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目的:观察桂枝芍药知母汤(GD)对尿酸钠致痛风性关节炎(GA)模型大鼠关节滑膜组织中Toll-髓性分化因子88(My D88)信号通路炎性信号表达的影响,探讨其相关的作用机制。方法:180只雄性SD大鼠随机分配到3个实验,分别为关节滑膜免疫组织化学技术(IHC)实验、酶联免疫吸附测定(ELISA)实验、蛋白质免疫印迹(Western blot)实验。各实验取大鼠60只,按体重随机分为6组,每组10只,分别为模型组,正常组,GD高、中、低剂量组(4,8,16 g·kg~(-1)),秋水仙碱阳性药组(3×10~(-4)g·kg~(-1))。实验组均ig给药,正常组、模型组给予等容积的蒸馏水,每天1次,连续给药7 d。第5天ig前,大鼠足踝关节注射尿酸钠悬液诱导GA。取大鼠关节滑膜组织,IHC检测受体Toll样受体-2(Toll-like receptor 2,TLR-2),TLR-4的表达,Image-Pro Plus 6.0图像分析系统测定平均积分吸光度IA,ELISA测定环氧化酶-2(cyclooxygenase-2,COX-2),转化生长因子-β_1(transforming growth factor-β_1,TGF-β_1)表达,Western blot检测My D88,核因子κB酶抑制剂-β(inhibitorκB kinaseβ,IκK-β),核因子κB抑制蛋白α(NF-κB inhibitorα,IκB-α),过氧化物酶体增殖物激活受体-γ(peroxisome proliferator-activated receptor-γ,PPAR-γ)表达水平。结果:造模72 h后,与正常组比较,GA模型大鼠关节滑膜组织中TLR-2,TLR-4平均IA,My D88,IκK-β蛋白及COX-2表达水平明显增高(P<0.05);与模型组比较,GD中、高剂量组TLR-2,TLR-4平均IA,My D88,IκK-β蛋白及COX-2含量表达均明显低于模型组(P<0.05);而TGF-β_1含量及IκB-α,PPAR-γ蛋白表达水平明显增高(P<0.05);GD各剂量组IκK-β蛋白无明显变化。结论:GD治疗GA的作用机制可能与降低TLR-2,TLR-4受体及My D88蛋白表达,增加PPAR-γ,IκB-α表达,抑制NF-κB活化,降低Toll-My D88信号通路炎性因子表达有关。
AIM: To observe the effect of Guizhishaoyaozhimu Tang (GD) on the inflammatory signal expression of Toll-myeloid differentiation factor 88 (My D88) signal pathway in synovial tissue of rats with urate induced gouty arthritis (GA) , To explore its related mechanism of action. Methods: One hundred and eighty male Sprague-Dawley rats were randomly assigned to three experiments: synovial immunohistochemistry (IHC), enzyme-linked immunosorbent assay (ELISA) and Western blot. Sixty rats in each experiment were randomly divided into 6 groups according to body weight, with 10 rats in each group. The rats in model group, normal group, GD high, middle and low dose groups (4, 8, 16 g · kg ~ (-1) ), And colchicine positive group (3 × 10 -4 g · kg -1). Experimental group were ig administration, normal group, model group given equal volume of distilled water, 1 day, continuous administration of 7 d. On the fifth day before ig, rats were injected with sodium urate suspension into the foot and ankle to induce GA. The expression of TLR-2 and TLR-4 in the synovial tissue of rats was detected by IHC. The average integral absorbance (IA) was measured by Image-Pro Plus 6.0 image analysis system. The expression of cyclooxygenase-2 (COX-2) and transforming growth factor-β1 (TGF-β1) was detected by ELISA. The expression of My D88, the inhibitor of nuclear factor κB (inhibitor ofκB kinaseβ, IκK-β), nuclear factor κB inhibitorα (IκB-α), peroxisome proliferator-activated receptor-γ (PPAR-γ) )The expression level. Results: Compared with the normal group, the expression of TLR-2 and TLR-4, IA, My D88, IκK-β and COX-2 in GA synovial tissue were significantly increased at 72 h after modeling 0.05). Compared with model group, the contents of IA, My D88, IκK-β and COX-2 in TLR-2 and TLR-4 were significantly lower than those in model group (P <0.05) The content of TGF-β 1 and the expression of IκB-α and PPAR-γ were significantly increased (P <0.05). There was no significant change in the expression of IκK-β in each dose of GD. Conclusion: The mechanism of GA in treating GA may decrease the expression of TLR-2, TLR-4 receptor and My D88, increase the expression of PPAR-γ and IκB-α, inhibit the activation of NF-κB and decrease the expression of Toll-My D88 signaling pathway Sex factor expression.