基于网络药理学的断藤益母汤治疗类风湿性关节炎的整合机制研究

As known, Rheumatoid arthritis (RA) has become a challenge of diagnosis and treatment to the western and traditional Chinese medicine both. In this research, we have always insisted on the combination of classic theory of TCM with clinical practice and developed a definitely effective empirical formula on RA─duanteng yimu tang (DTYMT), whose material basis is still completely unknown. By the applications of UFLC-Q-TOF-MS/MS and serum pharmacochemistry techniques, we preliminary obtained 18 possible active components in DTYMT and its 304 potential gene targets with the method of network pharmacology. On the basis of our early study, the aim of this research is as followed: Firstly, in the vitro experiment of RA, we shall further screen these active components by the observation of the proliferation and apoptosis changes in synovial cells and its target gene expression that the traditional Chinese medicine monomers might cause; Then, in the RA mice experiment in vitro, the curative effect of the screened active components in vivo experiment on the RA mice and expression changes of target genes; Finally, we will integrate and analyze the relations of “DTYMT—active components —target genes—disease (RA)” by the method of network pharmacology and identify the active components of DTYMT and its target gene and finally clarity the integrate mechanism of “multicomponent—multitarget—multipath” in DTYMT, which would not only lay foundations for the further original new drug for RA, but also supply new thread and method for the modernization of traditional Chinese compound recipe .

类风湿性关节炎（RA）是当今中西医学诊疗的疑难疾病。本课题组长期坚持经典理论结合临床实践，研制出治疗RA有确切疗效的经验方——断藤益母汤，但其物质基础尚未阐明。我们前期应用UFLC-Q-TOF-MS/MS技术和血清药物化学方法，并结合网络药理学方法，初步获得断藤益母汤的18个可能活性成分及其304个基因靶点。本项目拟在前期基础上，首先利用RA体外细胞模型，观察上述中药单体对滑膜细胞增殖、凋亡的影响及靶基因的表达变化，进一步筛选活性成分； 其次，应用RA小鼠模型，观察体外有效的活性成分对小鼠模型的整体疗效及靶基因的表达变化；最后，应用网络药理学方法进行“益母断藤汤—活性成分—基因靶点—疾病（RA）”的整合分析，明确断藤益母汤的活性成分及其作用靶点，最终阐明断藤益母汤的“多成分—多靶点—多途径”整合机制，为其临床应用提供科学依据，还将为进一步研发RA创新药物奠定基础。

类风湿关节炎（Rheumatoid Arthritis，RA）是一种临床常见的关节炎性疾病，以关节滑膜炎为主要病理特征，易造成关节破坏和关节畸形，发病率较高，我国RA人群有450~500 万左右。断藤益母汤是长期坚持经典理论结合临床实践总结出的经验方，临床疗效确切且毒副作用小，但是其重要活性成分和作用靶点及作用机制尚未阐明，制约了复方的深度开发与临床应用。.研究采用网络药理学方法分析断藤益母汤治疗RA的重要活性成分和关键靶点。以类风湿关节炎成纤维样滑膜细胞（RA FLSs）和破骨细胞及胶原诱导性关节炎（CIA）小鼠模型进行体外和体内实验，验证断藤益母汤及其重要活性成分治疗RA的作用及其具体机制。.初步筛选出断藤益母汤治疗RA的重要活性成分29个，重要的作用靶点42个。断藤益母汤重要活性成分益母草碱通过阻断MAPK和NF-κB通路减轻了RA成纤维样滑膜细胞的迁移和侵袭能力，并减轻了CIA小鼠关节炎。另一活性成分山奈酚通过阻断MAPK通路抑制类风湿关节炎成纤维细胞样滑膜细胞的迁移和侵袭，通过抑制PI3K/Akt通路作用于周期蛋白及凋亡蛋白等靶点诱导RA成纤维样滑膜细胞的凋亡，也减轻了CIA小鼠关节炎症和病理破坏。断藤益母汤复方介导NF-κB和PI3K/Akt通路抑制成纤维样滑膜细胞的活化、增殖、迁移侵袭与凋亡，在动物实验中减轻了CIA小鼠的关节炎症状。在体外培养的破骨细胞中断藤益母汤下调RANKL信号通路和Wnt信号通路抑制破骨细胞增殖与活化。.该项目的研究初步阐明了断藤益母汤通过 “多成分-多靶点-多途径”起到治疗RA的分子机制，为其临床应用提供了科学依据，也为研发RA创新药物奠定基础，使用网络药理学分析与实验验证相结合的研究方法也为中药复方和中药单体研究提供了借鉴。.