清热活血法治疗活动期类风湿关节炎临床与机制研究
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摘要
类风湿关节炎(rheumatoid arthritis, RA)是一类以关节滑膜慢性炎症为主要表现的自身免疫性疾病,是世界上主要致残性疾病之一。目前,世界的患病率为1%,我国的患病率为0.3-0.4%。本病可造成全身多关节肿胀疼痛,晚期可致关节畸形,功能丧失,患病10年的患者中至少50%不能坚持工作。
RA的首要治疗目标是通过控制炎症症状、阻止关节破坏、保持关节功能、维持正常社会活动,最大程度提高患者长期的健康、生活质量,延缓／阻断RA骨破坏是治疗的最终目标。RA疾病的活动状态是骨破坏主要预示因素,炎症过程不仅影响局部骨,也可导致远离部位的骨丢失,提示RA的骨破坏与滑膜炎有密切关系。我科室根据活动期RA的病机特点结合多年的临床经验,认为活动期RA的主要病机为湿热毒邪久恋,血凝痹阻经络,病邪深入骨骱,胶着不去,腐蚀筋骨气血,致关节肿痛僵硬,甚导致骨破坏,强调湿热瘀是活动期RA的重要致病机制及骨破坏的病理关键,清热活血法是活动期RA的基本治疗大法[8-9]。
本研究在RA清热活血法的指导下,以首都医学发展科研基金北京医学卫生科技联合攻关项目和国家自然科学基金资助课题为依托,以临床疗效确切的清热活血方药作为研究对象,采用免疫组化、ELISA、细胞共培养等技术,从理论研究、临床研究和实验研究三方面,验证了清热活血方药治疗活动期RA降低疾病活动度、改善关节功能及延缓骨破坏的临床疗效,并对其内在机制进行了探讨,取得了有学术价值的研究结果。
一、理论研究
1.通过整理中医对RA认识的历史沿革、相关病名、症状及病因病机的历代认识,梳理了病证结合治疗RA的诊疗思路。
2.通过对与“湿热瘀致痹”相关的历代文献及现代研究成果进行分析归纳,得出湿热瘀是活动期RA中医病机关键,并且是其骨破坏的病理关键,提出清热活血为活动期RA的治疗大法。
3.通过整理“骨”的生理病理在中医文献中的论述和RA骨破坏病因病机及治疗,并综述了中医药对RA骨破坏的临床及机制研究进展,提出RA骨破坏的临床及机制研究各种观点,丰富了中医治疗RA理论。
二、临床研究
目的评价清热活血方药降低湿热瘀阻证活动期RA疾病活动度及放射学进展评价。
方法分为中药组(清热活血方药组)、MTX组(甲氨蝶呤组)、中西药组(清热活血方药联合甲氨蝶呤组),进行了为期24周的随机对照研究和队列研究相结合的多中心临床研究,以评价RA疾病活动度(DAS28评分)的改善；并通过带药随访,采用Sharp/van der Heijde评分对1年期的双手X光片进行评价。
结果
1.本临床研究按计划共入组了213例受试者,中药组72例,中西药组71例,队列研究的MTX组70例,129例患者完成了24周的治疗。结果表明三组均可降低疾病活动度,中药组、中西药组、MTX组DAS28评分下降的均值分别为1.91、1.97、1.29,中药组和中西药组较甲氨蝶呤组疗效好。
2.中西药组治疗1个月后,关节功能较之疗前即开始出现明显改善,中药组和MTX组治疗3个月后,关节功能较之疗前开始出现明显改善；在改善关节功能方面,中西药组起效较快,中药组次之,MTX组最慢。中药组和中西药组对于改善RA关节功能方面较MTX组疗效好。
3.中药组、中西药组和MTX组在降低炎性实验室指标(ESR)方面均有较好疗效；三组比较中,中药+MTX的联合方案在降低ESR方面优于单独应用MTX方案。
4.随访进入放射学进展评价的合格病例共42例,中药组和中西药组各21例,两组Sharp总分、骨侵蚀评分及关节间隙狭窄评分均无统计学意义。两组Sharp评分变化率非常相似,中药组0.24±0.28/0.19(0.04,0.29),中西药组0.25±0.44/0.15(0.04,0.25)。两组不同进展程度的病例数也相似(p=0.46),无放射学进展者中药组7例,中西药组8例；放射学进展较明显者,中药组3例,中西药组1例。
5.安全性方面,中药组、中西药组和MTX组不良事件发生率有统计学差异,中西药组和MTX组出现的10例不良事件均与MTX密切相关,而纯中药组则未见明显不良反应。可见纯中药治疗较之中西药结合治疗、MTX治疗安全性更可靠。
三、实验研究
1.体内实验部分——清热活血方药抑制CIA大鼠Th/RANK通路以及骨破坏的实验研究
