艾拉莫德对类风湿关节炎成纤维样滑膜细胞增殖、迁移的影响及临床疗效观察
发布时间：2018-07-28 07:26
【摘要】：研究背景类风湿关节炎(RA)是一类以长病程、侵袭性破坏关节为特征的自身免疫性疾病,以多关节肿痛,滑膜炎为主要特点,未经正规治疗可导致关节残疾及功能障碍,可累及到各个脏器受累。其中,滑膜组织最容易受到累及,成纤维细胞样滑膜细胞(FLS)具有“类肿瘤”的特征,通过分泌炎性介质、增殖、侵袭及迁移等行为影响了 RA的病程,在关节稳态和RA发病机制中起重要作用,FLS的增殖、迁移行为受多种细胞因子及信号通路影响,而一种新型的抗类风湿药艾拉莫德(T-614)能作用于某些共同的细胞因子及信号通路。因此本文通过实验研究了解艾拉莫德是否能影响FLS的增殖及迁移行为,并通过临床观察艾拉莫德的疗效。研究目的1.体外原代培养RA患者滑膜细胞,了解艾拉莫德对FLS增殖及迁移功能的影响。2.观察了解不同艾拉莫德用药方法,并与传统DMARDs联合药物进行比较,了解对RA患者的临床血清学指标影响及病情改善情况。研究方法组织块法培养RA患者滑膜细胞,无菌条件下,通过关节镜手术选取RA患者滑膜组织,培养皿中并倒置放入培养箱内,观察不同浓度下艾拉莫德(100、200、400、800 ng/ml)作用滑膜细胞72小时后的细胞毒作用,是否引起部分细胞凋亡,以及观察滑膜细胞是否有迁移行为,完成细胞凋亡实验确定引起细胞凋亡的有效浓度并避免对随后实验的干扰,并通过Edu细胞增殖实验、细胞迁移Transwell实验了解艾拉莫德对RAFLS增殖和迁移的影响,并探讨可能的机制。临床实验筛选确诊为类风湿关节炎并且病情活动的患者90人,其中男性19人,女性71人,年龄21～67岁,平均年龄47.23±15.62岁,平均病程4.75±3.53年),患者随机分为3组,联合治疗组(31例)予艾拉莫德25mg/bid+MTX 10mg/qw,艾拉莫德单药组(28例)艾拉莫德25mg/bid,对照组(31例)予甲氨蝶呤(MTX)10mg/qw+来氟米特(LEF)20mg/qd。分别在用药0,1,3,6,12月观察抗CCP抗体、RF、ESR、CRP、PLT、病人VAS评分、医生VAS评分、DAS28-ESR、HAQ评分等指标综合改变情况。观察过程中因患者个人原因或药物不良反应退出观察研究8例,最后同时符合纳入标准、排除标准并完成整个临床治疗观察过程的患者共82例。本研究中所有定量资料在进行统计学分析之前均进行正态性检验,正态分布数据采用均数±标准差表示,非正态分布采用中位数M表示。用统计学方法分析治疗前及治疗中各时间点指标情况,是否具统计学意义,并比较每个指标在不同治疗组随时间变化情况,行组间比较,是否具有统计学意义。研究结果1.艾拉莫德(浓度达到800ng/ml时)能促进RAFLS凋亡,当浓度≤400ng/ml时对凋亡无明显影响;艾拉莫德(浓度达到400ng/ml时)能抑制滑膜RAFLS增殖。艾拉莫德(浓度达到200ng/ml时)能抑制滑膜RAFLS迁移,该抑制作用呈浓度依赖性。2.艾拉莫德能降低抗CCP抗体和RF水平,艾拉莫德联合MTX在降抗体方面疗效较传统DMARDs联合组(MTX+LEF)更有优势。3.传统DMARDs联合组(MTX+LEF)在血沉、CRP等指标的远期疗效方面较艾拉莫德单用组好。4.艾拉莫德与MTX联用有协同增效作用,具有较好的安全性,是RA治疗的一个值得推荐的方案。研究结论艾拉莫德能呈浓度依赖性抑制滑膜FLS增殖及迁移,能降低自身抗体水平,治疗RA改善炎症指标及病情,安全有效,与MTX联用能协同增效,值得推广。
[Abstract]:Background rheumatoid arthritis (RA) is a kind of autoimmune disease characterized by long course of disease and invasive destruction of joints. It is characterized by multiple joint swelling and pain and synovitis. Without regular treatment, joint disability and dysfunction can be caused by involvement of various organs. Among them, synovial tissue is the most vulnerable and fibroblasts. FLS, which has the characteristics of "tumor like", affects the course of RA by secreting inflammatory mediators, proliferation, invasion and migration, and plays an important role in the pathogenesis of joint homeostasis and RA. The proliferation of FLS, migration behavior is influenced by many cytokines and signal pathways, and a new anti rheumatoid drug Ira Maude (T-614 It can act on some common cytokines and signaling pathways. Therefore, this article is to investigate whether Ira Maude can affect the proliferation and migration of FLS and to observe the effect of Ira Maude by clinical observation. Objective 1. primary culture RA patients' synovial cells were cultured in vitro to understand the effect of Ira Maude on the proliferation and migration of FLS.2 To observe and understand the different Ira Maude medication methods, and compare with the traditional DMARDs combined drugs, to understand the effect of the clinical serological index and the condition improvement of the RA patients. The tissue block method was used to train the synovial cells of the patients with RA. Under the aseptic condition, the synovial tissue of the patients with RA was selected by arthroscopy, and the culture Petri dish and the inversion were placed in the culture dish. In the incubator, the cytotoxic