Background Obesity, a major global health problem, is associated with increased cardiometabolic morbidity and mortality. Protein glycosylation is a frequent posttranslational modification, highly responsive to inflammation and ageing. The prospect of biological age reduction, by changing glycosylation patterns through metabolic intervention, opens many possibilities. We have investigated whether weight loss interventions affect inflammation- and ageing-associated IgG glycosylation changes, in a longitudinal cohort of bariatric surgery patients. To support potential findings, BMI-related glycosylation changes were monitored in a longitudinal twins cohort. Methods IgG N-glycans were chromatographically profiled in 37 obese patients, subjected to low-calorie diet, followed by bariatric surgery, across multiple timepoints. Similarly, plasma-derived IgG N-glycan traits were longitudinally monitored in 1680 participants from the TwinsUK cohort. Results Low-calorie diet induced a marked decrease in the levels of IgG N-glycans with bisecting GlcNAc, whose higher levels are usually associated with ageing and inflammatory conditions. Bariatric surgery resulted in extensive alterations of the IgG N-glycome that accompanied progressive weight loss during 1-year follow-up. We observed a significant increase in digalactosylated and sialylated glycans, and a substantial decrease in agalactosylated and core fucosylated IgG N-glycans (adjusted p value range 7.38 × 10−04–3.94 × 10−02). This IgG N-glycan profile is known to be associated with a younger biological age and reflects an enhanced anti-inflammatory IgG potential. Loss of BMI over a 20 year period in the TwinsUK cohort validated a weight loss-associated agalactosylation decrease (adjusted p value 1.79 × 10−02) and an increase in digalactosylation (adjusted p value 5.85 × 10−06). Conclusions Altogether, these findings highlight that weight loss substantially affects IgG N-glycosylation, resulting in reduced glycan and biological age.

OBJECTIVE N-glycosylation is a functional posttranslational modification of immunoglobulins (Igs). We hypothesized that specific IgG N-glycans are associated with incident type 2 diabetes and cardiovascular disease (CVD). RESEARCH DESIGN AND METHODS We performed case-cohort studies within the population-based European Prospective Investigation into Cancer and Nutrition (EPIC)–Potsdam cohort (2,127 in the type 2 diabetes subcohort [741 incident cases]; 2,175 in the CVD subcohort [417 myocardial infarction and stroke cases]). Relative abundances of 24 IgG N-glycan peaks (IgG-GPs) were measured by ultraperformance liquid chromatography, and eight glycosylation traits were derived based on structural similarity. End point–associated IgG-GPs were preselected with fractional polynomials, and prospective associations were estimated in confounder-adjusted Cox models. Diabetes risk associations were validated in three independent studies. RESULTS After adjustment for confounders and multiple testing correction, IgG-GP7, IgG-GP8, IgG-GP9, IgG-GP11, and IgG-GP19 were associated with type 2 diabetes risk. A score based on these IgG-GPs was associated with a higher diabetes risk in EPIC-Potsdam and independent validation studies (843 total cases, 3,149 total non-cases, pooled estimate per SD increase 1.50 [95% CI 1.37–1.64]). Associations of IgG-GPs with CVD risk differed between men and women. In women, IgG-GP9 was inversely associated with CVD risk (hazard ratio [HR] per SD 0.80 [95% CI 0.65–0.98]). In men, a weighted score based on IgG-GP19 and IgG-GP23 was associated with higher CVD risk (HR per SD 1.47 [95% CI 1.20–1.80]). In addition, several derived traits were associated with cardiometabolic disease incidence. CONCLUSIONS Selected IgG N-glycans are associated with cardiometabolic risk beyond classic risk factors, including clinical biomarkers.

