类风湿关节炎滑膜成纤维细胞通过高表达RANKL促进破骨细胞分化及活化  

Rheumatoid synovial fibroblasts promote osteoclastogenesis via RANKL

作　　者：郑东辉[1] 戴冽[1] 韦秀宁[1] 朱浪静[1] 莫颖倩[1] 张白玉[1] 邹婵娟[1] 

机构地区：[1]中山大学孙逸仙纪念医院风湿免疫科,广州510120

出　　处：《中华关节外科杂志（电子版）》2011年第5期61-64,共4页Chinese Journal of Joint Surgery(Electronic Edition)

基　　金：国家自然科学基金面上项目(30972742);国家自然科学青年科学基金项目(81001334;30901333);广东省自然科学基金项目(9151008901000130;10451008901004542)

摘　　要：目的探讨类风湿关节炎滑膜成纤维细胞(RA-FLS)对破骨细胞(Oc)分化和活化的作用及机制。方法活动期RA滑膜体外分离培养FLS,以骨关节炎(OA)-FLS为对照,分别与健康人外周血单核细胞(MNC)共培养后,抗酒石酸酸性磷酸酶(TRAP)染色鉴定Oc并计数、甲苯胺蓝染色观察骨吸收陷窝情况。细胞免疫荧光染色检测FLS RANKL表达,Real-time PCR及W estern blot检测RANKL和OPG mRNA、蛋白表达。结果 RA-FLS与MNC共培养7 d时TRAP+且细胞核≥3个的Oc很少,14 d时见较多的Oc,21 d甲苯胺蓝染色示清晰的骨吸收陷窝,而OA-FLS与MNC共培养后未见Oc,也未见骨吸收陷窝。细胞免疫荧光染色示RA-FLS较OA-FLS高表达RANKL(P<0.05)。RA-FLS RANKL mRNA和蛋白表达较OA-FLS明显增高,而OPG mRNA和蛋白表达则明显降低,RANKL/OPG mRNA和蛋白比率较OA-FLS明显增高(均P<0.05)。结论 RA-FLS可能通过高表达RANKL,促进外周血MNC向Oc分化,并促进Oc的骨吸收功能。Objective To investigate the effects and mechanisms of rheumatoid fibroblast-like synoviocytes（FLS） in promoting osteoclastogenesis.Methods FLS were separated and cultured from synovium tissue of patients with active rheumatoid arthritis（RA） or osteoarthritis（OA）.FLS were then cocultured with normal monocytes from healthy volunteers＇ peripheral blood.Osteoclasts were determined and counted by trate-resistant acid phosphatase（TRAP） staining,and resorption lacunae on bone slices were identified by toluidine blue staining.The expression of receptor activator of NF-κB ligand（RANKL） on FLS was evaluated by immunofluorescence.Real-time PCR and Western blot were performed respectively to determine the mRNA and protein expression of RANKL and osteoprotegerin（OPG）.Results At the 7th day of cocultured RA-FLS with normal monocytes,few TRAP-positive cells with three or more nuclei were seen and these osteoclasts increased significantly by the 14th day.Resorption lacunae were observed at the 21st day.However,no osteoclast or resorption lacuna was observed when OA-FLS was cocultured with normal monocytes.Immunofluorescence staining showed significantly higher expression of RANKL in RA-FLS than in OA-FLS（P0.05）.Expression of RANKL mRNA and protein was significantly up-regulated in RA-FLS compared with that in OA-FLS,while expression of OPG mRNA and protein was significantly down-regulated.The ratio of RANKL/OPG mRNA and protein was also up-regulated in RA-FLS more obviously than in OA-FLS（all P0.05）.Conclusion RA-FLS may promote osteoclastogenesis and bone erosion in RA by RANKL up-regulation.

关 键 词：关节炎 类风湿 滑膜 成纤维细胞 破骨细胞 核因子κB受体激活配体 骨保护素 

分 类 号：R593.22[医药卫生—内科学]