目的从Th/RANKL通路阐明清热活血方药治疗RA可能的“护骨”机制。
方法以牛Ⅱ型胶原诱发SD大鼠关节炎模型,以甲氨喋呤治疗为阳性对照药,清热活血方药水煎剂和醇提剂灌胃给药6周。从x光片、HE染色、骨密度检查多个层次观察CIA大鼠骨关节滑膜增生炎症及骨破坏变化,ELISA法检测CIA大鼠血清骨破坏因子RANKL、IL-17、MIP-1a的水平。
结果CIA大鼠血清RANKL、IL-17、MIP-1a的水平较正常组均显著升高,清热活血方药水煎剂和醇提剂治疗组大鼠血清IL-17、MIP-1a水平较模型组均显著下降,RANKL水平有下降趋势。甲氨喋呤治疗组大鼠血清RANKL、IL-17水平较模型组均显著下降,MIP-1a有下降趋势。
2.体外实验部分——清热活血方药对体外培养的RA滑膜细胞OPG、RANKL分泌及破骨细胞分化影响的实验研究
目的观察清热活血方药干预体外细胞共培养体系中OPG、RANKL分泌及破骨细胞分化的作用并探讨其分子机制。
方法建立体外人血PBMC和正常人滑膜成纤维细胞(RA-HFLS)共培养体系,从健康人外周血中分离获得PBMC,采用IL-1诱导诱导IL-17,并采用ELISA、Western法及TRAP活性检测等检测方法对清热活血方药对RANKL诱导的RAW264.7破骨细胞分化及OPG分泌IL-17水平的影响进行初步研究。
结果清热活血含药血清(5%、10%和20%)可以剂量依赖性的抑制体外培养的RA-HFLS增殖、提高RA-HFLS培养上清中OPG的水平、抑制RA-HFLS上清中RANKL的表达和IL-17的水平。
四、结论
1.确立了湿热瘀是活动期RA中医病机关键及其骨破坏的病理关键,清热活血为治疗大法。
2.临床研究证实：清热活血方药具有降低RA疾病活动度、改善关节功能,延缓RA骨破坏的“护骨”的作用。
3.实验研究证实：清热活血方药通过抑制Th细胞因子(IL-17),使RANKL表达下调,OPG表达上调,从而调整了OPG/RANKL/RANK通路,延缓了骨破坏,证实了清热活血方药“护骨”的内在机制。
五、特色与创新之处
1.理论创新：首次提出湿热瘀血致活动期RA炎症病变、骨质破坏,清热活血方药可以“护骨”的科学假说,丰富和完善中医药治疗RA方证对应、病证结合治疗理论,阐明临床中药优效的科学基础。
2.思路创新：
2.1从干预RA关节骨破坏入手,采用Sharp评分,评价清热活血方药对活动期RA患者骨破坏的影响,为清热活血方药对RA疾病终点结局的影响提供循证医学依据。
2.2从新兴的“骨免疫学”角度,通过研究Th细胞因子(IL-17)、OPG及RANKL等靶点探讨清热活血方药对关节骨破坏的全身和局部作用机理,证实了清热活血方药可以“护骨”的科学假说。
总之,本研究从临床到基础,深层次的研究了清热活血方药对活动期RA疾病疗效和骨破坏的影响,揭示了中医清热活血法治疗活动期RA的科学内涵,为中医理论更好地指导临床实践提供了科学依据,为中药治疗RA的“护骨”机制探索出新的方法与思路。
Rheumatoid arthritis (RA) is a chronic, systemic inflammatory disorder that may affect many tissues and organs, but principally attacks synovial joints. The pathology of the disease process often leads to the destruction of articular cartilage and bone, finally the affected joints are ankylosis. About1%of the world's population is afflicted by RA. Onset is most frequent between the ages of40and50, but people of any age can be affected.70%patients with RA have joint erosion.90%patients with RA have joint destruction just2years after the onset of the symptoms.
