Article
Syndecan-4 modulates the WNT signaling pathway in chondrocytes and thereby controls the phenotype
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Authors
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Annelena Held
- Institut für Experimentelle Muskuloskelettale Medizin, Münster
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Charlotte Clarke
- Institut für Experimentelle Muskuloskelettale Medizin, Münster
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Richard Stange
- Dept. of Trauma, Hand and Reconstructive Surgery, Münster
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Giovanna Nalesso
- Centre for Experimental Medicine and Rheumatology Queen Mary London, London, UK
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Joanna Sherwood
- Centre for Experimental Medicine and Rheumatology Queen Mary London, London, UK
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Lars Godmann
- IEMM Universitätsklinikum Münster, Münster
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Frank Echtermeyer
- Dept. of Anaesthesiology and Intensive Care Medicine, Hannover
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Francesco Dell' Accio
- Centre for Experimental Medicine & Rheumatology William Harvey Research Institute, Barts and The London School of Medicine and Dentistry John Vane Science Centre, London EC1M 6BQ, UK, United Kingdom
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Thomas Pap
- Universitätsklinikum Münster, Westfälische Wilhelms-Universität, Institut für Experimentelle Muskuloskelettale Medizin, Münster
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Jessica Bertrand
- Universitätsklinikum Münster, Westfälische Wilhelms-Universität, Institut für Experimentelle Muskuloskelettale Medizin, Münster
| Published: | September 12, 2014 |
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Outline
Text
Background: Blockade of syndecan-4 (Sdc4) signaling protects mice from cartilage degradation in experimentally induced osteoarthritis (OA). OA induces hypertrophic differentiation of chondrocytes and matrix remodeling. Various signaling pathways including the WNT signaling pathway may trigger this induction of chondrocyte differentiation. We hypothesized that Sdc4 controls the chondrocyte phenotype by specific modulation of WNT signaling pathways.
Methods: In vitro analyses were performed using neonatal wild type (wt) and Sdc4-/- chondrocytes, or blocking antibodies against Sdc4. The influence of WNT3a on glycosaminoglycan (GAG) production was analyzed using alcian blue staining of micromass cultures. Expression of marker genes (aggrecan, collagen2, MMP13) was measured by quantitative RT-PCR. Effects of WNT3a on canonical and noncanonical WNT signaling were analyzed using Western Blot and TCF/LEF reporter assay. In vivo relevance was investigated upon induction of OA using the DMM model.
Results: Micromass cultures revealed a higher basal GAG production by Sdc4-/- chondrocytes. WNT3a stimulation led to a decrease in GAG production in wt cells, which was absent in Sdc4-/- chondrocytes. Quantitative RT-PCR showed a 10x higher basal production of aggrecan and collagen2 in Sdc4-/- chondrocytes. WNT3a increased the expression of both genes in Sdc-4 -/-, whereas it decreased their expression in wt chondrocytes. MMP13 was significantly less expressed in Sdc4-/- chondrocytes and, unlike in wt cells, was not upregulated upon WNT3a stimulation. Western blot showed that β-catenin is strongly reduced and not upregulated upon stimulation with WNT3a in Sdc4-/- chondrocytes. LRP6 was less phosporylated and the TCF/Lef promotor showed reduced activation upon WNT3a stimulation in Sdc4-/- chondrocytes. pCamKII was increased in Sdc4-/- under basal conditions, but was decreased upon WNT3a stimulation in Sdc4-/-. The same effects on canonical and noncanonical WNT signaling upon WNT stimulation were obtained by using a anti-Sdc-4 blocking antibody. In vivo analyses of canonical WNT signaling confirmed the decreased basal activation and the missing increase after induction of OA in Sdc4-/- mice.
Conclusion: Sdc4 is a major regulator of the cellular response to WNT signals through facilitating the induction of the canonical WNT signaling pathway. The blockade of Sdc-4 protects from OA induced changes in chondrocyte phenotype by inhibiting WNT induced differentiation of chondrocytes.