effect of Ella mod (100200400800 ng/ml) on the synovial cells after 72 hours was observed, whether some cell apoptosis was caused, and whether the synovial cells had migration behavior, the apoptosis experiment was completed to determine the effective concentration of apoptosis and avoid interference with the subsequent experiment. Edu cell proliferation test, cell migration Transwell experiment to understand the effect of Ira Maude on the proliferation and migration of RAFLS, and explore possible mechanisms. Clinical trials screening 90 patients with rheumatoid arthritis and disease activity, including 19 men, 71 women, 21~67 years old, the average age of 47.23 + 15.62 years, and the average course of 4.75 +. 3.53 years), the patients were randomly divided into 3 groups, and the combined treatment group (31 cases) was given Ira Maude 25mg/bid+MTX 10mg/qw, Ira Maude single drug group (28 cases) Ira Maude 25mg/bid, and the control group (31 cases) was given methotrexate (MTX) 10mg/qw+ leflunomide (LEF) 20mg/qd. to observe the anti CCP antibody in 0,1,3,6,12 month, respectively, RF, ESR, CRP, and the patient's score. DAS28-ESR, HAQ score and other indexes were changed. In the observation, 8 cases were withdrawn due to individual reasons or adverse drug reactions. At the same time, 82 cases were conformed to the inclusion criteria, excluding standards and completed the whole clinical observation process. The quantitative data in this study were all advanced before statistical analysis. The normal distribution of normal distribution data was represented by mean number + standard deviation, and the non normal distribution was represented by median M. Statistical analysis was used to analyze the indexes of each time point before and during the treatment. Whether there were statistical significance, and comparing each index at any time in different treatment groups, whether there was statistical difference between groups, was statistically significant. Results 1. Ella mod (when the concentration reaches 800ng/ml) can promote RAFLS apoptosis. When the concentration is less than 400ng/ml, there is no obvious effect on apoptosis; Ira Maude (when the concentration reaches 400ng/ml) can inhibit the proliferation of RAFLS in the synovial membrane. Ella mod (when the concentration reaches 200ng/ml) can inhibit the RAFLS migration of the synovium, which is a concentration dependent.2. Ella. Mod can reduce the level of anti CCP antibody and RF. The curative effect of Ira Maude combined with MTX in descending antibody is better than that of the traditional DMARDs combined group (MTX+LEF). The long-term effect of.3. traditional DMARDs combined group (MTX+LEF) in the erythrocyte sedimentation rate, CRP and other indexes is more synergistic than the combination of.4. Ella Maude and MTX, which has better safety. Full nature is a recommendable scheme for RA treatment. Conclusion Ira Maude can show the proliferation and migration of FLS in a concentration dependent suppressing synovial membrane. It can reduce the level of autoantibody and improve the inflammatory index and condition of RA. It is safe and effective. It can be synergized with the combination of MTX, and it is worth promoting.
