Low back pain (LBP) is the symptom of a group of syndromes with heterogeneous underlying mechanisms and molecular pathologies, making treatment selection and patient prognosis very challenging. Moreover, symptoms and prognosis of LBP are influenced by age, gender, occupation, habits, and psychological factors. LBP may be characterized by an underlying inflammatory process. Previous studies indicated a connection between inflammatory response and total plasma N-glycosylation. We wanted to identify potential changes in total plasma N-glycosylation pattern connected with chronic low back pain (CLBP), which could give an insight into the pathogenic mechanisms of the disease. Plasma samples of 1128 CLBP patients and 760 healthy controls were collected in clinical centers in Italy, Belgium and Croatia and used for N-glycosylation profiling by hydrophilic interaction ultra-performance liquid chromatography (HILIC-UPLC) after N-glycans release, fluorescent labeling and clean-up. Observed N-glycosylation profiles have been compared with a cohort of 126 patients with acute inflammation that underwent abdominal surgery. We have found a statistically significant increase in the relative amount of high-branched (tri-antennary and tetra-antennary) N-glycan structures on CLBP patients' plasma glycoproteins compared to healthy controls. Furthermore, relative amounts of disialylated and trisialylated glycan structures were increased, while high-mannose and glycans containing bisecting N-acetylglucosamine decreased in CLBP. Observed changes in CLBP on the plasma N-glycome level are consistent with N-glycosylation changes usually seen in chronic inflammation. To our knowledge, this is a first large clinical study on CLBP patients and plasma N-glycome providing a new glycomics perspective on potential disease pathology.

Immunosuppressants and biologicals are widely used therapeutics for various chronic inflammatory diseases (CID). To gain more detailed insight into their downstream effects, we examined their impact on serum immunoglobulin G (IgG) glycosylation. We analyzed IgG subclass-specific fragment crystallizable (Fc) N-glycosylation in patients suffering from various CID using the LC-MS approach. Firstly, we compared IgG Fc N-glycosylation between 128 CID patients and 204 healthy controls. Our results replicated previously observed CID-related decrease in IgG Fc galactosylation (adjusted p-value range 1.70 × 10⁻²–5.95 × 10⁻²²) and sialylation (adjusted p-value range 1.85 × 10⁻²–1.71 × 10⁻¹⁸). Secondly, to assess changes in IgG Fc N-glycosylation associated with therapy and remission status, we compared 139 CID patients receiving either azathioprine, infliximab, or vedolizumab therapy. We observed an increase in IgG Fc galactosylation (adjusted p-value range 1.98 × 10⁻²–1.30 × 10⁻¹⁵) and sialylation (adjusted p-value range 3.28 × 10⁻⁶–4.34 × 10⁻¹⁸) during the treatment. Furthermore, patients who reached remission displayed increased Fc galactosylation levels (p-value range 2.25 × 10⁻²–5.44 × 10⁻³) in comparison to patients with active disease. In conclusion, the alterations in IgG Fc glycosylation and the fact these changes are even more pronounced in patients who achieved remission, suggest modulation of IgG inflammatory potential associated with CID therapy.

Objectives: Hypertension and type 2 diabetes mellitus comorbidity (HDC) is common, which confers a higher risk of cardiovascular disease than the presence of either condition alone. Describing the underlying glycomic changes of immunoglobulin G (IgG) that predispose individuals to HDC may help develop novel protective immune-targeted and anti-inflammatory therapies. Therefore, we investigated glycosylation changes of IgG associated with HDC. Methods: The IgG N-glycan profiles of 883 plasma samples from the three northwestern Chinese Muslim ethnic minorities and the Han Chinese were analyzed by ultra-performance liquid chromatography instrument. Results: We found that 12 and six IgG N-glycan traits showed significant associations with HDC in the Chinese Muslim ethnic minorities and the Han Chinese, respectively, after adjustment for potential confounders and false discovery rate. Adding the IgG N-glycan traits to the baseline models, the area under the receiver operating characteristic curves (AUCs) of the combined models differentiating HDC from hypertension (HTN), type 2 diabetes mellitus (T2DM), and healthy individuals were 0.717, 0.747, and 0.786 in the pooled samples of Chinese Muslim ethnic minorities, and 0.828, 0.689, and 0.901 in the Han Chinese, respectively, showing improved discriminating performance than both the baseline models and the glycan-based models. Conclusion: Altered IgG N-glycan profiles were shown to associate with HDC, suggesting the involvement of inflammatory processes of IgG glycosylation. The alterations of IgG N-glycome, illustrated here for the first time in HDC, demonstrate a biomarker potential, which may shed light on future studies investigating their potential for monitoring or preventing the progression from HTN or T2DM towards HDC.

Aim: Alpha-1-acid glycoprotein (AGP) is a highly glycosylated protein in human plasma and one of the most abundant acute phase proteins in humans. Glycosylation plays a crucial role in its biological functions, and alterations in AGP N-glycome have been associated with various diseases and inflammatory conditions. However, large-scale studies of AGP N-glycosylation in the general population are lacking. Methods: Using recently developed high-throughput glycoproteomic workflow for site-specific AGP N-glycosylation analysis, 803 individuals from the Croatian island of Korcula were analyzed and their AGP N-glycome data associated with biochemical and physiological traits, as well as different environmental factors. Results: After regression analysis, we found that AGP N-glycosylation is strongly associated with sex, somewhat less with age, along with multiple biochemical and physiological traits (e.g. BMI, triglycerides, uric acid, glucose, smoking status, fibrinogen). Conclusion: For the first time we have extensively explored the inter-individual variability of AGP N-glycome in a general human population, demonstrating its changes with sex, age, biochemical, and physiological status of individuals, providing the baseline for future population and clinical studies.

Immunoglobulin G (IgG) is the most abundant serum antibody and is a key determinant of the humoral immune response. Its structural characteristics and effector functions are modulated through the attachment of various sugar moieties called glycans. IgG N-glycome patterns change with the age of individual and in different diseases. Variability of IgG glycosylation within a population is well studied and is affected by a combination of genetic and environmental factors. However, global inter-population differences in IgG glycosylation have never been properly addressed. Here we present population-specific N-glycosylation patterns of whole IgG, analysed in 5 different populations totalling 10,482 IgG glycomes, and of IgG’s fragment crystallisable region (Fc), analysed in 2,530 samples from 27 populations sampled across the world. We observed that country of residence associates with many N-glycan features and is a strong predictor of monogalactosylation variability. IgG galactosylation also strongly correlated with the development level of a country, defined by United Nations health and socioeconomic development indicators. We found that subjects from developing countries had low IgG galactosylation levels, characteristic for inflammation and ageing. Our results suggest that citizens of developing countries may be exposed to country-specific environmental factors that can cause low-grade chronic inflammation and the apparent increase in biological age.