Abstract
Rheumatoid arthritis is a common and debilitating systemic inflammatory condition affecting up to 1% of the world's population. This study aimed to investigate the immunological significance of O-glycans in chronic arthritis at a local and systemic level. O-Glycans released from synovial glycoproteins during acute and chronic arthritic conditions were compared and immune-reactive glycans identified. The sulfated core 1 O-glycan (Galβ1-3GalNAcol) was immune reactive, showing a different isomeric profile in the two conditions. From acute reactive arthritis, three isomers could be sequenced, but in patients with chronic rheumatoid arthritis, only a single 3-Gal sulfate-linked isomer could be identified. The systemic significance of this glycan epitope was investigated using the salivary mucin MUC7 in patients with rheumatoid arthritis and normal controls. To analyze this low abundance glycan, a selected reaction monitoring (SRM) method was developed to differentiate and relatively quantitate the core 1 O-glycan and the sulfated core 1 O-glycan Gal- and GalNAc-linked isomers. The acquisition of highly sensitive full scan linear ion trap MS/MS spectra in addition to quantitative SRM data allowed the 3- and 6-linked Gal isomers to be differentiated. The method was used to relatively quantitate the core 1 glycans from MUC7 to identify any systemic changes in this carbohydrate epitope. A statistically significant increase in sulfation was identified in salivary MUC7 from rheumatoid arthritis patients. This suggests a potential role for this epitope in chronic inflammation. This study was able to develop an SRM approach to specifically identify and relatively quantitate sulfated core 1 isomers and the unsulfated structure. The expansion of this method may afford an avenue for the high throughput investigation of O-glycans.
Highlights
Glycosylation is a prevalent and significant post-translational modification, exemplified by the existence of a number of congenital and genetic diseases; 64 gene defects have been identified that affect glycan biosynthesis [1]
Ulcerative colitis pathogenesis has been associated with an increase in sulfated O-glycans [13], and sulfated mucins have been identified in patients with Helicobacter pylori infection [14] and in severe ventilatorassociated pneumonia [15]
Identification of Immune-reactive Sulfated Glycans in synovial fluid (SF)— Glycans have been shown to be essential to a range of immune functions
Summary
Glycosylation is a prevalent and significant post-translational modification, exemplified by the existence of a number of congenital and genetic diseases; 64 gene defects have been identified that affect glycan biosynthesis [1]. This can result in quite devastating diseases, such as leukocyte-adhesion deficiency type II, which is caused by a loss of sialyl Lewis x production leading to mental retardation, or WiskottAldrich syndrome, which results from problems with O-linked glycans in lymphocytes, leading to immunodeficiency [2]. Glycosylation changes have been observed in many disease states, with several different carbohydrate groups identified as significant. It leads to an altered immune response and hyperproliferation of the synovial lining, causing debilitating joint pain and swelling [18, 19]
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