Understanding HS6ST1 and heparan sulfate 6-Osulfation in cartilage

McClurg, Oliver (2024) Understanding HS6ST1 and heparan sulfate 6-Osulfation in cartilage. Doctoral thesis, University of East Anglia.

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Abstract

Osteoarthritis (OA) is a common, degenerative musculoskeletal disease for which treatment options are lacking due to limited understanding of the molecular pathogenesis of joint degradation and repair. OA is characterised by loss of aggrecan and type II collagen from the joint, partly due to dysregulated growth factor signalling. Heparan sulfate (HS) proteoglycans in the pericellular matrix surrounding chondrocytes regulates growth factor signalling through their negatively-charged sulfate groups. HS 6-O-sulfation regulates the signalling of many growth factors and is therefore tightly regulated, being the only sulfate group that is modified during and after biosynthesis. In OA, HS 6-O-sulfation is increased due to increased expression of the 6-O-sulfotransferase, HS6ST1. Therefore, it has been hypothesised that increased 6-O-sulfation, driven by HS6ST1, contributes to OA progression.

In this thesis, I set out to investigate the effect of HS6ST1 on the C28/I2 chondrocyte-like cell line. HS6ST1 overexpression reduced sulfated glycosaminoglycan (sGAG) content in vitro, which could be reversed by addition of exogenous ligands (TGF-b1, BMP4). This suggests that 6-O-sulfated HS sequesters an endogenous anabolic ligand, which can be competed off by exogenous HS ligands. Label-free quantitative mass spectrometry of healthy and OA human cartilage identified proteins with high affinity for 6-O-sulfated HS, but candidates chosen for further study (endostatin, SERPINF1) could not account for the reduced sGAG. RNASeq functional enrichment analyses suggest that HS6ST1-overexpresison reduced cell-ECM interactions, driven by reduced integrin expression. This was accompanied by reduced phosphorylation of several signalling pathways, including SRC kinases and BMP4-induced SMAD1/5/8 phosphorylation.

Therefore, I propose that increased HS 6-O-sulfation is deleterious for cartilage as it inhibits growth factor signalling, and so downregulates expression of integrins, reducing mechano-responses of chondrocytes and impairing their attachment to the matrix and sGAG accumulation. Targeting HS 6-O-sulfation may therefore be of therapeutic benefit in OA.

Item Type:	Thesis (Doctoral)
Faculty \ School:	Faculty of Medicine and Health Sciences > Norwich Medical School
Depositing User:	Chris White
Date Deposited:	30 May 2024 07:37
Last Modified:	30 May 2024 07:37
URI:	https://ueaeprints.uea.ac.uk/id/eprint/95334
DOI:

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