Background/Purpose: Blockade of syndecan-4 (Sdc4) signaling protects mice from cartilage degradation in experimentally induced osteoarthritis (OA). Cartilage damage results in changes of chondrocyte phenotype induced by various signaling pathways including the activation of WNT signaling. Here, we hypothesized that Sdc4 modulates chondrocyte phenotype by the specific modulation of WNT signaling.

Methods: In vitro analyses were performed using neonatal wild type (wt) and Sdc4-/- chondrocytes, or a blocking Sdc4 antibody on wt chondrocytes. The influence of 100 ng/ ml WNT3a on glycosaminoglycan (GAG) production was analyzed using alcian blue staining of micromass cultures. The expression of chondrocyte marker genes (aggrecan, collagen 2, MMP13) was measured by quantitative RT-PCR. The effects of WNT3a on canonical and noncanonical WNT signaling were analyzed using Western Blot detection of ß-catenin, pLRP-6, pCamKII and pJNK and a TCF/LEF reporter assay. To investigate the in vivo relevance of the investigated pathways induction of OA in wt and Sdc4-/- mice was performed using the DMM model and stainings for ß-catenin and pCamKII were performed.

Results:

Micromass cultures revealed a higher basal GAG production by Sdc4-/- than wt chondrocytes. Furthermore, WNT3a stimulation led to a decrease in GAG in wt, which was not observed in Sdc4-/- chondrocytes. These findings were confirmed by a 10x higher basal production of aggrecan and collagen 2 in Sdc4-/- compared to wt chondrocytes. The expression of both genes was 10 fold increased by stimulation with WNT3a, whereas WNT3a led to a decrease in the expression of both genes in wt chondrocytes. Inverse results were found for MMP13, which was significantly less expressed in Sdc4-/- chondrocytes and was not upregulated upon WNT3a stimulation. Western blot analyses of canonical WNT signaling showed that ß catenin is strongly reduced and not upregulated upon stimulation with WNT3a in Sdc4-/- chondrocytes. Confirming these findings we also found less phosphorylation of LRP6 and less activation of the TCF/Lef reporter upon WNT3a stimulation in Sdc4-/- chondrocytes. Noncanonical WNT signaling was increased in Sdc4-/- under basal conditions, but decreased upon WNT3a stimulation in Sdc4-/- and increased in wt chondrocytes. The same blockade of canonical and downregulation of noncanonical WNT signaling upon WNT stimulation were obtained by using a blocking Sdc-4 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: We conclude from our data that Sdc4 is a major regulator of cellular response to WNT through the prevention of the induction of canonical WNT signaling. Therefore, we suggest that the blockade of Sdc-4 protects from OA induced changes in chondrocyte phenotype by inhibiting WNT induced differentiation of chondrocytes.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/syndecan-4-regulates-activation-of-wnt-signaling-in-chondrocytes/