The primary goal of RA treatment is through control inflammation symptoms, prevent joint destruction, keep joint function, maintain normal social activities and maximize the long-term health of the patients, quality of life, and delay/block RA bone destruction is the ultimate goal of treatment. RA disease activity is major factor of bone destroy, the inflammatory process affect not only local bone, can also lead to far away from the parts of the bone loss, hints that bone destruction and RA synovial membrane inflammation has close relationship. Our department according to active period of pathogenesis of RA characteristics combined with years of the clinical experience, emphasis on heat and dampness stasis is the important active RA pathogenic mechanism and pathological key of bone damage, Qingrehuoxue therapy is basic treatment method of active period RA.
This project under the guidance of Qingrehuoxue therapy, scientific development in the capital fund Beijing medical medical health technology project and the national natural science funds subject as the basis, used the immunohistochemical, ELISA, cell co-culture technologies, from the theory research, clinical research and the experimental research three aspects to explore effectiveness of Qingrehuoxue decoction treatment RA to reduce desease activity and delay bone destruction, and the internal mechanism was also be discussed.
Therapeutic Study
1. By sorting out the Traditional Chinese Medicine historical evolution, know RA related problems and symptoms and etiology, the history of the understanding, combed the disease and syndrome with combined therapy of diagnosis and treatment of RA ideas.
2. To analyze the "heat, dampness and blood stasis to bi" relevant literature and modern research results, and summarized heat, dampness and blood stasis are the key TCM pathogenesis of active RA, and is the key pathogenesis of bone destruction, put forward that Qingrehuoxue therapy is active RA treatment method.
3. Through sorting "bone" the physiology, pathology and treatment in the Traditional Chinese Medicine literature, discusses and RA bone destruction etiology and pathogenesis, the paper summarized the destruction of RA bone clinical and mechanism research progress, and puts forward the clinical and bone is destroyed RA mechanism study of the various ideas, enrich the theory of traditional Chinese medicine treatment RA.
Clinical Study
Objective:to evaluate Qingrehuoxue decoction reduces RA disease activity and radiological mobility progress evaluation.
Methods:Enrolled active RA patients were randomized into in Qingrehuoxue(QRHX) group and Methotrexate(MTX) group and QRHX+MTX group. A24weeks of randomized, controlled study and cohort study to assess RA disease activity (DAS28score) improvement; and by taking medicine follow-up, used the Sharp/van der Heijde method to evaluate one-year hands X-ray score.
Results:
1. We enrolled in this study213patients which were entered in Full Analysis Set (FAS),129patients completed24weeks of treatment. The results show that three groups all can reduce disease activity and the mean decline of DAS28score of QRHX group, MTX group and QRHX+MTX group were1.91,1.97and1.29, QRHX group and QRHX+MTX group have better curative effect than MTX group.
2. QRHX+MTX group can improve joint function just after one month treating, while QRHX group and MTX group improve joint function after three months treating; In improving joint function, QRHX+MTX group works faster, QRHX group is the second, and MTX group is the third. QRHX group and QRHX+MTX group have a better effect than MTX group to improving the RA joint function.
3. Three groups have good effect to reduce the inflammatory laboratory index (ESR); QRHX+MTX group works better than MTX group.
