玄胡索散成分分析及其治疗骨关节炎作用机制的网络药理学研究
发布时间：2019-05-29 21:36
【摘要】：骨关节炎是以进行性关节软骨损伤、软骨基质合成与降解失衡造成关节面完全被破坏为特征的慢性关节病。目前主要采用保守治疗、药物治疗和手术治疗方式,通过延缓疾病发展、预防关节损伤从而达到缓解病症的目的。患者多服用非甾体抗炎药、软骨保护剂等化药,但长期使用效果不佳,多有胃肠道反应等毒副作用。中医理论认为骨关节炎属"痹症",病因为气血痹阻、筋骨失养,治疗方法以补肾活血为主,辅以祛湿止痛。《普济方》中收载的玄胡索散,由延胡索、牛膝、当归、补骨脂组成,用于治疗肝肾不足,血凝气滞,腰腿疼痛等。虽然只有四味药,但组方精炼,体现了中医治疗骨关节炎的整体观念及遣方用药的配伍精髓。玄胡索散作为基础方在临床上应用广泛,并收载于以骨科闻名的中国中医科学院望京医院《骨科方剂药方管理系统》中,但目前尚无针对该复方的相关研究报道,其治疗骨关节炎的作用机制仍是空白,有必要进行深入研究,因此本试验对玄胡索散进行了以下研究:1.玄胡索散及其拆方对补骨脂素与异补骨脂素溶出量的影响采用高效液相色谱法分析玄胡索散及其拆方对补骨脂素与异补骨脂素溶出量的影响,结果发现各拆方组较全方组有显著差异(P0.05),其中当归、牛膝有助于二者的溶出,而延胡索抑制二者的溶出,为进一步揭示玄胡索散的配伍规律提供了依据。2.运用超高效液相色谱串联四极杆飞行时间质谱技术分析玄胡索散中所含化学成分基于超高效液相色谱串联四极杆飞行时间质谱技术,首次对玄胡索散中所含主要化学成分进行了快速分离与定性,利用优化后的分离与检测条件在13 min内对主要成分实现了有效分离,并通过与标准品、文献及数据库比较保留时间、碎片离子、裂解规律等信息,最终鉴定出57个化合物,为该药后期的药代动力学研究提供了依据。3.网络药理学预测玄胡索散治疗骨关节炎的作用机制基于网络药理学方法,在相关数据库及文献检索玄胡索散中四味中药所含化学成分、化学成分靶点、骨关节炎治疗靶点、蛋白相互作用关系等信息,并结合UPLC-Q-TOF-MS技术发现的配伍新成分,构建相互作用网络,进行网络药理学预测。最终经通路富集分析,发现了多条玄胡索散治疗骨关节炎的信号通路,如调节NO、IL-1β的生物合成,调节炎症反应、响应细胞因子或雌激素刺激,以及抗软骨细胞凋亡等。本研究在系统层面首次揭示了该药治疗骨关节炎的作用机制,并得到了相关化学成分靶点、治疗通路等之间的密切关系,为后期的试验研究提供了依据。4.玄胡索散对骨关节炎大鼠软骨基质的影响雄性SD大鼠随机分组,采用前交叉韧带横断联合部分半月板切除造骨关节炎模型,分别于给药3周、6周后处死大鼠取手术侧膝关节,进行HE染色切片评估软骨损伤程度与Masson染色切片评估胶原的降解。结果发现玄胡索散能明显抑制膝关节软骨损伤与胶原降解(P0.05),初步证明了玄胡索散治疗骨关节炎的潜在作用,为后期的体外细胞试验与网络药理学预测结果验证试验提供了依据。
[Abstract]:Osteoarthritis is a chronic arthropathy characterized by progressive joint cartilage injury, cartilage matrix synthesis and degradation imbalance resulting in the complete destruction of the articular surface. The invention mainly adopts the conservative treatment, the medicine treatment and the operation treatment mode, and the purpose of relieving the condition is achieved by delaying the development of the disease and preventing the joint injury. The patient has the advantages of no anti-inflammatory agent, chondroprotective agent and the like, but the long-term use effect is not good, and has the toxic and side effects such as gastrointestinal reaction and the like. The traditional Chinese medicine theory considers that the osteoarthritis belongs to the "a hand", and the disease is due to the obstruction of qi and blood, the loss of bones and bones, and the treatment method, which is mainly used for tonifying the kidney and promoting blood circulation, and is supplemented with the functions of eliminating And is prepared from Rhizoma Corydalis, Achyranthis Radix, Radix Angelicae Sinensis, and Fructus Psoraleae, and can be used for the treatment of deficiency of liver and kidney, blood coagulation and qi stagnation, and lumbago. Although only four medicines are used, the prescription is refined, and the integral concept of the traditional Chinese medicine for treating the osteoarthritis and the compatibility essence of the prescription drugs are reflected. It is widely used as a base in the clinical application, and is listed in the Wangjing Hospital of the Chinese Academy of Traditional Chinese Medicine, which is known as the Orthopaedics, and in the Prescription Management System of Orthopaedics. However, there is no relevant research report to the compound, and the mechanism for the treatment of osteoarthritis is still blank. It is necessary to carry out the in-depth study, so this test has carried