青藤碱对类风湿关节炎成纤维样滑膜细胞MyD88、TRAF-6表达的影响  

MyD88,TRAF-6 expression of sinomenine on rheumatoid arthritis fibroblast-like synoviocytes

作　　者：张洪长[1] 刘明昕[2] 张莹[3] 王恩鹏[1] 陈港[1] 姜鸿远 

机构地区：[1]长春中医药大学中医药与生物工程研究开发中心药理研究室,长春130117 [2]长春中医药大学图书馆参考咨询部,长春130117 [3]吉林大学药学院药理学教研室,长春130021

出　　处：《中国免疫学杂志》2015年第4期485-489,共5页Chinese Journal of Immunology

基　　金：吉林省科技厅科技发展计划项目资助(No.20140414051GH)

摘　　要：目的:探讨青藤碱(Sinomenine,SIN)对TLR信号转导通路及髓样分化因子88(Myeloid differentiation factor88,My D88)、肿瘤坏死因子受体相关因子6(Tumor necrosis factor receptor associated factor-6,TRAF-6)表达的影响,阐明SIN抑制类风湿关节炎(Rheumatoid arthritis,RA)成纤维样滑膜细胞(Fibroblast-like synoviocytes,FLS)增生导致RA软骨和软骨下骨破坏造成关节畸形的作用机制。方法:体外培养RA-FLS细胞,分为对照组与(0.125、0.25、0.5、1 mmol/L)SIN组,通过检测各组细胞内碱性磷酸酶(ALP)活性,确定体外用药最佳药物浓度;CCK-8法检测0.5 mmol/L SIN组的细胞增殖率;荧光定量PCR法测定对照组与0.5 mmol/L SIN组My D88和TRAF-6基因表达;Western blot法检测对照组与0.5 mmol/L SIN组My D88和TRAF-6蛋白表达。结果:SIN各组ALP活性均低于对照组,其中以0.5 mmol/L SIN组的ALP活性为最低(P<0.01)。CCK-8法检测,0.5 mmol/L SIN组RA-FLS细胞增殖率明显低于对照组(P<0.01),SIN诱导第4天细胞增殖率最高,进入平台期,之后细胞的增殖率开始下降。荧光定量PCR和Western blot法检测,0.5 mmol/L SIN组的My D88和TRAF-6基因及蛋白表达明显低于对照组,差异有统计学意义(P<0.01)。结论:SIN通过对TLR信号转导通路的影响有效地抑制RA-FLS细胞内My D88和TRAF-6的表达,这可能是其治疗RA防止软骨和软骨下骨破坏造成关节畸形发生的重要分子机制之一。Objective：To investigate SIN（Sinomenine）for TLR signal transduction pathway and MyD88（Myeloid Differentiation Factor 88）,TRAF-6（ Tumor necrosis factor receptor associated factor-6） expression,clarifying SIN inhibit RA（ Rheumatoid arthritis）-FLS（ Fibroblast-like synoviocytes） proliferation leads to joint deformity of RA cartilage and subchondral bone destruction caused by the role of mechanisms. Methods： RA-FLS cells for vitro were divided into a control group and（ 0. 125,0. 25,0. 5,1 mmol / L） SIN group,within each group were detected by alkaline phosphatase（ ALP） activity,to determine the best concentration for vitro drug;detect 0. 5 mmol / L SIN group in CCK-8 method to detect cell proliferation rate;fluorescence quantitative PCR method MyD88 SIN group and control group with 0. 5 mmol / L TRAF-6 gene expression;Western blot method to detect MyD88 SIN group and control group with 0. 5mmol / L TRAF-6 protein expression. Results： SIN the ALP activity of the lower than the control group,with the minimum ALP activity of 0. 5 mmol / L SIN group（ P〈0. 01）. CCK 8 method,0. 5 mmol / L RA-FLS cell proliferation rate SIN group was obviously lower than the control group（ P〈0. 01）,SIN induce cell proliferation rate was highest,4 days into the plateau,after cell proliferation rate began to fall. Fluorescence quantitative PCR and Western blot method to detect,0. 5 mmol / L SIN group of MyD88 and TRAF-6 gene and protein expression significantly lower than the control group,the difference was statistically significant（ P〈0. 01）. Conclusion： The SIN of TLR signalling pathways through effectively suppress the influence of RA-FLS MyD88 in the cell and the expression of TRAF-6,this could be a treatment of RA prevent cartilage and subchondral bone damage cause joint deformity happened one of the important molecular mechanism.

关 键 词：青藤碱 类风湿关节炎 成纤维样滑膜细胞 髓样分化因子88 肿瘤坏死因子受体相关因子6 

分 类 号：R967[医药卫生—药理学]