Protein glycosylation is the attachment of a carbohydrate moiety to a protein backbone affecting both structure and function of the protein. Abnormal glycosylation is associated with various diseases, and some of the changes in glycosylation are detectable even before symptom development. As such, glycans have emerged as compelling new biomarker candidates. A wide range of analytical methods exist for small-scale glycan analyses. However, there is a growing need for highly robust and reproducible high-throughput techniques that allow for large-scale glycoprofiling. Here we describe the evaluation of robustness and repeatability of immunoglobulin G (IgG) N-glycan analysis using the GlycoWorks RapiFluor-MS N-Glycan Kit followed by hydrophilic interaction ultra-high-performance liquid chromatography (HILIC-UHPLC) from 335 technical replicates of human plasma randomly distributed across 67 96-well plates. The data was collected over a five-month period using multiple UHPLC systems and chromatographic columns. Following relative IgG N-glycan quantification in acquired chromatograms, data analysis showed that the most abundant peaks that together made up for three fourths of the detected IgG N-glycome all had coefficients of variation (CVs) lower than 2 percent. The highest CVs ranging from 16 to 29 percent accompanied low abundance glycan peaks with the individual relative peak area below 1 percent that together made up for less than 2 percent of the detected IgG N-glycome. These results show that the tested method is very robust and repeatable, making it suitable for the IgG N-glycan analysis of a large number of samples in a high-throughput manner over a longer period of time.

Objectives: Glycosylation of immunoglobulin G (IgG) is an important regulator of the immune system and has been implicated in prevalent hypertension.The aim of this study is to investigate whether the IgG glycome begins to change prior to hypertension diagnosis by analysing the IgG glycome composition in a large population-based female cohort with two independent replication samples.Methods: We included 989 unrelated cases with incident hypertension and 1628 controls from the TwinsUK cohort (mean follow-up time of 6.3 years) with IgG measured at baseline by ultra-performance liquid chromatography and longitudinal BP measurement available. We replicated our findings in 106 individuals from the 10 001 Dalmatians and 729 from KORA S4. Cox regression mixed models were applied to identify changes in glycan traits preincident hypertension, after adjusting for age, mean arterial pressure, BMI, family relatedness and multiple testing (FDR < 0.1). Significant IgG-incident hypertension associations were replicated in the two independent cohorts by leveraging Cox regression mixed models in the 10 001 Dalmatians and logistic regression models in the KORA cohort.Results: We identified and replicated four glycan traits, incidence of bisecting GlcNAc, GP4, GP9 and GP21, that are predictive of incident hypertension after adjusting for confoundes and multiple testing [hazard ratio (95% CI) ranging from 0.45 (0.24-0.84) for GP21 to 2.9 (1.5-5.68) for GP4]. We then linearly combined the four replicated glycans and found that the glycan score correlated with incident hypertension, SBP and DBP. Conclusion:Our results suggest that the IgG glycome changes prior to the development of hypertension.

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Down syndrome (DS) is a genetic condition caused by trisomy 21 (T21) and characterized by a range of neurological symptoms including intellectual disability, early onset of neurodegeneration and dementia, some of which can be attributed to accelerated aging. N-glycosylation is a protein modification that plays a critical role in numerous biological processes and its dysregulation is associated with a wide range of diseases, in some even before the onset of symptoms. N-glycosylation of total plasma proteins, as well as specific plasma proteins, such as immunoglobulin G, has been shown to change in DS, displaying an accelerated aging phenotype consistent with the various symptoms of premature aging that occur in DS. However, little is known about how T21 affects the N-glycosylation of other cellular proteins. To better understand how T21 affects N-glycosylation during neural differentiation, we characterized and compared the total released N-glycans of induced pluripotent stem cells (iPSCs) and their neural stem cell (NSC) derivatives. We analyzed six different isogenic clones all derived from a single individual with mosaic DS and thus all sharing the same genetic background; however, three had a normal disomic karyotype (D21), while the other three had an additional copy of chromosome 21 (T21). We characterized the total cell N-glycosylation profiles using ultra high performance liquid chromatography (UHPLC) and subsequent tandem mass spectrometry analysis to determine proposed glycan structures. Our results revealed both qualitative and quantitative differences in the composition of N-glycomes between iPSCs and NSCs, with NSCs showing a higher amount of complex N-glycans and a lower amount of mannosidic N-glycans when compared to iPSCs. Moreover, we found differences in N-glycosylation patterns between D21 and T21 cells. Notably, T21 cells exhibited a significant increase in the amount of pseudohybrid N-glycans. Trisomy 21 also caused a significant decrease in the abundance of a hybrid monoantennary fucosylated glycan (H6N3F1). Our findings define the released N-glycan profile of total cells for both D21 and T21 iPSCs and NSCs and suggest that the presence of a third copy of chromosome 21 impacts N-glycosylation patterns already in the stem cell state.

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