures, suggesting that MD-generated HA–SA conformational ensembles could help researchers predict human-adaptive mutations, aiding surveillance of emerging pandemic threats.

Antibodies Against Unusual Forms of Sialylated Glycans.

Previous studies have shown that in the blood of healthy donors (1) there are no natural antibodies against sialylated glycoproteins, which contain Neu5Acα (N-acetylneuraminic acid) as the most widespread form of human sialic acid, and (2) there is a moderate level of antibodies capable of binding unnatural oligosaccharides, where Neu5Ac is beta-linked to a typical mammalian glycan core. In the present study, we investigated antibodies against βNeu5Ac in more detail and verified the presence of Kdn (2-keto-3-deoxy- D-glycero-D-galacto-nonulosonic acid) as a possible cause behind their appearance in humans, taking into account the expected cross-reactivity to Kdn glycans, which are found in bacterial glycoconjugates in both the α- and β-forms. We observed the binding of peripheral blood immunoglobulins to sialyllactosamines (where “sialyl” is Kdn or neuraminic acid) in only a very limited number of donors, while the binding to monosaccharide Kdn occurred in all samples, regardless of the configuration of the glycosidic bond of the Kdn moiety. In some individuals, the binding level of some of the immunoglobulins was high. This means that bacterial Kdn glycoconjugates are very unlikely to induce antibodies to βNeu5Ac glycans in humans. To determine the reason for the presence of these antibodies, we focused on noninfectious pathologies, as well as on a normal state in which a significant change in the immune system occurs: namely, pregnancy. As a result, we found that 2/3 of pregnant women have IgM in the blood against Neu5Acβ2-3Galβ1-4GlcNAcβ. Moreover, IgG class antibodies against Neu5Acβ2-3Galβ1-4GlcNAcβ and Neu5Acβ2-6Galβ1-4GlcNAcβ were also detected in eluates from the placenta. Presumably, these antibodies block fetal antigens.

Polymer-Stabilized Sialylated Nanoparticles: Synthesis, Optimization, and Differential Binding to Influenza Hemagglutinins.

During influenza infection, hemagglutinins (HAs) on the viral surface bind to sialic acids on the host cell’s surface. While all HAs bind sialic acids, human influenza targets terminal α2,6 sialic acids and avian influenza targets α2,3 sialic acids. For interspecies transmission (zoonosis), HA must mutate to adapt to these differences. Here, multivalent gold nanoparticles bearing either α2,6- or α2,3-sialyllactosamine have been developed to interrogate a panel of HAs from pathogenic human, low pathogenic avian, and other species’ influenza. This method exploits the benefits of multivalent glycan presentation compared to monovalent presentation to increase affinity and investigate how multivalency affects selectivity. Using a library-orientated approach, parameters including polymer coating and core diameter were optimized for maximal binding and specificity were probed using galactosylated particles and a panel of biophysical techniques [ultraviolet–visible spectroscopy, dynamic light scattering, and biolayer interferometry]. The optimized particles were then functionalized with sialyllactosamine and their binding analyzed against a panel of HAs derived from pathogenic influenza strains including low pathogenic avian strains. This showed significant specificity crossover, which is not observed in monovalent formats, with binding of avian HAs to human sialic acids and vice versa in agreement with alternate assay formats. These results demonstrate that precise multivalent presentation is essential to dissect the interactions of HAs and may aid the discovery of tools for disease and zoonosis transmission.

Biological Functions and Analytical Strategies of Sialic Acids in Tumor.

