Figure 4

IL-1β-mediated and load-mediated regulation of Wnt/β-catenin signaling in chondrocytes from FrzB^−/−and wild-type mice. Cultured articular chondrocytes from FrzB^−/− mice or wild-type (WT) mice were treated for 24 hours with interleukin (IL)-1β (1 ng/ml). Results from the IL-1β-treated samples were normalized to the control ones (Ctrl), so that the graphs represent the fold-induction in response to IL-1β. (A) Ctnnb1 gene expression (coding β-catenin) and Lef1 gene expression (a Wnt/β-catenin target gene) were analyzed by real-time polymerase chain reaction (PCR; n = 4 and n = 3, respectively). Ctnnb1 gene expression was not modulated by IL-1β in WT chondrocytes and Lef1 gene expression was decreased (P = 0.05). In contrast, the treatment induced Ctnnb1 and Lef1 gene expression in FrzB^−/− chondrocytes (P = 0.014 and P = 0.05, respectively). IL-1β-mediated regulation of β-catenin expression was thus different between FrzB^−/− and WT (P = 0.057 for Ctnnb1, P = 0.05 for Lef1). (B) Intracellular extracts were analyzed for total β-catenin by western blotting. Quantification of β-catenin blot suggested that IL-1β-mediated β-catenin accumulation was different between FrzB^−/− and WT (P = 0.10, n = 4). (C) FrzB^−/− cartilage explants or WT explants were subjected to dynamic compression for 6 hours (0.5 Hz, 1 MPa). Ctnnb1 gene expression was analyzed by real-time PCR (n = 3). Results from the loaded cartilage explants were normalized to those from the corresponding nonloaded explants (Ctrl), so that the graphs represent the fold-induction in response to compression. Ctnnb1 gene expression was not affected by load in WT explants but was increased 2.7 fold in compressed FrzB^−/− samples (P = 0.05). The load-induced increase in Ctnnb1 mRNA tends to be enhanced in FrzB^−/− explants compared with WT explants (P = 0.20). Bars represent the mean ± standard error of the mean, *P ≤ 0.05 versus Ctrl, #P ≤ 0.10 between FrzB^−/− and WT. FrzB, frizzled-related protein B; KO, knockout.