4. Follow-up into radiology qualified cases progress evaluation of42cases, QRHX group and QRHX+MTX group of each21cases, two groups of Sharp total score, bone erosion score and the joint space narrow scores are no statistical significance. Two groups of Sharp scoring rate very similar, QRHX group0.24±0.28/0.19(0.04,0.29), QRHX+MTX group0.25±0.44/0.15(0.04,0.25). Two different extent the number of cases also similar (p=0.46), no radiation study progress of QRHX group in7cases,8cases of QRHX+MTX medicine; Radiology progress the obvious, QRHX group in3, the other in1.5. Safety, QRHX group adverse reaction rate (0%) lower than QRHX+MTX group (1.55%) and MTX group (11.4%). The safety of pure Chinese herbal medicine treatment compared with MTX or combined with MTX treatments are more reliable.
Experimental Study
1. In vivo experimental study:QRHX decoction inhibit the CIA rat Th/RANK pathways and bone destruction.
Objects:Based on "osteoimmunology", through Th/RANKL pathways expounds QRHX decoction treatment RA possible "protect bone" mechanism.
Methods:to cattle II collagen type SD rats induced arthritis models to MTX for the positive control medicine, QRHX water decoction and ethanol-extraction agent to fill the stomach six weeks. From an X-ray, HE dyeing, bone mineral density check more levels observed CIA rats inflammation and bone joint synovial hyperplasia damage change, ELISA test levels of CIA rats bone damage factor RANKL, IL-17, the MIP-1.
Results:Serum levels of CIA rats RANKL, IL-17, the MIP-1of the normal group were significantly increased, QRHX water decoction and ethanol-extraction agent groups, serum levels of IL-17, the MIP-1were decreased significantly than model group, the RANKL level had decline trend. MTX group, serum levels RANKL, IL-17are decreased significantly than model group, the MIP-1had decline trend.2. In vitro experimental study:influence of QRHX decoction to RA synovial cell co-culture OPG, RANKL secretion, and broken bone cell differentiation
Objective:to observe influence of QRHX decoction to RA synovial cell co-culture OPG, RANKL secretion, and broken bone cell differentiation role and discuss the molecular mechanism.
Methods:to establish the in vitro human blood PBMC and normal synovial fibroblasts (RA-HFLS) of co-culture system, from healthy people in the peripheral blood separation get PBMC, using IL-1induction induction IL-17, and uses the ELISA, Western law and the TRAP active detection methods of QRHX decoction to observe the influence of the RAW264.7induction formulas RANKL broken bone cell differentiation and levels of OPG secretion, IL-17.
Results:blood serum containing QRHX (5%,10%and20%) can dose dependent in vitro inhibition of the RA-HFLS proliferation, improve RA-HFLS cultivation in the level of OPG, and inhibit of RA-HFLS RANKL in the expression and reduce IL-17level. Conclusion
1. Established the heat, dampness and blood stasis are the key pathogenesis and pathological of active RA bone destruction, Qingrehuoxue therapy is the basic principle.
2. Clinical study confirms:Qingrehuoxue decoction can reduce RA disease activity, and delay RA bone destruction, has a "protect bone" role.
3. Experimental study confirms:Qingrehuoxue decoction by inhibiting IL-17, cut down the RANKL expression, up the OPG level and adjusted the OPG/RANKL/RANK pathways, delay the bone destruction, confirm Qingrehuoxue decoction "protect bone" intrinsic mechanism. Characteristics and innovative points
1. The theory innovation:
first put forward heat, dampness and blood stasis are key points that lead to active RA inflammation lesion, bone destruction. Qingrehuoxue therapy can "protect bone" scientific hypothesis, enrich and perfect the Traditional Chinese Medicine for RA correspondence of prescription and syndrome, and correspondence of disease and syndrome therapy theory, clarify clinical Chinese medicine superior effect scientific basis.
2. The idea innovation:
2.1From intervention RA bone destruction, study the influence of Qingrehuoxue decoction treatment of RA patients bone destructive effects, to provide the influence of evidence-based medicine basis.
2.2Form" osteoimmunology " pespect, through the research on Th cell factors (IL-17), OPG and RANKL targets such as discussed Qingrehuoxue decoction invigorate the bone destruction systemic and local function mechanism, confirmed Qingrehuoxue decoction has "protect bone" scientific hypothesis.