【学位授予单位】：南方医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R593.22
本文编号：2149346
[Abstract]:Background rheumatoid arthritis (RA) is a kind of autoimmune disease characterized by long course of disease and invasive destruction of joints. It is characterized by multiple joint swelling and pain and synovitis. Without regular treatment, joint disability and dysfunction can be caused by involvement of various organs. Among them, synovial tissue is the most vulnerable and fibroblasts. FLS, which has the characteristics of "tumor like", affects the course of RA by secreting inflammatory mediators, proliferation, invasion and migration, and plays an important role in the pathogenesis of joint homeostasis and RA. The proliferation of FLS, migration behavior is influenced by many cytokines and signal pathways, and a new anti rheumatoid drug Ira Maude (T-614 It can act on some common cytokines and signaling pathways. Therefore, this article is to investigate whether Ira Maude can affect the proliferation and migration of FLS and to observe the effect of Ira Maude by clinical observation. Objective 1. primary culture RA patients' synovial cells were cultured in vitro to understand the effect of Ira Maude on the proliferation and migration of FLS.2 To observe and understand the different Ira Maude medication methods, and compare with the traditional DMARDs combined drugs, to understand the effect of the clinical serological index and the condition improvement of the RA patients. The tissue block method was used to train the synovial cells of the patients with RA. Under the aseptic condition, the synovial tissue of the patients with RA was selected by arthroscopy, and the culture Petri dish and the inversion were placed in the culture dish. In the incubator, the cytotoxic effect of Ella mod (100200400800 ng/ml) on the synovial cells after 72 hours was observed, whether some cell apoptosis was caused, and whether the synovial cells had migration behavior, the apoptosis experiment was completed to determine the effective concentration of apoptosis and avoid interference with the subsequent experiment. Edu cell proliferation test, cell migration Transwell experiment to understand the effect of Ira Maude on the proliferation and migration of RAFLS, and explore possible mechanisms. Clinical trials screening 90 patients with rheumatoid arthritis and disease activity, including 19 men, 71 women, 21~67 years old, the average age of 47.23 + 15.62 years, and the average course of 4.75 +. 3.53 years), the patients were randomly divided into 3 groups, and the combined treatment group (31 cases) was given Ira Maude 25mg/bid+MTX 10mg/qw, Ira Maude single drug group (28 cases) Ira Maude 25mg/bid, and the control group (31 cases) was given methotrexate (MTX) 10mg/qw+ leflunomide (LEF) 20mg/qd. to observe the anti CCP antibody in 0,1,3,6,12 month, respectively, RF, ESR, CRP, and the patient's score. DAS28-ESR, HAQ score and other indexes were changed. In the observation, 8 cases were withdrawn due to individual reasons or adverse drug reactions. At the same time, 82 cases were conformed to the inclusion criteria, excluding standards and completed the whole clinical observation process. The quantitative data in this study were all advanced before statistical analysis. The normal distribution of normal distribution data was represented by mean number + standard deviation, and the non normal distribution was represented by median M. Statistical analysis was used to analyze the indexes of each time point before and during the treatment. Whether there were statistical significance, and comparing each index at any time in different treatment groups, whether there was statistical difference between groups, was statistically significant. Results 1. Ella mod (when the concentration reaches 800ng/ml) can promote RAFLS apoptosis. When the concentration is less than 400ng/ml, there is no obvious effect on apoptosis; Ira Maude (when the concentration reaches 400ng/ml) can inhibit the proliferation of RAFLS in the synovial membrane. Ella mod (when the concentration reaches 200ng/ml) can inhibit the RAFLS migration of the synovium, which is a concentration dependent.2. Ella. Mod can reduce the level of anti CCP antibody and RF. The curative effect of Ira Maude combined with MTX in descending antibody is better than that of the traditional DMARDs combined group (MTX+LEF). The long-term effect of.3. traditional DMARDs combined group (MTX+LEF) in the erythrocyte sedimentation rate, CRP and other indexes is more synergistic than the combination of.4. Ella Maude and MTX, which has better safety. Full nature is a recommendable scheme for RA treatment. Conclusion Ira Maude can show the proliferation and migration of FLS in a concentration dependent suppressing synovial membrane. It can reduce the level of autoantibody and improve the inflammatory index and condition of RA. It is safe and effective. It can be synergized with the combination of MTX, and it is worth promoting.
【学位授予单位】：南方医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R593.22
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