祛痰化瘀利湿方对人滑膜细胞Wnt/β-catenin信号通路β-catenin、Cyclin D1、MMP-7的调控作用  

Regulatory effect of “Qutan Huayu Lishi Formula” on β-catenin,Cyclin D1,MMP-7 of the Wnt /β-catenin signaling pathway of human synoviocytes

作　　者：赵婧[1] 肖志锋[1] 阚卫兵[1] 宋朋飞[1] 姚丽[1] 姜玉祥[1] 谢殿洪[1] 于为国[1] 

机构地区：[1]上海中医药大学附属普陀医院伤骨科,上海200062

出　　处：《上海中医药杂志》2014年第7期91-94,共4页Shanghai Journal of Traditional Chinese Medicine

基　　金：国家中医药管理局中医药重点学科建设项目(201232)上海市卫生局课题资助(20114059);上海市教委创新课题资助项目(14YZ058);上海中医药大学附属普陀医院课题资助项目(2013GQ015Ⅱ)

摘　　要：目的观察祛痰化瘀利湿方对人滑膜细胞Wnt/β-catenin信号通路的调控作用。方法将GSK-3β选择性小分子抑制剂作用于正常人滑膜细胞,以激活Wnt/β-catenin信号通路。提取并检测胞核蛋白β-catenin的表达,从而确定最佳激活时间点。将对数生长期的滑膜细胞分为正常组、SB-216763激活组、祛痰化瘀利湿方组,分别采取相应的干预措施。Western blot方法检测滑膜细胞胞核蛋白β-catenin及其下游基因Cyclin D1和MMP-7的表达,酶联免疫吸附试验(ELISA)检测滑膜细胞培养上清液中Cyclin D1和MMP-7的表达。结果成功激活正常人滑膜细胞Wnt/β-catenin信号通路,SB-216763干预后激活组胞核蛋白β-catenin及其下游基因Cyclin D1和MMP-7表达较正常组显著升高(P<0.05);经中药含药血清干预后,祛痰化瘀利湿方组胞核蛋白β-catenin及其下游基因Cyclin D1和MMP-7表达与激活组相比显著下调(P<0.01)。结论印证了GSK-3β作为选择性小分子抑制剂能够激活人滑膜细胞Wnt/β-catenin信号通路;通路激活后,其下游基因Cyclin D1和MMP-7的表达显著升高;祛痰化瘀利湿方能够明显下调β-catenin、Cyclin D1、MMP-7的表达;提示从单一信号通路方面可阐释祛痰化瘀利湿方治疗骨性关节炎的分子作用机制。Objective To observe the regulatory effect of＂Qutan Huayu Lishi Formula＂on β-catenin,Cyclin D1,MMP-7 of the Wnt /β-catenin signaling pathway of human synoviocytes. Methods GSK-3β（Glycogen synthase kinase-3β） selective small molecule inhibitors in normal human synoviocytes（HS） were used to activate the Wnt /β-catenin signaling pathway. We extracted and detected nuclear β-catenin protein expression,thereby to determine the optimal activation time point. The synoviocytes in logarithmic phase were divided into the normal group,SB-216763 activation group,＂Qutan Huayu Lishi Formula＂group. The corresponding measures were used to each group. We detected the expressions of HS nuclear proteins β-catenin and its downstream gene Cyclin D1,MMP-7 were detected by Western Blotting. Expressions of Cyclin D1 and MMP-7 in culture supernatant fluid of synoviocytes were detected by ELISA. Results In this study,we have been successfully activated normal human synovial cells of Wnt /-catenin signaling pathway. After intervened by SB-216763,nuclear protein β-catenin and its downstream gene Cyclin D1 and MMP-7 expression of the activation group were significantly higher than those of the normal group（ P〈0. 05）; after intervened by serum containing＂Qutan Huayu Lishi Formula＂,the expressions of β-catenin,Cyclin D1 and MMP-7 of the＂Qutan Huayu Lishi Formula＂group were significantly down-regulated,with significant difference from that of the activation group（ P〈0. 01）. Conclusion This study confirms the GSK-3β as a selective small molecule inhibitor of human synovial cells capable of activating Wnt / β-catenin signaling pathway; after activating the pathway,its downstream genes Cyclin D1 and MMP-7 expressions were significantly increased; while＂Qutan Huayu Lishi Formula＂can significantly lowered β-catenin,Cyclin D1,MMP-7expression results in favor of the signal path from a single aspect of interpretation dampness phlegm stasis treatment of osteoarthritis molecular mechanisms of inflammatio

关 键 词：滑膜细胞 SB-216763 祛痰化瘀利湿方 Wnt Β-CATENIN信号通路 CYCLIN D1 MMP-7 实验研究 

分 类 号：R285[医药卫生—中药学]