out the following research on Xhosoxuan:1. The effect of Xuanhuoxue and its removal on the amount of psoralen and isopsoralen was analyzed by high performance liquid chromatography (HPLC). The results showed that there was a significant difference in the amount of psoralen and isopsoralen (P0.05). Achyranthis bidentata is helpful for the dissolution of the two, while the dissolution of the two can provide a basis for further revealing the compatibility of the Xuanhuoxue powder. The high performance liquid chromatography tandem quadrupole time-of-flight mass-spectrum technique is used to analyze the chemical components contained in the Xuanhuizu powder based on the ultra-high performance liquid chromatography tandem quadrupole time-of-flight mass spectrometry technology, and the first time the main chemical components contained in the Xhososol powder are rapidly separated and characterized, By using the optimized separation and detection conditions, the main components are effectively separated within 13 minutes, and the information such as retention time, fragment ion, cracking rule and the like is compared with the standard, the literature and the database, and the 57 compounds are finally identified, It provides a basis for the study of the pharmacokinetics of the drug in the later stage. The invention relates to a network pharmacology prediction function mechanism for treating osteoarthritis, which is based on the network pharmacology method, and the information such as the chemical components, the chemical components target point, the treatment target point of the osteoarthritis treatment, the protein interaction relation and the like contained in the four traditional Chinese medicines in the relevant database and the literature search, Combined with the new components of UPLC-Q-TOF-MS, the interaction network was constructed and the network pharmacological prediction was carried out. And finally, through the channel enrichment analysis, a plurality of signal paths for treating the osteoarthritis, such as regulating the biological synthesis of NO and IL-1, regulating the inflammatory reaction, responding to the cytokine or the estrogen stimulation, and the anti-chondrocyte apoptosis, and the like are found. In this study, the mechanism of the treatment of osteoarthritis was first revealed at the system level, and the close relationship between the relevant chemical components and the treatment path was obtained. The male SD rats were randomly divided into two groups: the anterior cruciate ligament transection and the joint part of the meniscectomy for osteoarthritis, and the knee joint of the operation side was sacrificed after 6 weeks of administration. The degree of cartilage damage was assessed by HE staining and the degradation of the collagen was assessed by the Masson staining section. The results showed that the treatment of the articular cartilage injury and the degradation of the collagen (P0.05). The potential role of the Xuanhuoxue powder in the treatment of osteoarthritis was preliminarily proved, and the basis was provided for the later in vitro cell test and the results of the validation of the network pharmacological prediction.
