新风胶囊通过结合Wnt5a经Wnt/β-catenin信号通路抑制类风湿性关节炎

Xinfeng capsule inhibits rheumatoid arthritis by binding to Wnt5a via Wnt/β-catenin signaling pathway

目的:本研究将探讨Wnt5a是否可以作为类风湿性关节炎(RA)潜在的诊断和治疗靶点,新风胶囊(XFC)如何通过Wnt5a/β-catenin信号通路改善RA。方法:在体内Adjuvant arthritis(AA)大鼠模型中采用ELISA和RT-qPCR检测炎症因子和病理基因研究XFC对AA大鼠疗效,RT-qPCR检测验证XFC调控通过网络药理学预测的核心基因和关键通路。在体外原代AA成纤维样滑膜细胞(FLS)中采用RT-qPCR、Western blot和免疫荧光等方法研究XFC对Wnt/β-catenin通路的调节机制。结果:XFC显著下调AA大鼠的关节炎评分和足爪肿胀,抑制AA大鼠关节炎症。XFC降低AA大鼠外周血中炎症因子TNF-α和IL-1水平,抑制AA大鼠关节滑膜和AA FLS中病理基因MMP3和fibronectin水平。网络药理学预测出Wnt通路与XFC治疗RA高度相关。细胞水平上,含XFC血清抑制Wnt通路相关基因β-catenin、CCND1和c-Myc的表达。分子对接结果显示XFC关键成分与Wnt5a的结合能力强,在AA FLS中Wnt5a过表达(Wnt5a-ove)干扰了XFC的作用。结论:Wnt5a在AA FLS和RA FLS中表达明显升高,XFC通过与Wn5a结合,抑制Wnt/β-catenin信号通路活化改善RA,为XFC改善RA提供新的治疗机制。

AIM:This study will clarify whether Wnt5a can be used as a potential diagnostic and therapeutic target for rheumatoid arthritis(RA)and how Xinfeng capsule(XFC)can improve RA through the Wnt5a/β-catenin signaling pathway.METHODS:ELISA and RT-qPCR were used to detect inflammatory factors and pathological genes in the rat model of AA in vivo to investigate the effect of XFC on AA rats.RT-qPCR was used to verify the core genes and key pathways of XFC regulation predicted by network pharmacology.The regulatory mechanism of XFC on Wnt/β-catenin pathway was elucidated by RT-qPCR.Western blot and immunofluorescence in primary AA fibroblast-like synovial cells(FLS)in vitro.RESULTS:XFC significantly decreased the arthritis score and paw swelling in AA rats,and inhibited joint inflammation in AA rats.XFC decreased the levels of inflammatory factors TNF-αand IL-1 in peripheral blood of AA rats,and inhibited the levels of pathological genes MMP3 and fibronectin in joint synovium and AA FLS of AA rats.Network pharmacology predicts that the Wnt pathway is highly correlated with XFC treatment of RA.At the cellular level,serum containing XFC inhibited the expression of Wnt pathway-related genesβ-catenin,CCND1 and c-Myc.The molecular docking results showed that the key components of XFC had strong binding ability to Wnt5a,and the overexpression of Wnt5a(Wnt5a-ove)in AA FLS interfered with the action of XFC.CONCLUSION:The expression of Wnt5a is significantly increased in AA FLS and RA FLS,and XFC can inhibit the activation of Wnt/β-catenin signaling pathway to improve RA by binding with Wn5a,providing a new therapeutic mechanism for XFC to improve RA.