痹祺提取物对CIA大鼠滑膜增殖及软骨破坏的影响

发布时间：2018-10-20 09:06

【摘要】：目的:类风湿关节炎(rheumatoid arthritis,RA)以关节肿痛及进行性破坏为主要临床表现,是一种长期的、慢性的、全身性的自身免疫性疾病,该病女性多发,且30-50岁为高发年龄,中国大陆的患病率为0.2%-4%。目前针对RA的纤维样滑膜细胞(Fibroblast like synoviocytes,FLS)的增生、转化及侵袭、滑膜新生血管增殖、血管翳的形成、软骨的降解与破坏、骨质侵蚀等病理方面的靶向性治疗为当前研究的重点与难点。痹祺提取物为目前临床治疗RA应用较多的药物,但其对RA的药理作用机制的研究尚不完整,故本实验以CIA大鼠为动物模型,模拟RA发病过程,观察痹祺提取物对CIA大鼠TNF-α、IL-18、OPN、COMP的血清水平及OPN、COMP分别在软骨、滑膜表达的影响,初步探讨痹祺胶囊干预CIA大鼠关节炎症、滑膜增生及软骨破坏的药理作用机制,为临床上治疗RA提供可靠的实验证据。方法:[1]建立CIA大鼠模型,观察并计算大鼠足肿胀率。[2]采用ELISA检测大鼠造模2W时血清TNF-α及灌药4W后血清TNF-α、IL-18、OPN、COMP水平。[3]采用IHC检测OPN、COMP在滑膜中的表达。[4]采用RT-PCR检测OPN、COMP在软骨中的表达。[5]显微镜下观察HE染色滑膜组织、软骨组织病理形态。结果:[1]成功建立CIA模型,CIA模型组较正常组大鼠足肿胀率明显升高,治疗组较模型组大鼠足肿胀率明显减低(P0.05)。[2]细胞因子血清水平比较:CIA模型组较正常组大鼠治疗后TNF-α、IL-18、OPN、COMP在血清中表达量升高(P0.05),治疗组较模型组表达量降低(P0.05)。[3]滑膜组织中细胞因子的蛋白表达:正常组滑膜区可见OPN少量表达,正常组滑膜区未见COMP的表达,模型组滑膜区可见大量阳性细胞,模型组较正常组阳性细胞增多(P0.05),治疗组滑膜区可见部分阳性细胞,较模型组减少(P0.05)。[4]软骨中细胞因子的蛋白表达:实时相对定量分析结果显示大鼠软骨OPN mRNA、COMP mRNA表达模型组较正常组升高(P0.05),治疗组较模型组降低(P0.05)。[5]HE染色病理图片观察:参与滑膜组织病理评分计算滑膜炎症程度、软骨病理评分计算软骨损伤程度,模型组较正常组病理评分升高(P0.05),治疗组较模型组病理评分下降(P0.05),中剂量组在炎性细胞浸润及滑膜细胞增生及软骨损伤方面较其他治疗组降低(P0.05)。结论:[1]痹祺提取物干预大鼠TNF-α、IL-18在血清中的表达,阻断炎性因子网络形成,从而干预CIA关节炎症的进一步发展。[2]痹祺提取物可能通过下调大鼠OPN在血清中及OPN、COMP在滑膜中的表达,干预滑膜肿瘤样增生,从而抑制滑膜血管新生及血管翳的形成。[3]痹祺提取物可能通过下调大鼠OPN、COMP在软骨中的表达,干预软骨基质降解,从而阻止软骨的破坏。[4]大鼠COMP血清水平模型组较正常组增加,反映CIA大鼠出现软骨破坏,经过痹祺提取物治疗的大鼠COMP血清水平较模型组下降,反映其治疗CIA大鼠软骨破坏有效。
[Abstract]:Objective: rheumatoid arthritis (rheumatoid arthritis,RA) is a long-term, chronic, systemic autoimmune disease with the main clinical manifestation of joint swelling and pain and progressive destruction. The prevalence rate in mainland China was 0.2-4. The proliferation, transformation and invasion of RA fibroid synovial cells (Fibroblast like synoviocytes,FLS), the proliferation of synovial neovascularization, the formation of pannus, the degradation and destruction of cartilage, Targeted treatment of bone erosion and other pathological aspects is the focus and difficulty of current research. Biqi extract is a widely used drug in clinical treatment of RA at present, but its pharmacological mechanism to RA is not complete. Therefore, CIA rats are used as animal models in this experiment to simulate the pathogenesis of RA. To observe the effects of Biqi extract on the serum levels of TNF- 伪 and IL-18,OPN,COMP and the expression of OPN,COMP in cartilage and synovium of CIA rats, and to explore the pharmacological mechanism of Biqi capsule in preventing arthritis, synovial hyperplasia and cartilage destruction in CIA rats. To provide reliable experimental evidence for clinical treatment of RA. Methods: [1] CIA rat model was established. The rate of rat foot swelling was observed and calculated. [2] the levels of serum TNF- 伪 and IL-18,OPN,COMP were detected by ELISA at 2 W and 4 W after administration. [3] IHC was used to detect the expression of OPN,COMP in synovium. [4] RT-PCR was used to detect the expression of OPN,COMP in cartilage. [5] Microscopy was used to detect the expression of OPN,COMP. HE staining of synovial tissue, Cartilage histopathology. Results: [1] CIA model was successfully established, and the rate of foot swelling in the CIA model group was significantly higher than that in the normal group. (2) comparison of serum levels of cytokines: the expression of TNF- 伪 and IL-18,OPN,COMP in serum of CIA model group was higher than that of normal group (P0.05), and that of treatment group was lower than that of model group (P0.05). [3] in synovial tissue, the expression of TNF- 伪 and IL-18,OPN,COMP in the treatment group was significantly lower than that in the model group (P0.05). Protein expression of cytokines: in normal group, a small amount of OPN was expressed in synovial region. The expression of COMP was not found in the synovial region of the normal group. A large number of positive cells were found in the synovial area of the model group, and the number of positive cells in the model group was higher than that in the normal group (P0.05). Some positive cells were found in the synovial area of the treatment group. Compared with the model group (P0.05). [4] the expression of cytokines in cartilage: the results of real-time relative quantitative analysis showed that the expression of OPN mRNA,COMP mRNA in the model group was higher than that in the normal group (P0.05), and that in the treatment group was lower than that in the model group (P0.05). [5] the pathological pictures of HE staining were observed. The degree of synovitis was calculated by participating in synovial tissue pathological score. The degree of cartilage injury in model group was higher than that in normal group (P0.05), the pathological score in treatment group was lower than that in model group (P0.05), and the inflammatory cell infiltration, synovial cell proliferation and cartilage injury in middle dose group were lower than that in other treatment group (P0.05). Conclusion: [1] Biqi extract interferes with the expression of TNF- 伪 and IL-18 in serum of rats, blocks the formation of inflammatory factor network, and interferes with the further development of CIA arthritis. [2] Biqi extract may down-regulate the expression of OPN in serum and OPN,COMP in synovium of rats. Intervention of synovial tumor-like hyperplasia, thereby inhibiting the formation of synovial angiogenesis and pannus. [3] Biqi extract may interfere with cartilage matrix degradation by down-regulating the expression of OPN,COMP in cartilage. [4] the serum level of COMP in the model group was higher than that in the normal group, reflecting cartilage destruction in the CIA rats, and the serum level of COMP in the rats treated with Biqi extract was lower than that in the model group. It is effective in the treatment of cartilage destruction in CIA rats.
【学位授予单位】：天津医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R593.22

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