晚期骨关节炎患者膝关节软骨下松质骨祖细胞的特性研究

Characteristics of progenitor cells derived from subchondral cortico-spongious bone in end-stage osteoarthritis patients

背景:骨关节炎(osteoarthritis,OA)又名退行性骨关节病,对其治疗目前局限于早期的保守治疗和晚期的关节置换。晚期OA患者软骨下是否具有间充质干细胞(mesenchymal stem cells,MSCs)决定了骨髓刺激技术可否用于晚期软骨缺损为主要表现的OA患者。但晚期OA患者软骨下松质骨是否具有MSCs仍不明确。目的:探讨晚期OA患者膝关节软骨下是否具有成软骨分化能力的松质骨祖细胞,为晚期膝OA患者软骨自身修复的研究奠定基础。方法:5例因晚期膝OA行全膝关节置换术患者的术中标本,取其软骨下松质骨,经分离培养后得到贴壁细胞,传代培养后行流式细胞仪分析细胞表面抗原;成骨、成脂、成软骨分化后分别行碱性磷酸酶活性检测、茜素红染色、油红O染色、阿利新蓝、Ⅱ型胶原和聚蛋白聚糖组织化学染色,Ⅰ型胶原、Ⅱ型胶原、聚蛋白聚糖及SOX-9基因表达水平实时聚合酶连反应(RealTime PCR,RT-PCR)测定。结果:从晚期膝OA患者膝关节软骨下松质骨内获得的贴壁细胞的细胞表面抗原基本符合MSCs标准,且具有成骨、成脂、成软骨分化能力。结论:晚期OA患者软骨下存在具有干细胞性质、可进行体外成软骨分化的松质骨祖细胞。该发现可为"一步式——微骨折处理-成软骨诱导材料填充-软骨修复"治疗理念提供一定依据。

Background：Bone marrow stimulation can be used in end-stage osteoarthritis patients in whom cartilage defects mainly presents only when there were mesenchymal stem cells （MSCs） in the subchondral zone of the knee. But the exis-tence of MSCs in the end-stage osteoarthritis patients is still unknown. Objective：To analyze multi-differentiation capacity of cells derived from human subchondral bone in patients with end-stage knee osteoarthritis. Methods：Human progenitor cells were isolated and derived from subchondral cortico-spongious bone chips from 5 patients undergoing total knee arthro-plasty due to end-stage osteoarthritis. Mesenchymal stem cell related cell surface antigens were analyzed by flow cytometry. After in vitro osteogenic, adipogenic and chondrogenic differentiation, alkaline phosphatase activity, Alizarin red staining, Oil Red O staining, Alcian blue staining and immunohistochemical staining for collagenⅡand Aggrecan were conducted. Real-time PCR was performed to confirm related gene expression of chondrogenic differentiation, such ascollagenⅠ, colla-genⅡ, Aggrecan and SOX-9. Results：Surface antigens of successfully-cultured cells from subchondral cortico-spongious bone chips were almost in accord with those of MSCs. And the cells had the ability of osteogenic, adipogenic and chondro-genic differentiation. Conclusions：Multi-potential progenitor cells do exist in subchondral bone in the knee of patients with end-stage osteoarthritis.