Sialic acids, a subset of nine carbon acidic sugars, often exist as the terminal sugars of glycans on either glycoproteins or glycolipids on the cell surface. Sialic acids play important roles in many physiological and pathological processes via carbohydrate-protein interactions, including cell–cell communication, bacterial and viral infections. In particular, hypersialylation in tumors, as well as their roles in tumor growth and metastasis, have been widely described. Recent studies have indicated that the aberrant sialylation is a vital way for tumor cells to escape immune surveillance and keep malignance. In this article, we outline the present state of knowledge on the metabolic pathway of human sialic acids, the function of hypersialylation in tumors, as well as the recent labeling and analytical techniques for sialic acids. It is expected to offer a brief introduction of sialic acid metabolism and provide advanced analytical strategies in sialic acid studies.

A glycal-based photoaffinity probe that enriches sialic acid binding proteins.

To identify sialic acid binding proteins from complex proteomes, three photocrosslinking affinity-based probes were constructed using Neu5Ac (5 and 6) and Neu5Ac2en (7) scaffolds. Kinetic inhibition assays and Western blotting revealed the Neu5Ac2en-based 7 to be an effective probe for the labeling of a purified gut microbial sialidase (BDI_2946) and a purified human sialic acid binding protein (hCD33). Additionally, LC-MS/MS affinity-based protein profiling verified the ability of 7 to enrich a low-abundance sialic acid binding protein (complement factor H) from human serum thus validating the utility of this probe in a complex context.

Human Sialic acid O-acetyl esterase (SIAE) \u2013 mediated changes in sensitivity to etoposide in a medulloblastoma cell line

Medulloblastoma (MB), the most common malignant paediatric brain tumour occurs in the cerebellum. Advances in molecular genomics have led to the identification of defined subgroups which are associated with distinct clinical prognoses. Despite this classification, standard therapies for all subgroups often leave children with life-long neurological deficits. New therapeutic approaches are therefore urgently needed to reduce current treatment toxicity and increase survival for patients. GD3 is a well-studied ganglioside which is known to have roles in the development of the cerebellum. Post-partum GD3 is not highly expressed in the brain. In some cancers however GD3 is highly expressed. In MB cells GD3 is largely acetylated to GD3A. GD3 is pro-apoptotic but GD3A can protect cells from apoptosis. Presence of these gangliosides has previously been shown to correlate with resistance to chemotherapy. Here we show that the GD3 acetylation pathway is dysregulated in MB and as a proof-of-principle we show that increased GD3 expression sensitises an MB cell line to etoposide.

Quantitation of Sialic Acids in Infant Formulas by Liquid Chromatography–Mass Spectrometry: An Assessment of Different Protein Sources and Discovery of New Analogues

Glycosidically bound, but not free, dietary sialic acids are used for the biosynthesis of new glycoconjugates in humans, making the quantitation of these two forms in infant food sources important, as in neonates the demand for sialic acid may exceed the de novo biosynthetic supply. Here, a rapid high-performance liquid chromatography-mass spectrometry method was developed to identify and quantitate glycosidically bound and free sialic acids in infant formulas. The sialic acid contents of eight commercially available infant formulas with varying protein source or manufacturer were investigated. The formula protein sources (whey vs casein) did not have a large impact on the ratios of free to bound sialic acids, nor did protein hydrolysis or sample form (solid vs liquid). Hydrolyzed bovine whey protein-based formulas were found to contain the highest amount of the most abundant human sialic acid, 5- N-acetylneuraminic acid (Neu5Ac). O-Acetylated Neu5Ac was quantified in all formulas tested and, for the first time, 2-keto-3-deoxy-d-glycero-d-galacto-nononic acid (Kdn) was detected in several infant formulas.

Examination of Signatures of Recent Positive Selection on Genes Involved in Human Sialic Acid Biology.