All in all, this project from clinical to the base, deeply researches on the Qingrehuoxue decoction treatment of active RA formulas blood disease curative effect and the influence of the RA bone destruction, and reveals the scientific connotation of Qingrehuoxue therapy treating active RA, guides clinical practice for Chinese medicine theory, and provides new methods and ideas of the scientific basis for Chinese medicine treatment of RA "protect bone" mechanism exploration.
RA的首要治疗目标是通过控制炎症症状、阻止关节破坏、保持关节功能、维持正常社会活动,最大程度提高患者长期的健康、生活质量,延缓／阻断RA骨破坏是治疗的最终目标。RA疾病的活动状态是骨破坏主要预示因素,炎症过程不仅影响局部骨,也可导致远离部位的骨丢失,提示RA的骨破坏与滑膜炎有密切关系。我科室根据活动期RA的病机特点结合多年的临床经验,认为活动期RA的主要病机为湿热毒邪久恋,血凝痹阻经络,病邪深入骨骱,胶着不去,腐蚀筋骨气血,致关节肿痛僵硬,甚导致骨破坏,强调湿热瘀是活动期RA的重要致病机制及骨破坏的病理关键,清热活血法是活动期RA的基本治疗大法[8-9]。
本研究在RA清热活血法的指导下,以首都医学发展科研基金北京医学卫生科技联合攻关项目和国家自然科学基金资助课题为依托,以临床疗效确切的清热活血方药作为研究对象,采用免疫组化、ELISA、细胞共培养等技术,从理论研究、临床研究和实验研究三方面,验证了清热活血方药治疗活动期RA降低疾病活动度、改善关节功能及延缓骨破坏的临床疗效,并对其内在机制进行了探讨,取得了有学术价值的研究结果。
一、理论研究
1.通过整理中医对RA认识的历史沿革、相关病名、症状及病因病机的历代认识,梳理了病证结合治疗RA的诊疗思路。
2.通过对与“湿热瘀致痹”相关的历代文献及现代研究成果进行分析归纳,得出湿热瘀是活动期RA中医病机关键,并且是其骨破坏的病理关键,提出清热活血为活动期RA的治疗大法。
3.通过整理“骨”的生理病理在中医文献中的论述和RA骨破坏病因病机及治疗,并综述了中医药对RA骨破坏的临床及机制研究进展,提出RA骨破坏的临床及机制研究各种观点,丰富了中医治疗RA理论。
二、临床研究
目的评价清热活血方药降低湿热瘀阻证活动期RA疾病活动度及放射学进展评价。
方法分为中药组(清热活血方药组)、MTX组(甲氨蝶呤组)、中西药组(清热活血方药联合甲氨蝶呤组),进行了为期24周的随机对照研究和队列研究相结合的多中心临床研究,以评价RA疾病活动度(DAS28评分)的改善；并通过带药随访,采用Sharp/van der Heijde评分对1年期的双手X光片进行评价。
结果
1.本临床研究按计划共入组了213例受试者,中药组72例,中西药组71例,队列研究的MTX组70例,129例患者完成了24周的治疗。结果表明三组均可降低疾病活动度,中药组、中西药组、MTX组DAS28评分下降的均值分别为1.91、1.97、1.29,中药组和中西药组较甲氨蝶呤组疗效好。
2.中西药组治疗1个月后,关节功能较之疗前即开始出现明显改善,中药组和MTX组治疗3个月后,关节功能较之疗前开始出现明显改善；在改善关节功能方面,中西药组起效较快,中药组次之,MTX组最慢。中药组和中西药组对于改善RA关节功能方面较MTX组疗效好。
3.中药组、中西药组和MTX组在降低炎性实验室指标(ESR)方面均有较好疗效；三组比较中,中药+MTX的联合方案在降低ESR方面优于单独应用MTX方案。
4.随访进入放射学进展评价的合格病例共42例,中药组和中西药组各21例,两组Sharp总分、骨侵蚀评分及关节间隙狭窄评分均无统计学意义。两组Sharp评分变化率非常相似,中药组0.24±0.28/0.19(0.04,0.29),中西药组0.25±0.44/0.15(0.04,0.25)。两组不同进展程度的病例数也相似(p=0.46),无放射学进展者中药组7例,中西药组8例；放射学进展较明显者,中药组3例,中西药组1例。
