蒙药忠伦阿汤对CIA模型大鼠免疫调节及诱导滑膜细胞凋亡作用机制的研究

Rheumatoid arthritis ( RA ) is systematic polyarthritis，the imbalance of immune regulation is the main pathogenesis, the basic pathological characteristics is Rheumatoid Synovium. It is recognized one of the refractory diseases. At present there are no radical drugs and methods to RA. Finding a new safe and effective drug from the Traditional Chinese Medicines and Mongolian Medicines has been the focus of the scholars. Mongolian medicine Zhong lun a Tang has the function of removing pathogenic heat from blood and with effectiveness of dredging the channels and alleviating pain. It used to treat the common Mongolian medicine disease such as huangshui disease, wandering pain Syndrome etc. There are reports that clinical application Zhong lun a decoction to RA treatment has significant effect. Our project group will further extend on the basis of previous studies. Through the establishment of the rats model of collagen-induced arthritis (CIA) and tripterygium group is control one, using modern molecular biology methods, such as flow cytometry , enzyme linked immunoassay, western blot(WB), with in vivo experiment, observe the effect of drugs on the immune organs ( thymus, spleen, lymph nodes ), the serum T lymphocyte subsets ( Th1, Th2 ), joint tissues, related cytokines and we will investigate the immunomodulatory effect; With in vitro experiments, using Cell culture technique and so on, observe the effect of drugs on induced apoptosis of synoviocytes in CIA rats. From different aspects, angles, a comprehensive system to reveal the key target of of Zhong lun a Tang to RA, provide an objective basis for clinical application, add the new contents of the Mongolian medicine for the treatment of RA, promote the modernization of Mongolian medicine.

类风湿关节炎(RA)为系统性多关节炎，免疫调节失衡是主要发病机制，关节滑膜炎为基本病理特征，是公认的难治性疾病之一，目前尚无根治性药物与方法，从中蒙医药中寻找安全有效的新药物一直是学者们关注的热点。忠伦阿汤可清热活血、舒筋止痛，治疗蒙医热性黄水病、游痛证（相当于RA）疗效确切。本项目组在前期研究基础上进一步深入扩展，拟建立胶原诱导性关节炎（CIA）大鼠模型,以雷公藤作对照药，利用现代分子生物学方法，如流式细胞技术、酶联免疫法、蛋白免疫印迹法等，通过体内实验，观察药物对免疫器官（胸腺、脾、淋巴结）、血清中T细胞亚群（Th1、Th2）、关节组织、相关细胞因子的影响，探讨其免疫调节作用；通过体外实验，采用细胞培养技术等，观察药物对诱导CIA大鼠滑膜细胞凋亡的影响，从不同层面、角度，全面系统揭示忠伦阿汤干预RA的关键靶标，为临床应用提供客观依据，为蒙医药治疗RA增添新内容，促进蒙医药现现代化进程。

蒙药忠伦阿汤有清热凉血、舒筋止痛功能，治疗蒙医常见热性黄水病、游痛证等，临床报道治疗类风湿关节炎（RA）等疗效显著。本课题研究忠伦阿汤对CIA关节炎大鼠模型免疫调节及诱导滑膜细胞凋亡的影响，以雷公藤作为对照，观察药物对大鼠免疫器官的影响，结果显示忠伦阿汤可下调胸腺指数、脾脏指数及小肠派氏小结计数； 观察药物对CIA大鼠组织病理学变化的影响，结果显示忠伦阿汤可下调大鼠滑膜增生、细胞浸润及骨破坏积分；观察忠伦阿汤对CIA大鼠血清和踝关节中TGF-β，IFN-γ，OPG，RANKL，MMP-3，MMP-9的影响，结果显示忠伦阿汤可下调促炎因子IFN－γ、抗CII抗体、RANKL、MMP-3和MMP-9的表达水平，上调抑炎因子TGF-β和OPG的表达；观察忠伦阿汤对CIA大鼠爪关节中IFN-γ、MMP-3、MMP-9的蛋白表达水平，结果显示忠伦阿汤可下调IFN-γ、MMP-3、MMP-9的蛋白表达水平；观察忠伦阿汤及苦参碱对CIA大鼠Bax、Bcl-2、caspase3、cyclin D1、P-JAK2、P-STAT1、P-STAT3的影响，结果显示忠伦阿汤可上调Bax、caspase3的表达，下调Bcl-2、cyclin D1、P-JAK2、P-STAT1、P-STAT3的表达；观察忠伦阿汤含药血清对CIA大鼠FLS细胞周期和凋亡的影响，结果显示忠伦阿汤可抑制FLS增殖速度并增加FLS凋亡率；观察苦参碱对FLS增殖与凋亡的影响，结果显示苦参碱可抑制FLS增殖速度并增加FLS凋亡率。 说明忠伦阿汤能够有效抑制免疫失调、滑膜炎及骨破坏，并能抑制滑膜细胞的增殖，诱导其凋亡，从而干预RA的发展，而其组分苦参的主要有效成分苦参碱有显著的抑制滑膜细胞增殖并诱导其凋亡的作用，值得进一步研究。本研究为临床蒙医药应用提供了科学依据。