Sialic acids are nine carbon sugars ubiquitously found on the surfaces of vertebrate cells and are involved in various immune response-related processes. In humans, at least 58 genes spanning diverse functions, from biosynthesis and activation to recycling and degradation, are involved in sialic acid biology. Because of their role in immunity, sialic acid biology genes have been hypothesized to exhibit elevated rates of evolutionary change. Consistent with this hypothesis, several genes involved in sialic acid biology have experienced higher rates of non-synonymous substitutions in the human lineage than their counterparts in other great apes, perhaps in response to ancient pathogens that infected hominins millions of years ago (paleopathogens). To test whether sialic acid biology genes have also experienced more recent positive selection during the evolution of the modern human lineage, reflecting adaptation to contemporary cosmopolitan or geographically-restricted pathogens, we examined whether their protein-coding regions showed evidence of recent hard and soft selective sweeps. This examination involved the calculation of four measures that quantify changes in allele frequency spectra, extent of population differentiation, and haplotype homozygosity caused by recent hard and soft selective sweeps for 55 sialic acid biology genes using publicly available whole genome sequencing data from 1,668 humans from three ethnic groups. To disentangle evidence for selection from confounding demographic effects, we compared the observed patterns in sialic acid biology genes to simulated sequences of the same length under a model of neutral evolution that takes into account human demographic history. We found that the patterns of genetic variation of most sialic acid biology genes did not significantly deviate from neutral expectations and were not significantly different among genes belonging to different functional categories. Those few sialic acid biology genes that significantly deviated from neutrality either experienced soft sweeps or population-specific hard sweeps. Interestingly, while most hard sweeps occurred on genes involved in sialic acid recognition, most soft sweeps involved genes associated with recycling, degradation and activation, transport, and transfer functions. We propose that the lack of signatures of recent positive selection for the majority of the sialic acid biology genes is consistent with the view that these genes regulate immune responses against ancient rather than contemporary cosmopolitan or geographically restricted pathogens.

Evidence for a carbohydrate-binding module (CBM) of Tannerella forsythia NanH sialidase, key to interactions at the host-pathogen interface.

Bacterial sialidases cleave terminal sialic acid from a variety of host glycoproteins, and contribute to survival and growth of many human-dwelling bacterial species, including various pathogens. Tannerella forsythia, an oral, Gram-negative, fastidious anaerobe, is a key organism in periodontal disease and possesses a dedicated sialic acid utilisation and scavenging (nan) operon, including NanH sialidase. Here, we describe biochemical characterisation of recombinant NanH, including its action on host-relevant sialoglycans such as sialyl Lewis A and sialyl Lewis X (SLeA/X), and on human cell-attached sialic acids directly, uncovering that it is a highly active broad specificity sialidase. Furthermore, the N-terminal domain of NanH was hypothesised and proved to be capable of binding to a range of sialoglycans and non-sialylated derivatives with Kd in the micromolar range, as determined by steady-state tryptophan fluorescence spectroscopy, but it has no catalytic activity in isolation from the active site. We consider this domain to represent the founding member of a novel subfamily of carbohydrate-binding module (CBM), involved in glycosidase-ligand binding. In addition, we created a catalytically inactive version of the NanH enzyme (FRIP → YMAP) that retained its ability to bind sialic acid-containing ligands and revealed for the first time that binding activity of a CBM is enhanced by association with the catalytic domain. Finally, we investigated the importance of Lewis-type sialoglycans on T. forsythia-host interactions, showing that nanomolar amounts of SLeA/X were capable of reducing invasion of oral epithelial cells by T. forsythia, suggesting that these are key ligands for bacterial-cellular interactions during periodontal disease.

A Single Mutation at Position 190 in Hemagglutinin Enhances Binding Affinity for Human Type Sialic Acid Receptor and Replication of H9N2 Avian Influenza Virus in Mice.

Influenza virus hemagglutinin (HA) is responsible for binding to host cell receptors and therefore influences the viral host range and pathogenicity in different species. We showed that the H9N2 avian influenza viruses harboring 190V in the HA exhibit enhanced virus replication in mice. Further studies demonstrate that 190V in the HA does not change virus receptor binding specificity but enhances virus binding affinity of the H9N2 virus to human cells and attachment to lung tissues from humans and mouse. Our findings suggest that more attention should be given to the H9N2 AIVs with HA-190V during surveillance due to their potential threat to mammals, including humans.

