Li Guangze, Hu Xiangyu, Ye Xiguang
Department of Orthopedics, Affiliated Aoyang Hospital of Jiangsu University, Zhangjiagang, China.
Department of Orthopedics, Affiliated Hospital of Hubei University of Chinese Medicine, Hubei Provincial Hospital of Traditional Chinese Medicine, Hubei Provincial Institute of Traditional Chinese Medicine, Wuhan, China.
Cell Biochem Biophys. 2025 Mar;83(1):647-656. doi: 10.1007/s12013-024-01497-1. Epub 2024 Dec 14.
Osteoarthritis (OA) is a joint disease closely related to aging and characterized by degeneration of articular cartilage. Robinin is a natural agent with various pharmacological properties. Recently, Robinin has been found to have the potential to improve the bone-related diseases. However, its effect on OA development remained unknown. Here, we discuss the specific role and underlying mechanisms of Robinin in interleukin-1beta (IL-1β)-treated chondrocytes and OA mouse model. Chondrocytes were isolated from the mouse to conduct in vitro assays. We evaluated cell viability and apoptosis using Cell Counting Kit-8 (CCK-8) assay and flow cytometry analysis, respectively. Western blotting assessed the levels of proteins related to apoptosis, extracellular matrix (ECM), and signaling pathways. Immunofluorescence staining was used to detect the expression of ECM and signaling markers. ELISA was conducted to assess the levels of inflammatory markers. The OA mice model was established using surgical destabilization of the medial meniscus (DMM), and then H&E staining and Safranin O staining were conducted to observe the histopathological changes in synovial tissues. TUNEL assay was used to detect cell apoptosis in vivo. Real-time RT-PCR was operated to measure mRNA level in vitro and in vivo. We discovered that Robinin reversed the IL-1β-induced decrease in chondrocyte viability. Robinin suppressed IL-1β-induced apoptosis of chondrocytes. The ECM destruction and inflammatory response induced by IL-1β were markedly reversed by Robinin incubation in the mouse chondrocytes. Besides, the upregulated cytokine mRNA levels in IL-1β-treated chondrocytes were reduced by Robinin treatment. The downregulation of COL2A1 level and upregulation of MMP13 and ADAMTS5 levels were counteracted by Robinin treatment. Robinin reduced the protein levels of Toll-like receptor 2 (TLR2) and Toll-like receptor 4 (TLR4) but enhanced the level of phosphorylated p65 (p-p65) in IL-1β-stimulated chondrocytes and OA mice. Robinin mitigated inflammation, cell apoptosis and cartilage destruction in synovial tissues from the OA mice. In conclusion, Robinin alleviated OA development in vitro and in vivo via TLR2/TLR4/NF-κB signaling pathway.
骨关节炎（OA）是一种与衰老密切相关的关节疾病，其特征为关节软骨退变。刺槐素是一种具有多种药理特性的天然药物。最近，已发现刺槐素具有改善骨相关疾病的潜力。然而，其对骨关节炎发展的影响尚不清楚。在此，我们探讨刺槐素在白细胞介素-1β（IL-1β）处理的软骨细胞和骨关节炎小鼠模型中的具体作用及潜在机制。从小鼠分离软骨细胞进行体外实验。我们分别使用细胞计数试剂盒-8（CCK-8）实验和流式细胞术分析评估细胞活力和凋亡。蛋白质印迹法检测与凋亡、细胞外基质（ECM）和信号通路相关的蛋白质水平。免疫荧光染色用于检测ECM和信号标志物的表达。进行酶联免疫吸附测定（ELISA）以评估炎症标志物水平。采用内侧半月板手术不稳定（DMM）建立骨关节炎小鼠模型，并进行苏木精-伊红（H&E）染色和番红O染色以观察滑膜组织的组织病理学变化。TUNEL实验用于检测体内细胞凋亡。实时逆转录聚合酶链反应（RT-PCR）用于测量体外和体内的mRNA水平。我们发现刺槐素可逆转IL-1β诱导的软骨细胞活力下降。刺槐素抑制IL-1β诱导软骨细胞凋亡。在小鼠软骨细胞中，刺槐素孵育可显著逆转IL-1β诱导的ECM破坏和炎症反应。此外，刺槐素处理可降低IL-1β处理的软骨细胞中细胞因子mRNA水平的上调。刺槐素处理可抵消COL2A1水平的下调以及MMP13和ADAMTS5水平的上调。刺槐素降低IL-1β刺激的软骨细胞和骨关节炎小鼠中Toll样受体2（TLR2）和Toll样受体4（TLR4）的蛋白质水平，但提高磷酸化p-p65的水平。刺槐素减轻骨关节炎小鼠滑膜组织中的炎症、细胞凋亡和软骨破坏。总之，刺槐素通过TLR2/TLR4/NF-κB信号通路在体外和体内减轻骨关节炎的发展。
