Publications by Year: 2019

2019
Byrd-Leotis L, Jia N, Dutta S, Trost JF, Gao C, Cummings SF, Braulke T, Müller-Loennies S, Heimburg-Molinaro J, Steinhauer DA, et al. Influenza binds phosphorylated glycans from human lung. Sci Adv. [Internet]. 2019;5 (2). Publisher's VersionAbstract
Influenza A viruses can bind sialic acid-terminating glycan receptors, and species specificity is often correlated with sialic acid linkage with avian strains recognizing α2,3-linked sialylated glycans and mammalian strains preferring α2,6-linked sialylated glycans. These paradigms derive primarily from studies involving erythrocyte agglutination, binding to synthetic receptor analogs or binding to undefined surface markers on cells or tissues. Here, we present the first examination of the N-glycome of the human lung for identifying natural receptors for a range of avian and mammalian influenza viruses. We found that the human lung contains many α2,3- and α2,6-linked sialylated glycan determinants bound by virus, but all viruses also bound to phosphorylated, nonsialylated glycans.
Gao C, Hanes MS, Byrd-Leotis L, Wei M, Jia N, Kardish RJ, McKitrick TR, Steinhauer DA, Cummings RD. Unique Binding Specificities of Proteins toward Isomeric Asparagine-Linked Glycans. Cell Chem Biol. [Internet]. 2019;26 (4) :535-347. Publisher's VersionAbstract
The glycan ligands recognized by Siglecs, influenza viruses, and galectins, as well as many plant lectins, are not well defined. To explore their binding to asparagine (Asn)-linked N-glycans, we synthesized a library of isomeric multiantennary N-glycans that vary in terminal non-reducing sialic acid, galactose, and N-acetylglucosamine residues, as well as core fucose. We identified specific recognition of N-glycans by several plant lectins, human galectins, influenza viruses, and Siglecs, and explored the influence of sialic acid linkages and branching of the N-glycans. These results show the unique recognition of complex-type N-glycans by a wide variety of glycan-binding proteins and their abilities to distinguish isomeric structures, which provides new insights into the biological roles of these proteins and the uses of lectins in biological applications to identify glycans.
Smith DF, Cummings RD, Song X. History and future of shotgun glycomics. Biochem Soc Trans. [Internet]. 2019;47 (1) :1-11. Publisher's VersionAbstract
Glycans in polysaccharides and glycoconjugates of the hydrophilic exterior of all animal cells participate in signal transduction, cellular adhesion, intercellular signaling, and sites for binding of pathogens largely through protein-glycan interactions. Microarrays of defined glycans have been used to study the binding specificities of biologically relevant glycan-binding proteins (GBP), but such arrays are limited by their lack of diversity or relevance to the GBP being investigated. Shotgun glycan microarrays are made up of structurally undefined glycans that were released from natural sources, labeled with bifunctional reagents so that they can be monitored during their purification using multidimensional chromatographic procedures, stored as a tagged glycan library (TGL) and subsequently printed onto microarrays at equal molar concentrations. The shotgun glycan microarray is then interrogated with a biologically relevant GBP and the corresponding glycan ligands can be retrieved from the TGL for detailed structural analysis and further functional analysis. Shotgun glycomics extended the defined glycan microarray to a discovery platform that supports functional glycomic analyses and may provide a useful process for ultimately defining the human glycome.
Mehta A, Cummings RD. GLAD: GLycan Array Dashboard, a Visual Analytics Tool for Glycan Microarrays. Bioinformatics [Internet]. 2019. Publisher's VersionAbstract

MOTIVATION:

Traditional glycan microarray data is typically presented as excel files with limited visualization and interactivity. Thus, comparisons and analysis of glycan array data have been difficult, and there is need for a tool to facilitate data mining of glycan array data.

RESULTS:

GLAD (GLycan Array Dashboard) is a web-based tool to visualize, analyze, present, and mine glycan microarray data. GLAD allows users to input multiple data files to create comparisons. GLAD extends the capability of the microarray data to produce more comparative visualizations in the form of grouped bar charts, heatmaps, calendar heatmaps, force graphs and correlation maps in order to analyze broad sets of samples. Additionally, it allows users to filter, sort and normalize the data and view glycan structures in an interactive manner, to facilitate faster visual data mining.

AVAILABILITY:

GLAD is freely available for use on the Web at https://glycotoolkit.com/Tools/GLAD/ with all major modern browsers (Edge, Firefox, Chrome, Safari).

SUPPLEMENTARY INFORMATION:

Full documentation and video tutorials for GLAD can be found on https://glycotoolkit.com/GLAD.