5.安全性方面,中药组、中西药组和MTX组不良事件发生率有统计学差异,中西药组和MTX组出现的10例不良事件均与MTX密切相关,而纯中药组则未见明显不良反应。可见纯中药治疗较之中西药结合治疗、MTX治疗安全性更可靠。
三、实验研究
1.体内实验部分——清热活血方药抑制CIA大鼠Th/RANK通路以及骨破坏的实验研究
目的从Th/RANKL通路阐明清热活血方药治疗RA可能的“护骨”机制。
方法以牛Ⅱ型胶原诱发SD大鼠关节炎模型,以甲氨喋呤治疗为阳性对照药,清热活血方药水煎剂和醇提剂灌胃给药6周。从x光片、HE染色、骨密度检查多个层次观察CIA大鼠骨关节滑膜增生炎症及骨破坏变化,ELISA法检测CIA大鼠血清骨破坏因子RANKL、IL-17、MIP-1a的水平。
结果CIA大鼠血清RANKL、IL-17、MIP-1a的水平较正常组均显著升高,清热活血方药水煎剂和醇提剂治疗组大鼠血清IL-17、MIP-1a水平较模型组均显著下降,RANKL水平有下降趋势。甲氨喋呤治疗组大鼠血清RANKL、IL-17水平较模型组均显著下降,MIP-1a有下降趋势。
2.体外实验部分——清热活血方药对体外培养的RA滑膜细胞OPG、RANKL分泌及破骨细胞分化影响的实验研究
目的观察清热活血方药干预体外细胞共培养体系中OPG、RANKL分泌及破骨细胞分化的作用并探讨其分子机制。
方法建立体外人血PBMC和正常人滑膜成纤维细胞(RA-HFLS)共培养体系,从健康人外周血中分离获得PBMC,采用IL-1诱导诱导IL-17,并采用ELISA、Western法及TRAP活性检测等检测方法对清热活血方药对RANKL诱导的RAW264.7破骨细胞分化及OPG分泌IL-17水平的影响进行初步研究。
结果清热活血含药血清(5%、10%和20%)可以剂量依赖性的抑制体外培养的RA-HFLS增殖、提高RA-HFLS培养上清中OPG的水平、抑制RA-HFLS上清中RANKL的表达和IL-17的水平。
四、结论
1.确立了湿热瘀是活动期RA中医病机关键及其骨破坏的病理关键,清热活血为治疗大法。
2.临床研究证实：清热活血方药具有降低RA疾病活动度、改善关节功能,延缓RA骨破坏的“护骨”的作用。
3.实验研究证实：清热活血方药通过抑制Th细胞因子(IL-17),使RANKL表达下调,OPG表达上调,从而调整了OPG/RANKL/RANK通路,延缓了骨破坏,证实了清热活血方药“护骨”的内在机制。
五、特色与创新之处
1.理论创新：首次提出湿热瘀血致活动期RA炎症病变、骨质破坏,清热活血方药可以“护骨”的科学假说,丰富和完善中医药治疗RA方证对应、病证结合治疗理论,阐明临床中药优效的科学基础。
2.思路创新：
2.1从干预RA关节骨破坏入手,采用Sharp评分,评价清热活血方药对活动期RA患者骨破坏的影响,为清热活血方药对RA疾病终点结局的影响提供循证医学依据。
2.2从新兴的“骨免疫学”角度,通过研究Th细胞因子(IL-17)、OPG及RANKL等靶点探讨清热活血方药对关节骨破坏的全身和局部作用机理,证实了清热活血方药可以“护骨”的科学假说。
总之,本研究从临床到基础,深层次的研究了清热活血方药对活动期RA疾病疗效和骨破坏的影响,揭示了中医清热活血法治疗活动期RA的科学内涵,为中医理论更好地指导临床实践提供了科学依据,为中药治疗RA的“护骨”机制探索出新的方法与思路。
Rheumatoid arthritis (RA) is a chronic, systemic inflammatory disorder that may affect many tissues and organs, but principally attacks synovial joints. The pathology of the disease process often leads to the destruction of articular cartilage and bone, finally the affected joints are ankylosis. About1%of the world's population is afflicted by RA. Onset is most frequent between the ages of40and50, but people of any age can be affected.70%patients with RA have joint erosion.90%patients with RA have joint destruction just2years after the onset of the symptoms.
