塞来昔布对人滑膜细胞和人软骨细胞共培养体系相关因子的调控作用  

Effects of celecoxib on co-culture system of human chondrocytes and synovial cells

作　　者：王德刚[1] 许传勇 姜玉祥[1] 阚卫兵[1] 肖志锋[1] 姚丽[1] 刘婉[1] 谢殿洪[1] 汤豪[1] 张玉婷[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 [2]山东省威海市威海卫人民医院脊柱外科,264200

出　　处：《广东医学》2016年第9期1278-1282,共5页Guangdong Medical Journal

基　　金：上海市教委科研创新项目(编号:14YZ058);上海中医药大学预算内科研项目(编号:2014YSN64)

摘　　要：目的观察塞来昔布对Wnt/β-catenin信号通路活化的人滑膜细胞和人软骨细胞共培养体系相关因子的调控作用,初步阐释塞来昔布治疗骨关节炎的分子作用机制。方法通过β-catenin慢病毒载体转染正常人滑膜细胞,激活其Wnt/β-catenin信号通路。Wnt/β-catenin信号通路活化的人滑膜细胞与正常人软骨细胞置于Transwell小室中共培养。倒置显微镜观察Wnt/β-catenin信号通路活化的人滑膜细胞对人软骨细胞形态学变化的影响。塞来昔布混悬液干预共培养体系,采用Western blot方法检测塞来昔布对滑膜细胞β-catenin蛋白表达的影响;采用ELISA方法检测塞来昔布对滑膜细胞、软骨细胞培养上清液基质金属蛋白酶-7(MMP-7)、Ⅱ型胶原羧基端肽(CTX-Ⅱ)、软骨寡聚基质蛋白(COMP)表达的影响。结果β-catenin慢病毒载体转染正常人滑膜细胞后成功激活Wnt信号通路并与软骨细胞共培养。随着培养时间的延长,软骨细胞生长逐渐受到抑制,软骨细胞胞体边缘与板壁接触处发白,折光度增加,细胞贴壁强度降低。塞来昔布干预后,Western blot检测发现滑膜细胞中β-catenin蛋白的表达明显下调(P<0.01);ELISA法检测发现滑膜与软骨细胞上清液中MMP-7、CTX-Ⅱ、COMP的表达与正常对照相比明显升高(P<0.01)。结论滑膜细胞Wnt/β-catenin信号通路的激活可以使软骨细胞生长受到抑制。塞来昔布可以降低滑膜细胞β-catenin表达,且与干预时间长短相关。塞来昔布可以抑制滑膜细胞Wnt/β-catenin信号通路,下调MMP-7、CTX-Ⅱ、COMP的表达。塞来昔布可能通过抑制OA滑膜Wnt/β-catenin信号通路,改善滑膜炎症、调控滑膜-软骨共同体系微环境、达到抑制软骨降解,促进软骨细胞功能恢复而治疗骨关节炎。Objective To observe the effects of celecoxib on co - culture system of human chondrocytes and syn- ovial cells with activated Wnt/β - catenin signal pathway, and to clarify the molecular mechanism of celeeoxib in the treat- ment of osteoarthritis （OA）. Methods β- catenin lentivirus vector was constructed to transfeet synovial cells of healthy individuals, so as to activate the Wnt/β-catenin signal pathway. These synovial cells and chondrocytes were co -cul- tured in transwell chamber. Inverted microscope was used to observe the effects of activated Wnt/β- catenin signal path- way in human synovial cells on morphological changes of human chondrocytes. The co - cuture system was treated with celecoxib suspensions. Western blot was used to detect the influence of eelecoxib on protein expression of β - catenin in synovial cells. ELISA was used to detect the influence of celecoxib on expressions of MMP - 7, CTX - 11 and COMP in the culture supernatant. Results After being transfected by lentivirus vector, Wnt/β - catenin signal pathway was suc- cessfully activated in the co - culture system with chondrocytes. Growth of chondrocytes was gradually inhibited with in- creasing time of culture. The contact surface between edges of chondrocytes and walls were white, with increased refraction and decreased adhesion. Western bolt showed that expression of β -catenin in synovial cells decreased significantly after have being transfeeted （ P 〈 0. 01 ）. ELISA results showed that celecoxib significantly upregulated the level of MMP - 7, CTX - II , COMP in co - culture cells supernatant （ P 〈 0. 01 ）. Conclusion The growth of chondrocytes can be inhibi- ted by synovial cells with activated Wnt/β - catenin signal pathway. β- catenin expression can be down - regulated by celeeoxib in a time- dependent manner. Celecoxib can inhibit Wnt/β-eatenin signal pathway, down -regulate the ex-pression level of MMP - 7, CTX -Ⅱ, COMP. Celecoxib inhibits the Wnt/β- catenin signal pathway in synovial cells, alleviates

关 键 词：Wnt信号通路 细胞共培养 MMP-7 CTX-Ⅱ COMP 塞来昔布 

分 类 号：R967[医药卫生—药理学]