Department of Orthopaedic Surgery, Otto-von-Guericke University, Magdeburg, Germany.
Chemical Genomics Centre of the Max-Planck Society, Dortmund, Germany.
Osteoarthritis Cartilage. 2018 Jun;26(6):818-823. doi: 10.1016/j.joca.2018.02.908. Epub 2018 Mar 17.
The canonical Wnt signaling pathway has been shown to be involved in regulating chondrocyte hypertrophic differentiation during Osteoarthritis (OA). The aim of this study was to test the therapeutic potential of two stapled peptide canonical Wnt inhibitors - SAH-Bcl9 and StAx-35R - in preventing Wnt induced cartilage changes in OA.
Primary neonatal murine chondrocytes and cartilage explants from OA patients undergoing total joint replacement for knee OA, were used for microscopy to determine matrix and cell penetrating capacity of fluorescein isothiocyanate FITC-tagged SAH-Bcl9 and StAx-35R peptides. T cell factor/lymphoid enhancer-binding factor (TCF/LEF) reporter assays were used to monitor the inhibition of Wnt3a induced β-catenin signaling by each peptide. Changes in chondrocyte phenotypic marker gene expression were analyzed by qRT PCR.
Both peptides localized intercellular in primary murine chondrocytes and cartilage explants. They inhibited Wnt3a induced TCF/LEF promoter activity in primary murine chondrocytes. Both inhibitors did not rescue Wnt3a altered expression of chondrocyte phenotypic genes (Sox9, Col2a1, Acan) and hypertrophy marker gene (Col10a1) at high doses (100 ng/ml). Upon application of 10 ng/ml Wnt3a, StAx-35R partially reversed the Wnt effect on Sox9 and Col2a1 gene expression. Both peptides, however, reversed the downregulation of SOX9 and aggrecan (ACAN), and decrease of COL10A1 gene expression in preserved human OA cartilage explants.
These data indicate that blockade of canonical Wnt signaling might be a therapeutic strategy to treat early OA cases and protect further cartilage degradation by preventing chondrocyte hypertrophic differentiation.
经典 Wnt 信号通路已被证明参与调节骨关节炎（OA）中软骨细胞的肥大分化。本研究旨在测试两种订书肽经典 Wnt 抑制剂 - SAH-Bcl9 和 StAx-35R - 在预防 OA 中 Wnt 诱导的软骨变化中的治疗潜力。
使用原代新生鼠软骨细胞和来自接受全膝关节置换术的 OA 患者的软骨外植体进行显微镜检查，以确定荧光素异硫氰酸酯 FITC 标记的 SAH-Bcl9 和 StAx-35R 肽的基质和细胞穿透能力。使用 T 细胞因子/淋巴增强因子结合因子（TCF/LEF）报告基因测定来监测每种肽对 Wnt3a 诱导的β-连环蛋白信号的抑制作用。通过 qRT-PCR 分析软骨细胞表型标记基因的表达变化。
两种肽均在原代鼠软骨细胞和软骨外植体中定位于细胞间。它们抑制 Wnt3a 诱导的原代鼠软骨细胞中 TCF/LEF 启动子活性。两种抑制剂在高剂量（100ng/ml）下均不能挽救 Wnt3a 改变的软骨细胞表型基因（Sox9、Col2a1、Acan）和肥大标志物基因（Col10a1）的表达。在应用 10ng/ml Wnt3a 时，StAx-35R 部分逆转了 Wnt 对 Sox9 和 Col2a1 基因表达的影响。然而，两种肽均逆转了 SOX9 和聚集蛋白（ACAN）的下调，以及保存的人 OA 软骨外植体中 COL10A1 基因表达的降低。
这些数据表明，阻断经典 Wnt 信号可能是治疗早期 OA 病例并通过防止软骨细胞肥大分化来保护进一步软骨降解的治疗策略。
