Genes and signal molecules involved in the rhizobialeguminoseae symbiosis

Microbial Biotechnolog

Serum Protein N-Glycosylation Changes with Rheumatoid Arthritis Disease Activity during and after Pregnancy

Proteomic

Serum N-glycan profiles differ for various breast cancer subtypes

Breast cancer is the most prevalent cancer in women. Early detection of this disease improves survival and therefore population screenings, based on mammography, are performed. However, the sensitivity of this screening modality is not optimal and new screening methods, such as blood tests, are being explored. Most of the analyses that aim for early detection focus on proteins in the bloodstream. In this study, the biomarker potential of total serum N-glycosylation analysis was explored with regard to detection of breast cancer. In an age-matched case-control setup serum protein N-glycan profiles from 145 breast cancer patients were compared to those from 171 healthy individuals. N-glycans were enzymatically released, chemically derivatized to preserve linkage-specificity of sialic acids and characterized by high resolution mass spectrometry. Logistic regression analysis was used to evaluate associations of specific N-glycan structures as well as N-glycosylation traits with breast cancer. In a case-control comparison three associations were found, namely a lower level of a two triantennary glycans and a higher level of one tetraantennary glycan in cancer patients. Of note, various other N-glycomic signatures that had previously been reported were not replicated in the current cohort. It was further evaluated whether the lack of replication of breast cancer N-glycomic signatures could be partly explained by the heterogenous character of the disease since the studies performed so far were based on cohorts that included diverging subtypes in different numbers. It was found that serum N-glycan profiles differed for the various cancer subtypes that were analyzed in this study.Surgical oncolog

Impact of asialoglycoprotein receptor and mannose receptor deficiency on murine plasma N-glycome profiles

The asialoglycoprotein receptor (ASGPR) and the mannose receptor C -type 1 (MRC1) are well known for their selective recognition and clearance of circulating glycoproteins. Terminal galactose and N-Acetylgalactosamine are recognized by ASGPR, while terminal mannose, fucose, and N-Acetylglucosamine are recognized by MRC1. The effects of ASGPR and MRC1 deficiency on the N-glycosylation of individual circulating proteins have been studied. However, the impact on the homeostasis of the major plasma glycoproteins is debated and their glycosylation has not been mapped with high molecular resolution in this context. Therefore, we evaluated the total plasma N-glycome and plasma proteome of ASGR1 and MRC1 deficient mice. ASGPR deficiency resulted in an increase in O-acetylation of sialic acids accompanied by higher levels of apolipoprotein D, haptoglobin, and vitronectin. MRC1 deficiency decreased fucosylation without affecting the abundance of the major circulating glycoproteins. Our findings confirm that concentrations and Nglycosylation of the major plasma proteins are tightly controlled and further suggest that glycan-binding receptors have redundancy, allowing compensation for the loss of one major clearance receptor.Proteomic

Autoimmune hepatitis displays distinctively high multi-antennary sialylation on plasma N-glycans compared to other liver diseases

BackgroundChanges in plasma protein glycosylation are known to functionally affect proteins and to associate with liver diseases, including cirrhosis and hepatocellular carcinoma. Autoimmune hepatitis (AIH) is a liver disease characterized by liver inflammation and raised serum levels of IgG, and is difficult to distinguish from other liver diseases. The aim of this study was to examine plasma and IgG-specific N-glycosylation in AIH and compare it with healthy controls and other liver diseases.MethodsIn this cross-sectional cohort study, total plasma N-glycosylation and IgG Fc glycosylation analysis was performed by mass spectrometry for 66 AIH patients, 60 age- and sex-matched healthy controls, 31 primary biliary cholangitis patients, 10 primary sclerosing cholangitis patients, 30 non-alcoholic fatty liver disease patients and 74 patients with viral or alcoholic hepatitis. A total of 121 glycans were quantified per individual. Associations between glycosylation traits and AIH were investigated as compared to healthy controls and other liver diseases.ResultsGlycan traits bisection (OR: 3.78 [1.88–9.35], p-value: 5.88 × 10− 3), tetraantennary sialylation per galactose (A4GS) (OR: 2.88 [1.75–5.16], p-value: 1.63 × 10− 3), IgG1 galactosylation (OR: 0.35 [0.2–0.58], p-value: 3.47 × 10− 5) and hybrid type glycans (OR: 2.73 [1.67–4.89], p-value: 2.31 × 10− 3) were found as discriminators between AIH and healthy controls. High A4GS differentiated AIH from other liver diseases, while bisection associated with cirrhosis severity.ConclusionsCompared to other liver diseases, AIH shows distinctively high A4GS levels in plasma, with potential implications on glycoprotein function and clearance. Plasma-derived glycosylation has potential to be used as a diagnostic marker for AIH in the future. This may alleviate the need for a liver biopsy at diagnosis. Glycosidic changes should be investigated further in longitudinal studies and may be used for diagnostic and monitoring purposes in the future.Cellular mechanisms in basic and clinical gastroenterology and hepatolog

