BCP crystals promote chondrocyte hypertrophic differentiation in OA cartilage by sequestering Wnt3a
BCP crystals promote chondrocyte hypertrophic differentiation in OA cartilage by sequestering Wnt3a
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DOI：
10.1136/annrheumdis-2019-216648
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发表时间：
2020-07-01
影响因子：
27.4
通讯作者：
Pap, Thomas
中科院分区：
文献类型：
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作者：
Bertrand, Jessica;Kraft, Tabea;Pap, Thomas
目的碱性磷酸钙(BCP)结晶的软骨钙化是骨关节炎(OA)过程中的常见现象。它与疾病的严重程度直接相关，并已知与软骨细胞的肥大分化有关。WNT3a是调控肥大软骨细胞分化的一种形态因子。方法在6~22周的小鼠和不同级别的人软骨中观察软骨细胞外基质的钙化和硫酸盐化。WNT3a和β-catenin在人和小鼠软骨中均有表达。用定量逆转录聚合酶链式反应(RT-PCR)分析硫酸调节酶(HS2St1、HS6St1)的表达。应用标记基因Axin2、Sox9、Col2、MMP13、ColX和Aggrecan的定量RT-PCR检测BCP晶体对软骨细胞表型的影响。用β-连环蛋白和pLRP6的蛋白质印迹法研究了Wnt信号的激活。用免疫组织化学染色和Western blotting分析BCP与WNT3a的结合能力。结果在这里，我们报道了人和小鼠骨关节炎细胞周围基质硫酸盐化增加。WNT3a与硫酸乙酰肝素蛋白多糖共定位于骨性关节炎软骨细胞的细胞周基质中，激活了典型的Wnt信号。在体外，BCP晶体与WNT3a物理结合。有趣的是，根据LRP6的磷酸化和β-连环蛋白的稳定化，BCP晶体足以诱导典型的Wnt信号，并诱导软骨细胞表型的肥厚性转变。结论因此，我们的数据证实BCP晶体是Wnt3a在细胞周围基质中的集中因子和软骨细胞肥大的诱导者。
Objective Calcification of cartilage with basic calcium phosphate (BCP) crystals is a common phenomenon during osteoarthritis (OA). It is directly linked to the severity of the disease and known to be associated to hypertrophic differentiation of chondrocytes. One morphogen regulating hypertrophic chondrocyte differentiation is Wnt3a.Methods Calcification and sulfation of extracellular matrix of the cartilage was analysed over a time course from 6 to 22 weeks in mice and different OA grades of human cartilage. Wnt3a and beta-catenin was stained in human and murine cartilage. Expression of sulfation modulating enzymes (HS2St1, HS6St1) was analysed using quantitative reverse transcription PCR (RT-PCR). The influence of BCP crystals on the chondrocyte phenotype was investigated using quantitative RT-PCR for the marker genes Axin2, Sox9, Col2, MMP13, ColX and Aggrecan. Using western blot for beta-catenin and pLRP6 we investigated the activation of Wnt signalling. The binding capacity of BCP for Wnt3a was analysed using immunohistochemical staining and western blot.Results Here, we report that pericellular matrix sulfation is increased in human and murine OA. Wnt3a co-localised with heparan sulfate proteoglycans in the pericellular matrix of chondrocytes in OA cartilage, in which canonical Wnt signalling was activated. In vitro, BCP crystals physically bound to Wnt3a. Interestingly, BCP crystals were sufficient to induce canonical Wnt signalling as assessed by phosphorylation of LRP6 and stabilisation of beta-catenin, and to induce a hypertrophic shift of the chondrocyte phenotype.Conclusion Consequently, our data identify BCP crystals as a concentrating factor for Wnt3a in the pericellular matrix and an inducer of chondrocyte hypertrophy.