Lu LL, Smith MT, Yu KKQ, Luedemann C, Suscovich TJ, Grace PS, Cain A, Yu WH, McKitrick TR, Lauffenburger D, et al. IFN-γ-independent immune markers of Mycobacterium tuberculosis exposure. Nat Med. [Internet]. 2019;9 (6) :977-987. Publisher's VersionAbstract
Exposure to Mycobacterium tuberculosis (Mtb) results in heterogeneous clinical outcomes including primary progressive tuberculosis and latent Mtb infection (LTBI). Mtb infection is identified using the tuberculin skin test and interferon-γ (IFN-γ) release assay IGRA, and a positive result may prompt chemoprophylaxis to prevent progression to tuberculosis. In the present study, we report on a cohort of Ugandan individuals who were household contacts of patients with TB. These individuals were highly exposed to Mtb but tested negative disease by IFN-γ release assay and tuberculin skin test, 'resisting' development of classic LTBI. We show that 'resisters' possess IgM, class-switched IgG antibody responses and non-IFN-γ T cell responses to the Mtb-specific proteins ESAT6 and CFP10, immunologic evidence of exposure to Mtb. Compared to subjects with classic LTBI, 'resisters' display enhanced antibody avidity and distinct Mtb-specific IgG Fc profiles. These data reveal a distinctive adaptive immune profile among Mtb-exposed subjects, supporting an expanded definition of the host response to Mtb exposure, with implications for public health and the design of clinical trials.
Pettinato G, Lehoux S, Ramanathan R, Salem MM, He LX, Muse O, Flaumenhaft R, Thompson MT, Rouse EA, Cummings RD, et al. Generation of fully functional hepatocyte-like organoids from human induced pluripotent stem cells mixed with Endothelial Cells. Sci Rep. [Internet]. 2019;9 (1). Publisher's VersionAbstract
Despite advances in stem cell research, cell transplantation therapy for liver failure is impeded by a shortage of human primary hepatocytes (HPH), along with current differentiation protocol limitations. Several studies have examined the concept of co-culture of human induced pluripotent cells (hiPSCs) with various types of supporting non-parenchymal cells to attain a higher differentiation yield and to improve hepatocyte-like cell functions both in vitro and in vivo. Co-culturing hiPSCs with human endothelial cells (hECs) is a relatively new technique that requires more detailed studies. Using our 3D human embryoid bodies (hEBs) formation technology, we interlaced Human Adipose Microvascular Endothelial Cells (HAMEC) with hiPSCs, leading to a higher differentiation yield and notable improvements across a wide range of hepatic functions. We conducted a comprehensive gene and protein secretion analysis of our HLCs coagulation factors profile, showing promising results in comparison with HPH. Furthermore, a stage-specific glycomic analysis revealed that the differentiated hepatocyte-like clusters (HLCs) resemble the glycan features of a mature tissue rather than cells in culture. We tested our HLCs in animal models, where the presence of HAMEC in the clusters showed a consistently better performance compared to the hiPSCs only group in regard to persistent albumin secretion post-transplantation.
Tuccinardi D, Farr OM, Upadhyay J, Oussaada SM, Klapa MI, Candela M, Rampelli S, Lehoux S, Lázaro I, Sala-Vila A, et al. Mechanisms Underlying the Cardiometabolic Protective Effect of Walnut Consumption in Obese Subjects: A Cross-Over, Randomized, Double-Blinded, Controlled Inpatient Physiology Study. Diabetes Obes Metab. [Internet]. 2019. Publisher's VersionAbstract

AIMS:

To assess the effects of walnuts on cardiometabolic outcomes in obese subjects and to explore underlying mechanisms using novel methods including metabolomic, lipidomic, glycomic, and microbiome analysis integrated with lipid particle fractionation, appetite-regulating hormones and hemodynamic measurements.

MATERIALS AND METHODS:

10 obese subjects were enrolled in this cross-over, randomized, double-blind, placebo-controlled clinical trial. Patients participated in two 5-day inpatient stays during which they consumed a smoothie containing 48g walnuts or a macronutrient-matched placebo smoothie without nuts, with a one-month washout period between the two visits.

RESULTS:

Walnut consumption improved aspects of the lipid profile, i.e. reduced fasting small and dense LDL particles (p<.02) and increased postprandial large HDL particles (p<.01). Lipoprotein Insulin Resistance Score, glucose and insulin AUC decreased significantly after walnut consumption (p<.01, p<.02, p<.04, respectively). Consuming walnuts significantly increased 10 N-glycans, with 8 of them carrying a fucose core. Lipidomic analysis showed a robust reduction in harmful ceramides, hexosylceramides and sphingomyelins, which have been shown to mediate effects on cardiometabolic risk. Peptide YY AUC significantly increased after walnut consumption (p<.03). No major significant changes in hemodynamic, metabolomic analysis or in host health-promoting bacteria such as Faecalibacterium were found.

CONCLUSIONS:

These data provide a more comprehensive mechanistic perspective of the effect of dietary walnut consumption on cardiometabolic parameters. Lipidomic and lipid nuclear magnetic resonance spectroscopy analysis showed an early but significant reduction in ceramides and other atherogenic lipids with walnut consumption that may explain the longer-term benefits of walnuts on insulin resistance, cardiovascular risk and mortality. This article is protected by copyright. All rights reserved.

Woodward AM, Lehoux S, Mantelli F, Di Zazzo A, Brockhausen I, Bonini S, Argüeso P. Inflammatory Stress Causes N-Glycan Processing Deficiency in Ocular Autoimmune Disease. Am J Pathol. [Internet]. 2019;189 (2) :283-294. Publisher's VersionAbstract
High levels of proinflammatory cytokines have been associated with a loss of tissue function in ocular autoimmune diseases, but the basis for this relationship remains poorly understood. Here we investigate a new role for tumor necrosis factor α in promoting N-glycan-processing deficiency at the surface of the eye through inhibition of N-acetylglucosaminyltransferase expression in the Golgi. Using mass spectrometry, complex-type biantennary oligosaccharides were identified as major N-glycan structures in differentiated human corneal epithelial cells. Remarkably, significant differences were detected between the efficacies of cytokines in regulating the expression of glycogenes involved in the biosynthesis of N-glycans. Tumor necrosis factor α but not IL-1β had a profound effect in suppressing the expression of enzymes involved in the Golgi branching pathway, including N-acetylglucosaminyltransferases 1 and 2, which are required for the formation of biantennary structures. This decrease in gene expression was correlated with a reduction in enzymatic activity and impaired N-glycan branching. Moreover, patients with ocular mucous membrane pemphigoid were characterized by marginal N-acetylglucosaminyltransferase expression and decreased N-glycan branching in the conjunctiva. Together, these data indicate that proinflammatory cytokines differentially influence the expression of N-glycan-processing enzymes in the Golgi and set the stage for future studies to explore the pathophysiology of ocular autoimmune diseases.