Automated Plasma Glycomics with Linkage-Specific Sialic Acid Esterification and Ultrahigh Resolution MS

Surgical oncolog

Large-scale analysis of apolipoprotein CIII glycosylation by ultrahigh resolution mass spectrometry

Apolipoprotein-CIII (apo-CIII) is a glycoprotein involved in lipid metabolism and its levels are associated with cardiovascular disease risk. Apo-CIII sialylation is associated with improved plasma triglyceride levels and its glycosylation may have an effect on the clearance of triglyceride-rich lipoproteins by directing these particles to different metabolic pathways. Large-scale sample cohort studies are required to fully elucidate the role of apo-CIII glycosylation in lipid metabolism and associated cardiovascular disease. In this study, we revisited a high-throughput workflow for the analysis of intact apo-CIII by ultrahigh-resolution MALDI FT-ICR MS. The workflow includes a chemical oxidation step to reduce methionine oxidation heterogeneity and spectrum complexity. Sinapinic acid matrix was used to minimize the loss of sialic acids upon MALDI. MassyTools software was used to standardize and automate MS data processing and quality control. This method was applied on 771 plasma samples from individuals without diabetes allowing for an evaluation of the expression levels of apo-CIII glycoforms against a panel of lipid biomarkers demonstrating the validity of the method. Our study supports the hypothesis that triglyceride clearance may be regulated, or at least strongly influenced by apo-CIII sialylation. Interestingly, the association of apo-CIII glycoforms with triglyceride levels was found to be largely independent of body mass index. Due to its precision and throughput, the new workflow will allow studying the role of apo-CIII in the regulation of lipid metabolism in various disease settings.Proteomic

Serum N-glycome analysis reveals pancreatic cancer disease signatures

Background &Aims Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer type with loco-regional spread that makes the tumor surgically unresectable. Novel diagnostic tools are needed to improve detection of PDAC and increase patient survival. In this study we explore serum proteinN-glycan profiles from PDAC patients with regard to their applicability to serve as a disease biomarker panel. Methods Total serumN-glycome analysis was applied to a discovery set (86 PDAC cases/84 controls) followed by independent validation (26 cases/26 controls) using in-house collected serum specimens. ProteinN-glycan profiles were obtained using ultrahigh resolution mass spectrometry and included linkage-specific sialic acid information.N-glycans were relatively quantified and case-control classification performance was evaluated based on glycosylation traits such as branching, fucosylation, and sialylation. Results In PDAC patients a higher level of branching (OR 6.19,P-value 9.21 x 10(-11)) and (antenna)fucosylation (OR 13.27,P-value 2.31 x 10(-9)) ofN-glycans was found. Furthermore, the ratio of alpha 2,6- vs alpha 2,3-linked sialylation was higher in patients compared to healthy controls. A classification model built with three glycosylation traits was used for discovery (AUC 0.88) and independent validation (AUC 0.81), with sensitivity and specificity values of 0.85 and 0.71 for the discovery set and 0.75 and 0.72 for the validation set. Conclusion SerumN-glycome analysis revealed glycosylation differences that allow classification of PDAC patients from healthy controls. It was demonstrated that glycosylation traits rather than singleN-glycan structures obtained in this clinical glycomics study can serve as a basis for further development of a blood-based diagnostic test.Surgical oncolog

Solid-Phase Extraction Strategies to Surmount Body Fluid Sample Complexity in High-Throughput Mass Spectrometry-Based Proteomics

Proteomic

Automated imaging MS: Toward high throughput imaging mass spectrometry

The term molecular histology has been used to convey the potential of imaging mass spectrometry to describe tissue by its constituent peptides and proteins, and to link this with established histological features. The low throughput of imaging mass spectrometry has been one of the factors inhibiting a full investigation of the clinical potential of molecular histology. Here we report the development of an automated set-up, consisting of a controlled environment sample storage chamber, a sample loading robot, and a MALDI-TOF/TOF mass spectrometer, all controlled by a single user interface. The automated set-up is demonstrated to have the positional stability and experimental reproducibility necessary for its clinical application. (C) 2009 Elsevier B.V. All rights reserved.Proteomic