Laboratory for Experimental Orthopedics, Department of Orthopedic Surgery, Maastricht University, Maastricht, the Netherlands.
Experimental Rheumatology, Department of Rheumatology, Radboud University Medical Center, Nijmegen, the Netherlands.
Osteoarthritis Cartilage. 2023 Jan;31(1):60-71. doi: 10.1016/j.joca.2022.09.004. Epub 2022 Sep 21.
Since the joint microenvironment and tissue homeostasis are highly dependent on synovial fluid, we aimed to compare the essential chondrocyte signaling signatures of non-osteoarthritic vs end-stage osteoarthritic knee synovial fluid. Moreover, we determined the phenotypic consequence of the distinct signaling patterns on articular chondrocytes.
Protein profiling of synovial fluid was performed using antibody arrays. Chondrocyte signaling and phenotypic changes induced by non-osteoarthritic and osteoarthritic synovial fluid were analyzed using a phospho-kinase array, luciferase-based transcription factor activity assays, and RT-qPCR. The origin of osteoarthritic synovial fluid signaling was evaluated by comparing the signaling responses of conditioned media from cartilage, synovium, infrapatellar fat pad and meniscus. Osteoarthritic synovial fluid induced pathway-phenotype relationships were evaluated using pharmacological inhibitors.
Compared to non-osteoarthritic synovial fluid, osteoarthritic synovial fluid was enriched in cytokines, chemokines and growth factors that provoked differential MAPK, AKT, NFκB and cell cycle signaling in chondrocytes. Functional pathway analysis confirmed increased activity of these signaling events upon osteoarthritic synovial fluid stimulation. Tissue secretomes of osteoarthritic cartilage, synovium, infrapatellar fat pad and meniscus activated several inflammatory signaling routes. Furthermore, the distinct pathway signatures of osteoarthritic synovial fluid led to accelerated chondrocyte dedifferentiation via MAPK/ERK signaling, increased chondrocyte fibrosis through MAPK/JNK and PIK/AKT activation, an elevated inflammatory response mediated by cPKC/NFκB, production of extracellular matrix-degrading enzymes by MAPK/p38 and PIK/AKT routes, and enabling of chondrocyte proliferation.
This study provides the first mechanistic comparison between non-osteoarthritic and osteoarthritic synovial fluid, highlighting MAPKs, cPKC/NFκB and PIK/AKT as crucial OA-associated intracellular signaling routes.
由于关节微环境和组织内稳态高度依赖滑液，我们旨在比较非骨关节炎和终末期骨关节炎膝关节滑液中基本软骨细胞信号特征。此外，我们还确定了不同信号模式对关节软骨细胞表型的影响。
使用抗体芯片对滑液进行蛋白质谱分析。通过磷酸激酶阵列、基于荧光素酶的转录因子活性测定和 RT-qPCR 分析非骨关节炎和骨关节炎滑液诱导的软骨细胞信号和表型变化。通过比较软骨、滑膜、髌下脂肪垫和半月板的条件培养基的信号反应来评估骨关节炎滑液信号的来源。使用药理学抑制剂评估骨关节炎滑液诱导的途径-表型关系。
与非骨关节炎滑液相比，骨关节炎滑液富含细胞因子、趋化因子和生长因子，这些因子在软骨细胞中引发了不同的 MAPK、AKT、NFκB 和细胞周期信号。功能途径分析证实，骨关节炎滑液刺激后这些信号事件的活性增加。骨关节炎软骨、滑膜、髌下脂肪垫和半月板的组织分泌液激活了几个炎症信号途径。此外，骨关节炎滑液的独特途径特征通过 MAPK/ERK 信号导致软骨细胞去分化加速，通过 MAPK/JNK 和 PIK/AKT 激活导致软骨细胞纤维化增加，通过 cPKC/NFκB 介导的炎症反应增加，通过 MAPK/p38 和 PIK/AKT 途径产生细胞外基质降解酶，并使软骨细胞增殖。
本研究首次对非骨关节炎和骨关节炎滑液进行了机制比较，突出了 MAPKs、cPKC/NFκB 和 PIK/AKT 作为骨关节炎相关的关键细胞内信号途径。
