Changes of IgG N-Glycosylation in Thyroid Autoimmunity: The Modulatory Effect of Methimazole in Graves’ Disease and the Association With the Severity of Inflammation in Hashimoto’s Thyroiditis

Abstract

The N-glycome of immunoglobulin G (IgG), the most abundant glycoprotein in human blood serum, reflects pathological conditions of autoimmunity and is sensitive to medicines applied in disease therapy. Due to the high sensitivity of N-glycosylation, the IgG N-glycan profile may serve as an indicator of an ongoing inflammatory process. The IgG structure and its effector functions are strongly dependent on the composition of N-glycans attached to the Fc fragment, and the binding of antigens is regulated by Fab sugar moieties. Because of the crucial role of N-glycans in IgG function, remodeling of its N-oligosaccharides can induce pathological changes that ultimately contribute to the development of autoimmunity; restoration of their physiological structure is critical to the reduction of disease symptoms. Our recently published data have shown that the pathology of autoimmune thyroid diseases (AITDs), including Hashimoto’s thyroiditis (HT) and Graves’ disease (GD), is accompanied by alterations of the composition of IgG N-glycans. The present study is a more in-depth investigation of IgG glycosylation in both AITDs, designed to determine the relationship between the severity of thyroid inflammation and IgG N-glycan structures in HT, and to assess the impact of immunosuppressive therapy on the N-glycan profile in GD patients. The study material consisted of human serum samples collected from donors with elevated anti-thyroglobulin (Tg) and/or anti-thyroperoxidase (TPO) IgGs without symptoms of hypothyroidism (n=68), HT patients characterized by high autoantibody titers and advanced destruction of the thyroid gland (n=113), GD patients with up-regulated IgG against thyroid-stimulating hormone receptor (TSHR) before (n=62) and after (n=47) stabilization of TSH level as a result of methimazole therapy (study groups), and healthy donors (control group, n=90). IgG was isolated from blood serum using protein G affinity chromatography. N-glycans were released from IgG by PNGase F digestion and analyzed by ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) after 2-aminobenzamide (2-AB) labeling. UPLC-MS chromatograms were integrated into 25 peaks (GP) in the Waters UNIFI Scientific Information System, and N-glycans were assigned based on the glucose unit values and mass-to-charge ratios (m/z) of the detected ions. The Kruskal-Wallis non-parametric test was used to determine the statistical significance of the results (p<0.05). The obtained results suggest that modifications of IgG sialylation, galactosylation and core-fucosylation are associated with the severity of HT symptoms. Methimazole therapy implemented in GD patients affected the IgG N-glycan profile; as a result, the content of the sialylated and galactosylated oligosaccharides with core fucose differed after treatment. Our results suggest that N-glycosylation of IgG undergoes dynamic changes during the intensification of thyroiditis in HT, and that in GD autoimmunity it is affected significantly by immunosuppressive therapy.

The role of N-glycosylation in B-cell biology and IgG activity. The aspects of autoimmunity and anti-inflammatory therapy

“…Changes in IgG Fuc content during progression and remission of autoimmunity have been shown in SLE. Glycosylation analysis using Aurelia aurantia (AAL) and Lens culinaris (LCA) lectins in modified ELISA showed higher levels of the Fuc in IgG from SLE patients compared to healthy donors, and normalisation of IgG fucosylation in disease remission (145). Our study of immunoglobulin glycome in thyroid autoimmune diseases (AITD) by ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) revealed the remodeling of IgG fucosylation in Hashimoto's thyroiditis patients.…”

Galactia lindenii lectin type-II. Proposal of its potential use in diagnostic tools

“…The common autoimmune thyroid disorder HT is characterized by marked lymphocyte and plasma cell in ltration of the parenchyma and antibodies speci c to thyroid antigens [28, [154][155][156]. The thyroid structure can be destroyed by activated T-lymphocytes inducing chronic in ammation and the late-stage disease can resemble the histology of lymphatic tissue [156,157]. In ammatory in ltrates in tumor tissue may represent a condition preceding the development of malignancy, and that is how HT can be frequently observed in PTC, and immune dysregulation is involved in both disorders [158].…”

“…Increased levels of terminal mono-and disaccharide α/β-GalNAc glycotopes are present in HT patient´s thyroid tissue and blood [16,28,77]. However, unlike HT tissues, low levels of α1,2-Fuc have been detected in the glycan antennas of HT peripheral blood mononuclear cells (PBMCs) [90], and high levels of sialylated glycans are present in the sera of HT patients with advanced thyroid destruction [157]. The increase in serum glycoconjugate sialylation is related to the presence of the β-galactoside α2,6sialyltransferase 1 (ST6Gal1), which is active in the bloodstream of the HT patients, independently from the classical pathway of cellular glycosylation in ER and Golgi [157,161].…”

“…However, unlike HT tissues, low levels of α1,2-Fuc have been detected in the glycan antennas of HT peripheral blood mononuclear cells (PBMCs) [90], and high levels of sialylated glycans are present in the sera of HT patients with advanced thyroid destruction [157]. The increase in serum glycoconjugate sialylation is related to the presence of the β-galactoside α2,6sialyltransferase 1 (ST6Gal1), which is active in the bloodstream of the HT patients, independently from the classical pathway of cellular glycosylation in ER and Golgi [157,161]. In this regard, it has been pointed out that the process of IgG sialylation may explain the great dynamics of in ammatory processes mediated by antibodies in HT [161].…”

Study on the Treatment of ITP Mice with IVIG Sourced from Distinct Sex-Special Plasma (DSP-IVIG)

“…The main component of IVIG is immunoglobulin G (IgG). Following papain hydrolysis, classical IgG antibody consists of one molecule of fragment crystallizable (Fc) and two molecules of fragment of antigen binding (Fab) fragments [4,5].…”