Glycan structures of lubricin, a lubricating component of synovial fluid, are different in rheumatoid arthritis and osteoarthritis, with implications for inflammatory processes.
Osteoarthritis (OA) and rheumatoid arthritis (RA) cause debilitating joint pain, and degeneration of cartilage can be detected in both diseases by the deposition of proteoglycan fragments into synovial fluid. However, their etiologies and disease manifestations differ significantly; RA is an autoimmune disease characterized by inflammation of the synovium, whereas OA lacks an inflammatory component and is a consequence of age-related wear. Niclas Karlsson and colleagues investigated the acidic glycoprotein and proteoglycan fraction from synovial fluid samples from OA and RA patients, and unexpectedly identified lubricin (proteoglycan 4, PRG4) as the dominant component. Furthermore, they found that glycosylation of PRG4 is different in OA and RA.
The acidic protein fraction — isolated by anion exchange chromatography — formed less than 2% of the total protein content of the synovial fluid. Of this fraction, one major band after gel electrophoresis was stained by Alcian Blue, which detects acidic-type oligosaccharides and sulfated glycosaminoglycans. With the use of liquid chromatography coupled with tandem mass spectrometry (LC–MS2), this was identified as PRG4, a highly glycosylated boundary lubricant that covers the cartilage surface.
PRG4 was named as a proteoglycan in part due to a single consensus sequence for the attachment of chondroitin sulfate. It is known to contain mucin-type core 1 oligosaccharides, but detailed information about its glycosylation was lacking. The authors analyzed PRG4 glycans by LC–MS2 and identified at least nine different O-linked structures, including disialylated core 1 and core 2, and sulfated core 2 oligosaccharides.
In RA samples, there was a higher proportion of disialylated tetrasaccharide compared with singly sialylated tetrasaccharide on PRG4, with the reverse true of OA samples. This suggests a higher expression level of sialyltransferases in RA, which leads to a less complex type of O-linked glycosylation because the disialylated structures cannot be modified further in the Golgi. This is a common feature of cancer and inflammation, and in the case of PRG4 in RA it may be a protective response to combat disease progression: increased sialylation increases the negative charge around PRG4, increasing the repellant force and enhancing lubrication of joints. Alternatively, decreased sialylation in OA may be an exacerbating factor in the disease.
Sulfated core 2 oligosaccharides from PRG4 were detected in the OA samples, possibly reflecting the greater capacity for modification compared with the more highly sialylated RA oligosaccharides. A glycan epitope that forms part of the L-selectin ligand structure was also found on PRG4, but no fucosylated structures were detected. Leukocyte rolling along endothelial cell surfaces is regulated by fucosylation of L-selectin ligands, but it is not known whether sulfation of oligosaccharides has a role in inflammation. The non-fucosylated sulfated PRG4 oligosaccharide might be unique to inflammation of joint tissues.
Proteoglycans such as aggrecan are the focus of arthritic biomarker research and thus it was surprising that PRG4 was more abundant in synovial fluid. The authors speculate that loss of PRG4 at the cartilage surface could occur before the loss of extra cellular matrix components. The sample size of this study was small, and, as healthy synovial samples are difficult to obtain, there was no control available for comparison. However, this research could lead to novel biomarkers for RA, and will help to understand the enigmatic cause of synovial inflammation.
Original research paper
- Estrella, R. P., Whitelock, J. M., Packer, N. H. and Karlsson, N. G.The glycosylation of human synovial lubricin: implications for its role in inflammation. Biochem. J. (published online 5 May 2010) doi:10.1042/bj20100360 | Article
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