Truncated lubricin glycans in osteoarthritis stimulate the synoviocyte secretion of VEGFA, IL-8, and MIP-1α: Interplay between O-linked glycosylation and inflammatory cytokines

Abstract

The primary aim of the study was to identify inflammatory markers relevant for osteoarthritis (OA)-related systemic (plasma) and local (synovial fluid, SF) inflammation. From this, we looked for inflammatory markers that coincided with the increased amount of O-linked Tn antigen (GalNAcα1-Ser/Thr) glycan on SF lubricin. Inflammatory markers in plasma and SF in OA patients and controls were measured using a 44-multiplex immunoassay. We found consistently 29 markers detected in both plasma and SF. The difference in their concentration and the low correlation when comparing SF and plasma suggests an independent inflammatory environment in the two biofluids. Only plasma MCP-4 and TARC increased in our patient cohort compared to control plasma. To address the second task, we concluded that plasma markers were irrelevant for a direct connection with SF glycosylation. Hence, we correlated the SF-inflammatory marker concentrations with the level of altered glycosylation of SF-lubricin. We found that the level of SF-IL-8 and SF-MIP-1α and SF-VEGFA in OA patients displayed a positive correlation with the altered lubricin glycosylation. Furthermore, when exposing fibroblast-like synoviocytes from both controls and OA patients to glycovariants of recombinant lubricin, the secretion of IL-8 and MIP-1α and VEGFA were elevated using lubricin with Tn antigens, while lubricin with sialylated and nonsialylated T antigens had less or no measurable effect. These data suggest that truncated glycans of lubricin, as found in OA, promote synovial proinflammatory cytokine production and exacerbate local synovial inflammation.

Tryptase β regulation of joint lubrication and inflammation via proteoglycan-4 in osteoarthritis

“…7j) demonstrated that adding tryptase β alone to human synovial fluid is sufficient to induce a change a gene expression in NF-κB1 and RELA. In addition, we previously demonstrated that altered truncated PRG4 glycans can stimulate the synoviocyte secretion of vascular endothelial growth factor a (VEGFA), IL8 and CCL3 65 . Recombinant PRG4 can induce NF-κB activation and requires tryptase β activity.…”

A Novel Fluorescent Immuno-Lectin Assay to Identify Osteoarthritis Associated Glycoforms of Lubricin

“…The change of plasma glycosylation in OA with decreased amount of SNA epitopes is quite different compared to OA regarding SF lubricin where the glycans are shown to be more truncated and expected to have more Tn antigens compared to controls 10 . The origin of this lubricin SNA depleted glycoform found in the OA-cohort ‘s plasma is unknown and whether it is caused by alterations in catabolic and/or anabolic processes.…”

“…In OA, there is an increase of truncated structures such as Tn-antigen (GalNAcα1-Ser/Thr) in SF lubricin 10 . Lubricin with truncated glycosylation acts as a signaling molecule that stimulates synoviocytes influencing the level of inflammatory cytokines, chemokines, and growth factors 10 .…”

“…In this report we set out to investigate the different glycoforms of lubricin found in SF 10 and plasma, in respect to OA. In order to detect differences in glycosylation we developed and validated a Fluorescent Immuno-Lectin Assay (FILA), to ensure that we had the sensitivity required.…”

“…The glycosylation of lubricin occurs on its STP-rich domain, that have shown to harbor more than 160 binding sites for O- linked glycosylation including sialylated/non sialylated core-1 O -glycans (Galβ1-3GalNAcα1-), and core-2 structures such as NeuAcα2-3Galβ1-3(NeuAcα2-3Galβ1-4GlcNAcβ1-6)GalNAcα1- 28,29 . In OA, there is an increase of truncated structures such as Tn-antigen (GalNAcα1-Ser/Thr) in SF lubricin 10 . Lubricin with truncated glycosylation acts as a signaling molecule that stimulates synoviocytes influencing the level of inflammatory cytokines, chemokines, and growth factors 10 .…”