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Objective: Maturity-onset diabetes of the young (MODY) due to variants in HNF1A is the most common type of monogenic diabetes. Frequent misdiagnosis results in missed opportunity to use sulfonylureas as first-line treatment. A nongenetic biomarker could improve selection of subjects for genetic testing and increase diagnosis rates. We previously reported that plasma levels of antennary fucosylated N -glycans and high-sensitivity C-reactive protein (hs-CRP) are reduced in individuals with HNF1A-MODY. In this study, we examined the potential use of N -glycans and hs-CRP in discriminating individuals with damaging HNF1A alleles from those without HNF1A variants in an unselected population of young adults with nonautoimmune diabetes., Research Design and Methods: We analyzed the plasma N -glycan profile, measured hs-CRP, and sequenced HNF1A in 989 individuals with diabetes diagnosed when younger than age 45, persistent endogenous insulin production, and absence of pancreatic autoimmunity. Systematic assessment of rare HNF1A variants was performed., Results: We identified 29 individuals harboring 25 rare HNF1A alleles, of which 3 were novel, and 12 (in 16 probands) were considered pathogenic. Antennary fucosylated N -glycans and hs-CRP were able to differentiate subjects with damaging HNF1A alleles from those without rare HNF1A alleles. Glycan GP30 had a receiver operating characteristic curve area under the curve (AUC) of 0.90 (88% sensitivity, 80% specificity, cutoff 0.70%), whereas hs-CRP had an AUC of 0.83 (88% sensitivity, 69% specificity, cutoff 0.81 mg/L)., Conclusions: Half of rare HNF1A sequence variants do not cause MODY. N -glycan profile and hs-CRP could both be used as tools, alone or as adjuncts to existing pathways, for identifying individuals at high risk of carrying a damaging HNF1A allele., (© 2018 by the American Diabetes Association.)

Background: Statins are among the most widely prescribed medications worldwide and usually many individuals involved in clinical and population studies are on statin therapy. Immunoglobulin G (IgG) glycosylation has been associated with numerous cardiometabolic risk factors., Methods: The aim of this study was to investigate the possible association of statin use with N-glycosylation of IgG. The association was analyzed in two large population cohorts (TwinsUK and KORA) using hydrophilic interaction liquid chromatography (HILIC-UPLC) in the TwinsUK cohort and reverse phase liquid chromatography coupled with electrospray mass spectrometry (LC-ESI-MS) in the KORA cohort. Afterwards we investigated the same association for only one statin (rosuvastatin) in a subset of individuals from the randomized double-blind placebo-controlled JUPITER study using LC-ESI-MS for IgG glycome and HILIC-UPLC for total plasma N-glycome., Results: In the TwinsUK population, the use of statins was associated with higher levels of core-fucosylated biantennary glycan structure with bisecting N-acetylglucosamine (FA2B) and lower levels of core-fucosylated biantennary digalactosylated monosialylated glycan structure (FA2G2S1). The association between statin use and FA2B was replicated in the KORA cohort. In the JUPITER trial we found no statistically significant differences between the randomly allocated placebo and rosuvastatin groups., Conclusions: In the TwinsUK and KORA cohorts, statin use was associated with a small increase of pro-inflammatory IgG glycan, although this finding was not confirmed in a subset of participants from the JUPITER trial., General Significance: Even if the association between IgG N-glycome and statins exists, it is not large enough to pose a problem for glycomic studies., (Crown Copyright © 2017. Published by Elsevier B.V. All rights reserved.)

Glycosylation of membrane proteins plays a crucial role in various physiological events, including intercellular recognition and intermolecular interactions on the cell surface (Gornik et al., Biochim Biophys Acta 1820:1318-1326, 2012). To study composition and function of N-glycans on membrane proteins one has to have an efficient and reproducible analytical method, which includes protein extraction and analysis of glycans. In this chapter we provide an analytical approach that includes cloud-point extraction (CPE) of total membrane proteins with the non-ionic detergent Triton X-114 and subsequent analysis of their N-glycans using hydrophilic interaction liquid chromatography (HILIC)-UPLC/HPLC. The protocol presented here can be used for parallel analysis of both membrane and intracellular proteins.

Low back pain (LBP) is a common debilitating condition which aetiology and pathogenesis are poorly understood. We carried out a first so far analysis of associations between LBP and plasma IgG N-glycome in a sample of 4511 twins from TwinsUK database assessed for LBP, lumbar disc degeneration (LDD) as its possible cause, and IgG-glycan levels. Using weighted correlation network analysis, we established a correlation between LBP and glycan modules featured by glycans that either promote or block antibody-dependent cell-mediated cytotoxicity (ADCC). The levels of four glycan traits representing two of those modules were statistically significantly different in monozygotic twins discordant for LBP. Also, the trend to higher prevalence of systemic inflammatory disorders was shown for twins with low level of fucosylated glycans and high level of non-fucosylated glycans. Core fucosylation of IgG is a "safety switch" reducing ADCC, thus our results suggest the involvement of ADCC and associated inflammation in pathogenesis of LBP. No correlation between LDD scores and glycans was found assuming that the inflammation may not be a part of LDD. These data provide a new insight into understanding the complex pathophysiology of LBP and suggest glycan levels as a possible biomarker for inflammation-related subtypes of LBP.

Glycans constitute the most abundant and diverse form of the post-translational modifications, and animal studies have suggested the involvement of IgG glycosylation in mechanisms of renal damage. Here, we explored the associations between IgG glycans and renal function in 3274 individuals from the TwinsUK registry. We analyzed the correlation between renal function measured as eGFR and 76 N-glycan traits using linear regressions adjusted for covariates and multiple testing in the larger population. We replicated our results in 31 monozygotic twin pairs discordant for renal function. Results from both analyses were then meta-analyzed. Fourteen glycan traits were associated with renal function in the discovery sample (P<6.5×10(-4)) and remained significant after validation. Those glycan traits belong to three main glycosylation features: galactosylation, sialylation, and level of bisecting N-acetylglucosamine of the IgG glycans. These results show the role of IgG glycosylation in kidney function and provide novel insight into the pathophysiology of CKD and potential diagnostic and therapeutic targets., (Copyright © 2016 by the American Society of Nephrology.)

Protein glycosylation is the most common epiproteomic modification involved in numerous physiological and pathological processes. Previous studies reported strong associations between human plasma N-glycans and age, prompting us to evaluate the potential application of this biological phenomenon in the field of forensics. Blood from 526 blood donors from different parts of Croatia was collected on bloodstain cards during the period 2004-2007 and stored at 4°C for 6-9 years. Glycosylation profiles of the bloodstains were analysed using hydrophilic interaction ultra performance liquid chromatography (HILIC-UPLC) and divided into 38 glycan groups (GP1-GP38). A statistically significant correlation between N-glycan profiles of bloodstains and chronological age was found and a statistical model that can be used for the age prediction was designed (Age = 75.59 - 5.15 × (GP4)(2)+ 17.07 × GP6 - 5.30 × (GP10)(2) - 16.56 × GP16 + 20.07 × GP20 - 7.54 × (GP20)(2) + 16.47 × GP22). This model explains 47.78% of the variation in age, with a prediction error of 9.07 years. Our findings demonstrate that analysing the N-glycan profile could be a new tool in forensics, offering an approximate human age estimation from dried bloodstains found at a crime scene.