Dietary fish oil at low intakes increases DHA incorporation and reduces low frequency fatigue in rat hindlimb skeletal muscle

Abstract of a presentation. Publication Details Henry, R., Peoples, G. E. & McLennan, P. (2016). Dietary fish oil at low intakes increases DHA incorporation and reduces low frequency fatigue in rat hindlimb skeletal muscle. In The Nutrition Society of Australia and New Zealand 2015 Annual Scientific Meeting, 1-4 Dec 2015, Wellington, New Zealand. Journal of Nutrition & Intermediary Metabolism, 4 36-37. This conference paper is available at Research Online: http://ro.uow.edu.au/smhpapers/4019 CONCENTRATION AND DISTRIBUTION OF SIALIC ACID IN SOW MILK DURING LACTATION M. Jahan, P. Wynn, B. Wang. EH Graham Centre for Agricultural Innovation, Charles Sturt University, NSW, Australia E-mail address: mjahan@csu.edu.au (M. Jahan) Background/Aims: Human milk sialic acids has been proposed as a bioactive compound promoting immune function, gut maturation and neurodevelopment of the newborn. Porcine milk however, has received little attention. The aims of the present study were to quantify and compare the levels of N-acetylneuraminic acid (Neu5Ac), N-glycolylneuraminic acid (Neu5Gc) and ketodeoxynonulpsonic acid (KDN) in sow milk during course of lactation. Method:Milk samples from 22 sows were collected by manual expression on 3 occasions, day 1 (colostrum), day 3 (transition milk) and day 15-21 (mature milk) respectively. The concent-ration of Neu5Ac, Neu5Gc and KDN were analyzed using UHPLC. Results: Sow milk contained significant amounts of Sia with the highest concentration found in colostrum (1238.50 mg/L) followed by transition milk and then mature milk. Neu5Ac was the major form of Sia (93-96%) and then Neu5Gc (3-6%), KDN however contained as little as 1-2%. This distribution was common to each milk fraction and to each time point in lactation. Conclusions: Porcine milk contained a rich source of sialylated glycoconjugate. The predominately form of sialic acid is Neu5Ac. The high concentrations of Sia in porcine milk suggest that Sia is an important nutrient that may contribute to the optimization of immune function, neuro-development and growth and development of piglets. Funding source(s): International Postgraduate Research Scholarships, Charles Sturt University. MOLECULAR MECHANISMS DRIVING AIRWAY INFLAMMATION FOLLOWING A HIGH-FAT MIXED MEAL IN ASTHMA Q. Li , K.J. Baines , P.G. Gibson , L.G. Wood . Centre for Asthma and Respiratory Diseases, Hunter Medical Research Institute, University of Newcastle, NSW, Australia; Department of Respiratory and Sleep Medicine, John Hunter Hospital, NSW, Australia E-mail address: c3156151@uon.edu.au (Q. Li) Background/Aims: A high fat meal is associatedwith airway inflammation in asthma, but the mechanisms are not well understood. This aim of this study was to use microarray techniques to examine the molecular mechanisms of fat-induced airway inflammation in asthma compared with healthy controls. Methods: Subjects with asthma (n 1⁄4 11) and healthy controls (n 1⁄4 8) were provided with a high-fat, high energy meal, containing total energy (TE) of 3846 kJ and 48 g (49% of TE) total fat, including 20.5 g (21% of TE) saturated fat. Sputum was induced at 0 and 4 hours and gene expression was examined by microarray and quantitative real-time PCR (qPCR). Results: Following the high fat dietary challenge, 168 entities were significantly differentially expressed greater than 1.5 fold in subjects with asthma. Five genes involved in immune system processes were selected for qPCR analysis (S100P, S100A16, MAL, MUC1 and NLPR12). qPCR confirmed that S100P, S100A16, MAL and MUC1 were significantly increased in the asthma group post-meal. There was a moderate and significant correlation between the change in S100P andMUC1 gene expression and the change in sputum %neutrophils following the high fat meal (r 1⁄4 0.552, p 1⁄4 0.024; r 1⁄4 0.495, p 1⁄4 0.045 respectively). NLRP12 gene expression at 4 hours strongly correlated with the change in total and saturated non-esterified plasma fatty acid levels at 2 hours (r 1⁄4 0.555, p 1⁄4 0.028; r 1⁄4 0.53, p 1⁄4 0.037 respectively). Conclusions: Our data identifies several genes that contribute to neutrophilic airway inflammation following consumption of a high fat meal in asthmatics, which may prove to be therapeutic targets for immunomodulation. Funding sources: NHMRC, HMRI, Thoracic Society of Australia and New Zealand. DOES INCREASED INTAKE OF FOLIC ACID INCREASE CANCER RISK? D. Mackerras, J. Tan, C. Larter. Food Standards Australia New Zealand, ACT,

