Evaluating Method For The Effect Of Traditional Chinese Medicine On Synovial Angiogenesis Of Rheumatoid Arthritis

Rhuematoid arthritis (RA) is an autoimmune diseases characterized by chronic synovitis, cartilage erosion and bone joint destruction. The prevalence of rheumatoid arthritis is about0.5-1%in the world, and about0.32～0.36%in our country. Pathogenesis of RA has not yet been fully elucidated up to now. And, there is no effective therapy for RA.Angiogenesis accompanied by synovial hyperplasia and inflammation is the basis of pannus formation and joint destruction of RA. Clinical study has showed that the neovascularization in joints of RA is correlated with inflammation. The drugs with inhibiting angiogenesis can alleviate the syndrom of the RA. Therefore, blocking synovial angiogenesis has become one of the important target in the treatment of RA.RA is the disease of "rheumatism" of traditional Chinese medicine (TCM), but also collateral disease. Many clinical practice suggests a lot of Chinese herbal medicine have the effect of inhibiting synovial angiogenesis of RA. However, there is no evaluation method for the drugs of anti-RA synovial angiogenesis which is suitable for the multi-target, multi-level and multi-component of TCM wordly. The study is to establish the evaluation method for anti-RA synovial angiogenesis of TCM.In this study, evaluation methods of anti-RA synovial angiogenesis are built including animal models (collagen-induced arthritis model (CIA), animal models which can integrate disease and syndrome disease-syndromes, HUVEC and RA-HFLS), efficacy evaluation, mechanism, detection analysis. Then, triptolide,Qingluoyin,Wenluoyin, and Aconite Decoction are used to verify the scientific nature of the methods.1Screening models of anti-RA synovial angiogenesisCIA is the ideal model that study RA wordly. CIA is established in DA and SD rats respectively. The results show that the incidence of CIA in DA rats is100%, and in SD rats87.5%. CIA in DA and SD rats are suitable for study RA.In order to make animal models reflecting the characteristics of traditional Chinese medicine (TCM)"card", the RA models of the cold and hot syndrome are established by "wind, cold/hot and wet" stimulation based on CIA model.To investigate of the formation of blood vessels in the complex process, we established screening models in vivo and in vtro, Including:proliferation, migration, adhesion and lumen formed experiments of blood umbilical vein endothelial cells (HUVEC) and RA synovial fibroblasts (RA-HFLS). matrix glue bolt experiment in mice are used also.2Pharmacodynamic evaluation index of anti-RA synovial angiogenesisThe degree of swelling, incidence clinical score, the first of the disease time, the synovial inflammatory hyperplasia and the degree of the inflammatory cell infiltration as well as histopathological examination and X-ray are used to evaluate the effect of RA. The synovial tissue angiogenesis (HE dyeing and immunohistochemistry for CD31) in CIA rats and in mice.3Evaluation index of mechanism of Anti-RA synovial angiogenesis based on the angiogenesis processThe study investingates the HUVEC/RA-HFLS proliferation, migration, adhesion and lumen formation in vitro to examine HUVEC and RA HFLS functions and observes the regulatory pathway of VEGF/of VEGFR and Ang1/Tie2and inflammation-related regulatory factor expression in vitro and/or in vivo.4The practices of TCM with anti-RA synovial angiogenesisWith the methods above, we demonstrate for the first time that triptolide, an active compound of TWHF, has anti-RA synovial angiogenesis and its mechanism is inhibition of the proliferation, migration, adhesion of HUVEC and RA HFLS, lumen formation in mice and regulation of the VEGF/VEGFR and Ang/Tie2information pathway. The practice demonstrate the rationality and reliability of our methods.By CIA rats and Matrigel plug model in mice, the study shows that Qingluoyin and Wenluoyin have the effect of inhibition of angiogenesis, its mechanism may be related to inhibition the expression of IL-1β, TNFα and VEGF in CIA rats. The effect of Qingluoyin is better to Wenluoyin, but the target is similar. This suggest that CIA model may be suitbal to TCM with cold character. Aconite decoction is the classic prescription for cold dampness arthralgia syndrome from Zhongjing Zhang. With the RA models with the cold and hot syndrome and CIA, we investigating that Aconite decoction has the effect of inhibition of angiogenesis. And the effect of it on the cold model is the best, the effect on the CIA is second. Aconite decoction has no effect on hot model. This results that the cold model is suitbale for the TCM with hot character.The highlights:This method is based on improving the standard of disease-evaluating, and studying the function and mechanism of TCM against RA from a key pathology link of synovial angiogenesis, which has the advanced idea and higher international approval.The research mode of incorporating in vivo and in vitro, local and system, form and function, efficacy and mechanism coincide with the characteristics of TCM.The disease animal model is still the main model of RA efficacy evaluation of traditional Chinese medicine at present, and it is widely used in the efficacy evaluation and pharmacological mechanism research of TCM and the extract of anti-RA. The animals combined with disease and syndrome is is an ideal method of compound efficacy evaluation,which has typical of TCM syndrome and obvious drug properties of hot and cold. But it is limited by the maturity of syndrome animal models.In the practice, triptolide, formula (Qingluoyin and Wenluoyin) and classic formula of anti-RA cold syndrome (Aconite decoction) are combined, the designed is suitable and the results of validation is true.