【学位授予单位】：北京中医药大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R285
[Abstract]:Osteoarthritis is a chronic arthropathy characterized by progressive joint cartilage injury, cartilage matrix synthesis and degradation imbalance resulting in the complete destruction of the articular surface. The invention mainly adopts the conservative treatment, the medicine treatment and the operation treatment mode, and the purpose of relieving the condition is achieved by delaying the development of the disease and preventing the joint injury. The patient has the advantages of no anti-inflammatory agent, chondroprotective agent and the like, but the long-term use effect is not good, and has the toxic and side effects such as gastrointestinal reaction and the like. The traditional Chinese medicine theory considers that the osteoarthritis belongs to the "a hand", and the disease is due to the obstruction of qi and blood, the loss of bones and bones, and the treatment method, which is mainly used for tonifying the kidney and promoting blood circulation, and is supplemented with the functions of eliminating And is prepared from Rhizoma Corydalis, Achyranthis Radix, Radix Angelicae Sinensis, and Fructus Psoraleae, and can be used for the treatment of deficiency of liver and kidney, blood coagulation and qi stagnation, and lumbago. Although only four medicines are used, the prescription is refined, and the integral concept of the traditional Chinese medicine for treating the osteoarthritis and the compatibility essence of the prescription drugs are reflected. It is widely used as a base in the clinical application, and is listed in the Wangjing Hospital of the Chinese Academy of Traditional Chinese Medicine, which is known as the Orthopaedics, and in the Prescription Management System of Orthopaedics. However, there is no relevant research report to the compound, and the mechanism for the treatment of osteoarthritis is still blank. It is necessary to carry out the in-depth study, so this test has carried out the following research on Xhosoxuan:1. The effect of Xuanhuoxue and its removal on the amount of psoralen and isopsoralen was analyzed by high performance liquid chromatography (HPLC). The results showed that there was a significant difference in the amount of psoralen and isopsoralen (P0.05). Achyranthis bidentata is helpful for the dissolution of the two, while the dissolution of the two can provide a basis for further revealing the compatibility of the Xuanhuoxue powder. The high performance liquid chromatography tandem quadrupole time-of-flight mass-spectrum technique is used to analyze the chemical components contained in the Xuanhuizu powder based on the ultra-high performance liquid chromatography tandem quadrupole time-of-flight mass spectrometry technology, and the first time the main chemical components contained in the Xhososol powder are rapidly separated and characterized, By using the optimized separation and detection conditions, the main components are effectively separated within 13 minutes, and the information such as retention time, fragment ion, cracking rule and the like is compared with the standard, the literature and the database, and the 57 compounds are finally identified, It provides a basis for the study of the pharmacokinetics of the drug in the later stage. The invention relates to a network pharmacology prediction function mechanism for treating osteoarthritis, which is based on the network pharmacology method, and the information such as the chemical components, the chemical components target point, the treatment target point of the osteoarthritis treatment, the protein interaction relation and the like contained in the four traditional Chinese medicines in the relevant database and the literature search, Combined with the new components of UPLC-Q-TOF-MS, the interaction network was constructed and the network pharmacological prediction was carried out. And finally, through the channel enrichment analysis, a plurality of signal paths for treating the osteoarthritis, such as regulating the biological synthesis of NO and IL-1, regulating the inflammatory reaction, responding to the cytokine or the estrogen stimulation, and the anti-chondrocyte apoptosis, and the like are found. In this study, the mechanism of the treatment of osteoarthritis was first revealed at the system level, and the close relationship between the relevant chemical components and the treatment path was obtained. The male SD rats were randomly divided into two groups: the anterior cruciate ligament transection and the joint part of the meniscectomy for osteoarthritis, and the knee joint of the operation side was sacrificed after 6 weeks of administration. The degree of cartilage damage was assessed by HE staining and the degradation of the collagen was assessed by the Masson staining section. The results showed that the treatment of the articular cartilage injury and the degradation of the collagen (P0.05). The potential role of the Xuanhuoxue powder in the treatment of osteoarthritis was preliminarily proved, and the basis was provided for the later in vitro cell test and the results of the validation of the network pharmacological prediction.
【学位授予单位】：北京中医药大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R285
【参考文献】
相关期刊论文 前10条
1 王美玲;张清清;付爽;刘月红;梁钏镭;陈娜;丁永胜;王海丽;瞿晓梅;赵保胜;高晓燕;;UPLC-Q-TOF MS~E技术结合UNIFI数据库筛查方法快速分析巴戟天化学成分[J];质谱学报;2017年01期
2 谷艳超;刘世清;夏韶强;廖琦;王文静;;骨关节炎发病机制和治疗的研究进展[J];中国骨与关节杂志;2016年10期
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