The primary goal of RA treatment is through control inflammation symptoms, prevent joint destruction, keep joint function, maintain normal social activities and maximize the long-term health of the patients, quality of life, and delay/block RA bone destruction is the ultimate goal of treatment. RA disease activity is major factor of bone destroy, the inflammatory process affect not only local bone, can also lead to far away from the parts of the bone loss, hints that bone destruction and RA synovial membrane inflammation has close relationship. Our department according to active period of pathogenesis of RA characteristics combined with years of the clinical experience, emphasis on heat and dampness stasis is the important active RA pathogenic mechanism and pathological key of bone damage, Qingrehuoxue therapy is basic treatment method of active period RA.
This project under the guidance of Qingrehuoxue therapy, scientific development in the capital fund Beijing medical medical health technology project and the national natural science funds subject as the basis, used the immunohistochemical, ELISA, cell co-culture technologies, from the theory research, clinical research and the experimental research three aspects to explore effectiveness of Qingrehuoxue decoction treatment RA to reduce desease activity and delay bone destruction, and the internal mechanism was also be discussed.
Therapeutic Study
1. By sorting out the Traditional Chinese Medicine historical evolution, know RA related problems and symptoms and etiology, the history of the understanding, combed the disease and syndrome with combined therapy of diagnosis and treatment of RA ideas.
2. To analyze the "heat, dampness and blood stasis to bi" relevant literature and modern research results, and summarized heat, dampness and blood stasis are the key TCM pathogenesis of active RA, and is the key pathogenesis of bone destruction, put forward that Qingrehuoxue therapy is active RA treatment method.
3. Through sorting "bone" the physiology, pathology and treatment in the Traditional Chinese Medicine literature, discusses and RA bone destruction etiology and pathogenesis, the paper summarized the destruction of RA bone clinical and mechanism research progress, and puts forward the clinical and bone is destroyed RA mechanism study of the various ideas, enrich the theory of traditional Chinese medicine treatment RA.
Clinical Study
Objective:to evaluate Qingrehuoxue decoction reduces RA disease activity and radiological mobility progress evaluation.
Methods:Enrolled active RA patients were randomized into in Qingrehuoxue(QRHX) group and Methotrexate(MTX) group and QRHX+MTX group. A24weeks of randomized, controlled study and cohort study to assess RA disease activity (DAS28score) improvement; and by taking medicine follow-up, used the Sharp/van der Heijde method to evaluate one-year hands X-ray score.
Results:
1. We enrolled in this study213patients which were entered in Full Analysis Set (FAS),129patients completed24weeks of treatment. The results show that three groups all can reduce disease activity and the mean decline of DAS28score of QRHX group, MTX group and QRHX+MTX group were1.91,1.97and1.29, QRHX group and QRHX+MTX group have better curative effect than MTX group.
2. QRHX+MTX group can improve joint function just after one month treating, while QRHX group and MTX group improve joint function after three months treating; In improving joint function, QRHX+MTX group works faster, QRHX group is the second, and MTX group is the third. QRHX group and QRHX+MTX group have a better effect than MTX group to improving the RA joint function.
3. Three groups have good effect to reduce the inflammatory laboratory index (ESR); QRHX+MTX group works better than MTX group.
4. Follow-up into radiology qualified cases progress evaluation of42cases, QRHX group and QRHX+MTX group of each21cases, two groups of Sharp total score, bone erosion score and the joint space narrow scores are no statistical significance. Two groups of Sharp scoring rate very similar, QRHX group0.24±0.28/0.19(0.04,0.29), QRHX+MTX group0.25±0.44/0.15(0.04,0.25). Two different extent the number of cases also similar (p=0.46), no radiation study progress of QRHX group in7cases,8cases of QRHX+MTX medicine; Radiology progress the obvious, QRHX group in3, the other in1.5. Safety, QRHX group adverse reaction rate (0%) lower than QRHX+MTX group (1.55%) and MTX group (11.4%). The safety of pure Chinese herbal medicine treatment compared with MTX or combined with MTX treatments are more reliable.
