骨痹通方调控Wnt/β-catenin信号通路影响骨关节炎软骨细胞肥大和凋亡的机制研究

Osteoarthritis (OA) is joint disease with high rate of disability which lacks of effective treatments. The pathogenesis of pain, swelling and deformation in OA is deficiency of liver and kidney, binding of phlegm and blood stasis. Our previous studies showed that Gu Bi Tong Recipe with the effect of invigorating liver and kidney, reducing phlegm and resolving masses can relieve pain and swelling of the joints of OA. The mechanism is related to inhibition of abnormal apoptosis of hypertrophic chondrocytes and reduction of expression of β-catenin in cartilage.Therefore, we put forward the scientific hypothesis: Gu Bi Tong Recipe could protect articular cartilage and delay the progress of the disease by inhibiting the abnormal hypertrophy and apoptosis of chondrocytes based on regulation of Wnt/-catenin signaling pathway. Animal experiments will be applied in this study to observe the effect of Gu Bi Tong Recipe on expression of key upstream molecules of Wnt/ beta-catenin pathway to clarify its target; and then cell experiments will be used with signaling pathway blockers and activators to explore the regulation of Wnt/ beta-catenin pathway to clarify the effect of Gu Bi Tong Recipe on articular cartilage hypertrophy and apoptosis and in order to detect the scientific connotation of treatment of OA by Gu Bi Tong Recipe which is invigorating liver and kidney, reducing phlegm and resolving masses.

骨关节炎（OA）在关节疾病中致残率高，缺乏有效的治疗手段。OA关节疼痛、肿胀变形的病机是肝肾亏虚、痰瘀互结。课题组前期研究表明，具有补益肝肾、化痰散结作用的骨痹通方可有效缓解OA关节疼痛和肿胀。其机制与抑制软骨细胞异常肥大和凋亡、降低软骨β-catenin表达有关。因此，我们提出科学假说：骨痹通方通过调节Wnt/β-catenin信号通路抑制关节软骨细胞异常肥大和凋亡，进而发挥软骨保护作用，延缓病情进展。本研究拟通过动物实验，观察骨痹通方对Wnt/β-catenin通路上游关键信号分子表达的影响，明确其作用靶点；进而通过细胞实验，借助该信号通路阻断剂及激动剂，探索骨痹通方调控Wnt/β-catenin通路抑制关节软骨异常肥大和凋亡、保护软骨的机制，进而阐释骨痹通方补益肝肾、化痰散结治疗OA的科学内涵。

骨痹通方是名老中医经验方，可有效缓解OA关节疼痛和肿胀。其机制与抑制软骨细胞异常肥大和凋亡、降低软骨β-catenin表达有关。本研究通过动物实验，观察骨痹通方对Wnt/β-catenin通路上游关键信号分子表达的影响，明确其作用靶点；进而通过细胞实验，借助该信号通路阻断剂，探索骨痹通方调控Wnt/β-catenin通路、抑制关节软骨异常肥大和凋亡、保护软骨的机制，阐释骨痹通方补益肝肾、化痰散结治疗OA的科学内涵。动物实验证实骨痹通方能够有效减轻Hulth手术建立的OA大鼠模型软骨基质退变及软骨细胞凋亡，并通过降低Wnt5a、β-catenin、Bax、MMP-3的表达来抑制Wnt/β-catenin信号通路的异常激活。细胞试验发现骨痹通方能够有效抑制经IL-1β激活的Wnt/β-catenin信号通路，降低Wnt5a、β-catenin、MMP3的表达，促进DKK-1、FRZB的表达，从而抑制了Wnt/β-catenin信号通路的激活。使用XAV-939阻断Wnt/β-catenin信号通路后，骨痹通方仍能够进一步抑制β-catenin、MMP3的表达，提示骨痹通方除抑制Wnt/β-catenin通路外可能仍具有其他途径发挥对软骨的保护作用。进一步探究表明，骨痹通方对NF-κB信号通路也具有一定的抑制作用。综上所述，骨痹通方通过降低Wnt5a、β-catenin的表达，抑制OA造成的Wnt/β-catenin 信号通路异常激活，下调MMP3及Bax在蛋白和基因水平的表达，从而发挥保护关节软骨及减少软骨细胞凋亡的作用。