The pH Effect on Backbone Dynamics of the Antifreeze‐like Domain of Human Sialic Acid Synthase

The pH Effect on Backbone Dynamics of the Antifreeze‐like Domain of Human Sialic Acid Synthase

Human sialic acid acetyl esterase: Towards a better understanding of a puzzling enzyme.

Sialic acid acetyl esterase (SIAE) removes acetyl moieties from the hydroxyl groups in position 9 and 4 of sialic acid. Recently, a dispute has been opened on its association to autoimmunity. In order to get new insights on human SIAE biology and to clarify its seemingly contradictory molecular properties, we combined in silico characterization, phylogenetic analysis and homology modeling with cellular studies in COS7 cells. Genomic and phylogenetic analysis revealed that in most tissues only the "long" isoform, originally referred to lysosomal sialic acid esterase, is detected. Using the homology modeling approach, we predicted a model of SIAE 3D structure, which fulfills the topological features of SGNH-hydrolase family. In addition, the model and site-directed mutagenesis experiments allowed the definition of the residues involved in catalysis. SIAE transient expression revealed that the protein is glycosylated and is active in vitro as an esterase with a pH optimum corresponding to 8.4-8.5. Moreover, glycosylation influences the biological activity of the enzyme and is essential for release of SIAE into the culture medium. According to these findings, co-localization experiments demonstrated the presence of SIAE in membranous structures corresponding to endoplasmic reticulum and Golgi complex. Thus, at least in COS7 cells, SIAE behaves as a typical secreted enzyme, subjected to glycosylation and located along the classical secretory route or in the extracellular space. In these environments, the enzyme could act on 9-O-acetylated sialic acid residues, contributing to the fine-tuning of the various functions played by this acidic sugar.

Preexisting human antibodies neutralize recently emerged H7N9 influenza strains.

The emergence and seasonal persistence of pathogenic H7N9 influenza viruses in China have raised concerns about the pandemic potential of this strain, which, if realized, would have a substantial effect on global health and economies. H7N9 viruses are able to bind to human sialic acid receptors and are also able to develop resistance to neuraminidase inhibitors without a loss in fitness. It is not clear whether prior exposure to circulating human influenza viruses or influenza vaccination confers immunity to H7N9 strains. Here, we demonstrate that 3 of 83 H3 HA-reactive monoclonal antibodies generated by individuals that had previously undergone influenza A virus vaccination were able to neutralize H7N9 viruses and protect mice against homologous challenge. The H7N9-neutralizing antibodies bound to the HA stalk domain but exhibited a difference in their breadth of reactivity to different H7 influenza subtypes. Mapping viral escape mutations suggested that these antibodies bind at least two different epitopes on the stalk region. Together, these results indicate that these broadly neutralizing antibodies may contribute to the development of therapies against H7N9 strains and may also be effective against pathogenic H7 strains that emerge in the future.