Experimental Study
1. In vivo experimental study:QRHX decoction inhibit the CIA rat Th/RANK pathways and bone destruction.
Objects:Based on "osteoimmunology", through Th/RANKL pathways expounds QRHX decoction treatment RA possible "protect bone" mechanism.
Methods:to cattle II collagen type SD rats induced arthritis models to MTX for the positive control medicine, QRHX water decoction and ethanol-extraction agent to fill the stomach six weeks. From an X-ray, HE dyeing, bone mineral density check more levels observed CIA rats inflammation and bone joint synovial hyperplasia damage change, ELISA test levels of CIA rats bone damage factor RANKL, IL-17, the MIP-1.
Results:Serum levels of CIA rats RANKL, IL-17, the MIP-1of the normal group were significantly increased, QRHX water decoction and ethanol-extraction agent groups, serum levels of IL-17, the MIP-1were decreased significantly than model group, the RANKL level had decline trend. MTX group, serum levels RANKL, IL-17are decreased significantly than model group, the MIP-1had decline trend.2. In vitro experimental study:influence of QRHX decoction to RA synovial cell co-culture OPG, RANKL secretion, and broken bone cell differentiation
Objective:to observe influence of QRHX decoction to RA synovial cell co-culture OPG, RANKL secretion, and broken bone cell differentiation role and discuss the molecular mechanism.
Methods:to establish the in vitro human blood PBMC and normal synovial fibroblasts (RA-HFLS) of co-culture system, from healthy people in the peripheral blood separation get PBMC, using IL-1induction induction IL-17, and uses the ELISA, Western law and the TRAP active detection methods of QRHX decoction to observe the influence of the RAW264.7induction formulas RANKL broken bone cell differentiation and levels of OPG secretion, IL-17.
Results:blood serum containing QRHX (5%,10%and20%) can dose dependent in vitro inhibition of the RA-HFLS proliferation, improve RA-HFLS cultivation in the level of OPG, and inhibit of RA-HFLS RANKL in the expression and reduce IL-17level. Conclusion
1. Established the heat, dampness and blood stasis are the key pathogenesis and pathological of active RA bone destruction, Qingrehuoxue therapy is the basic principle.
2. Clinical study confirms:Qingrehuoxue decoction can reduce RA disease activity, and delay RA bone destruction, has a "protect bone" role.
3. Experimental study confirms:Qingrehuoxue decoction by inhibiting IL-17, cut down the RANKL expression, up the OPG level and adjusted the OPG/RANKL/RANK pathways, delay the bone destruction, confirm Qingrehuoxue decoction "protect bone" intrinsic mechanism. Characteristics and innovative points
1. The theory innovation:
first put forward heat, dampness and blood stasis are key points that lead to active RA inflammation lesion, bone destruction. Qingrehuoxue therapy can "protect bone" scientific hypothesis, enrich and perfect the Traditional Chinese Medicine for RA correspondence of prescription and syndrome, and correspondence of disease and syndrome therapy theory, clarify clinical Chinese medicine superior effect scientific basis.
2. The idea innovation:
2.1From intervention RA bone destruction, study the influence of Qingrehuoxue decoction treatment of RA patients bone destructive effects, to provide the influence of evidence-based medicine basis.
2.2Form" osteoimmunology " pespect, through the research on Th cell factors (IL-17), OPG and RANKL targets such as discussed Qingrehuoxue decoction invigorate the bone destruction systemic and local function mechanism, confirmed Qingrehuoxue decoction has "protect bone" scientific hypothesis.
All in all, this project from clinical to the base, deeply researches on the Qingrehuoxue decoction treatment of active RA formulas blood disease curative effect and the influence of the RA bone destruction, and reveals the scientific connotation of Qingrehuoxue therapy treating active RA, guides clinical practice for Chinese medicine theory, and provides new methods and ideas of the scientific basis for Chinese medicine treatment of RA "protect bone" mechanism exploration.
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