| Summary: | Disruption of the Wnt canonical signalling leads to the development of osteoarthritis both in human and in mice but the underlying molecular mechanisms are poorly understood. Both forced activation and blockade of Wnt/β catenin signalling lead to cartilage breakdown. This study attempts to unravel the mechanisms leading to such paradox and is based on the hypothesis that WNT-3A triggers multiple signalling pathways simultaneously, with distinct outcomes. WNT-3A-stimulation induced activation of Wnt/β-catenin pathway in articular chondrocytes and promoted proliferation and loss of chondrocytes phenotype markers, such as COL2A1, Aggrecan and SOX9 mRNA. However, whereas the inhibition of the Wnt/β-catenin pathway by DKK1 rescued the proliferative effect of WNT-3A, it did not rescue the loss of chondrocyte phenotype but, paradoxically, it further enhanced it. Therefore I tested the possibility that WNT-3A-induced chondrocyte de-differentiation could be mediated by other WNT pathways independent of β-catenin. Indeed, in AHAC WNT-3A induced intracellular calcium mobilization and phosphorylation and nuclear localization of CaMKII in a G-protein dependent manner, suggesting the activation of the Wnt/CaMKII pathway. Inhibition of the Wnt/CaMKII pathway rescued the loss of the phenotypic markers SOX9 and COL2A1 induced by WNT-3A, indicating that this pathway drives WNT-3A-induced chondrocyte de-differentiation. Finally, my data show that the Wnt/β-catenin and the Wnt/CaMKII pathways are mutually inhibitory, explaining why both exogenous WNT-3A and its inhibitor DKK1 lead to chondrocyte de-differentiation: the first through direct activation of CaMKII, and the second indirectly by removal of the inhibition of CaMKII exerted by the β-catenin pathway. My results show for the first time that a single WNT ligand can simultaneously activate at least two different pathways in the same cells with different outcomes. These findings highlight the possibility to therapeutically target individual outcomes of Wnt signalling, for instance to prevent chondrocyte de-differentiation without affecting crucial anabolic processes such as cell proliferation.
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