Suppression of β-catenin and matrix metalloproteinase expression levels in chondrocyte-specific Lrp5-knockout mice. (A) Col2a1-Cre-transgenic mice were backcrossed with Lrp5fl/fl mice to generate Lrp5fl/fl;Col2a1-Cre mice. RT-PCR was used to examine deletion of the Lrp5 gene in cartilage versus brain, heart and bone. WT: Wild type. (B) Cartilage destruction in Lrp5fl/fl;Col2a1-cre and control LRP5fl/fl mice subjected to sham operations and destabilization of the medial meniscus (DMM) surgery was evaluated by Safranin O staining and Mankin score (n = 15 independent experimental animals). Scale bar: 50 μm. (C) The expression levels of LRP5, β-catenin, matrix metalloproteinase 3 (MMP3) and MMP13 in sham-operated control cartilage and DMM-operated osteoarthritic cartilage of LRP5fl/fl;Col2a1-cre and LRP5fl/fl mice were determined by immunohistochemical staining. Scale bar: 50 μm. (D) Primary cultures of articular chondrocytes isolated from WT and Lrp5-/- mice were treated with the indicated amounts of anti-Fas antibody for 6 hours or treated with 1 ng/ml IL-1β for 24 hours and with 0.1 μg/ml of anti-Fas antibody for an additional 6 hours. Apoptotic cells were quantified by fluorescence-activated cell sorting analysis (n = 5), Con: Control; IgG: Immunoglobulin G. (E) Apoptotic chondrocytes were visualized by 4′,6-diamidino-2-phenylindole (DAPI) staining and terminal deoxynucleotidyl transferase deoxyuridine triphosphate nick end labeling (TUNEL) assays of cartilage samples from Lrp5fl/fl;Col2a1-cre and control LRP5fl/fl mice after sham operations and DMM surgery. Values are expressed as means ± SEM (*P < 0.05, **P < 0.001). Scale bar: 50 μm.