Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing, 100700, China.
School of Pharmacy, Anhui University of Traditional Chinese Medicine, Hefei, 230012, China.
J Ethnopharmacol. 2022 Nov 15;298:115611. doi: 10.1016/j.jep.2022.115611. Epub 2022 Aug 8.
A Chinese patent medicine derived from a classical traditional Chinese medicine formula, Yu-Xue-Bi tablet (YXB) is widely used in the clinic to treat rheumatoid arthritis (RA). During the progression of RA, angiogenesis plays a central role in fostering the production of inflammatory cells, leading to synovial hyperplasia and bone destruction. However, whether YXB attenuates the angiogenesis during RA progression remains to be defined.
We aimed to evaluate the anti-angiogenic activity of YXB and explore its mechanism of action in collagen-induced arthritis (CIA) rats and VEGF-induced HUVECs.
Transcriptional regulatory network analysis and a network pharmacology approach were employed to explore mechanism of YXB in RA angiogenesis. The antiarthritic effect of YXB was evaluated by determining the arthritis incidence, and score, and by micro-CT analysis. The anti-angiogenic effect of YXB in vivo was assessed by histological and immunohistochemical analyses. The anti-angiogenic effect of YXB in vitro was assessed by wound healing, Transwell migration, Transwell invasion, and tube formation assays. Western-blotting and immunohistochemical analysis were employed to explore the molecular mechanisms of YXB.
YXB reduced disease severity and ameliorated pathological features in CIA rats. YXB markedly decreased bone destruction and synovial angiogenesis. Consistently, we also demonstrated that YXB effectively suppressed angiogenesis marker CD31 and VEGF expression. In vitro, YXB effectively inhibited HUVEC migration, invasion, and tube formation. Following the identification of transcriptional expression profiles, "YXB putative targets-known RA-related genes-genes associated with the therapeutic effect of YXB" interaction network was constructed and analyzed. After that, the LOX/Ras/Raf-1 signaling axis, which is involved in RA angiogenesis, was identified as one of the candidate mechanisms of YXB against RA. Experimentally, YXB dose-dependently decreased the expression levels of LOX, Ras, and Raf-1, as well as the phosphorylation of MEK and ERK in CIA rats, and these effects were better than the inhibitory effects of methotrexate (MTX), an FDA approved drug used for some autoimmune diseases such as RA. In addition, YXB may function as a potent angiogenesis inhibitor and significantly suppress the VEGF-induced activation of LOX/Ras/Raf-1 signaling in vitro.
We provide evidence that YXB may decrease the disease severity of RA and reduce bone erosion by suppressing angiogenesis via inhibition of LOX/Ras/Raf-1 signaling.
愈血痹片（YXB）是一种源自经典中药配方的中成药，广泛应用于临床治疗类风湿关节炎（RA）。在 RA 的进展过程中，血管生成在促进炎性细胞产生、导致滑膜增生和骨破坏方面起着核心作用。然而，YXB 是否能减轻 RA 进展过程中的血管生成仍有待确定。
我们旨在评估 YXB 的抗血管生成活性，并探讨其在胶原诱导性关节炎（CIA）大鼠和 VEGF 诱导的 HUVECs 中的作用机制。
采用转录调控网络分析和网络药理学方法探讨 YXB 在 RA 血管生成中的作用机制。通过测定关节炎发病率和评分以及 micro-CT 分析来评估 YXB 的抗关节炎作用。通过组织学和免疫组织化学分析评估 YXB 在体内的抗血管生成作用。通过划痕愈合、Transwell 迁移、Transwell 侵袭和管形成实验评估 YXB 在体外的抗血管生成作用。采用 Western-blotting 和免疫组织化学分析探讨 YXB 的分子机制。
YXB 降低了 CIA 大鼠的疾病严重程度并改善了病理特征。YXB 显著减少了骨破坏和滑膜血管生成。一致地，我们还表明 YXB 有效抑制了血管生成标志物 CD31 和 VEGF 的表达。体外，YXB 有效抑制了 HUVEC 的迁移、侵袭和管形成。在鉴定转录表达谱后，构建并分析了“YXB 假定靶点-已知 RA 相关基因-YXB 治疗作用相关基因”相互作用网络。之后，确定了 LOX/Ras/Raf-1 信号通路，该通路参与 RA 血管生成，是 YXB 治疗 RA 的候选机制之一。实验中，YXB 剂量依赖性地降低了 CIA 大鼠中 LOX、Ras 和 Raf-1 的表达水平以及 MEK 和 ERK 的磷酸化水平，其作用优于甲氨蝶呤（MTX）的抑制作用，MTX 是一种 FDA 批准的用于治疗某些自身免疫性疾病（如 RA）的药物。此外，YXB 可能作为一种有效的血管生成抑制剂，通过抑制 LOX/Ras/Raf-1 信号通路显著抑制 VEGF 诱导的体外激活。
我们提供的证据表明，YXB 可能通过抑制 LOX/Ras/Raf-1 信号通路来降低 RA 的疾病严重程度并减少骨质侵蚀。
