Cell therapy: A therapeutic alternative to treat focal cartilage lesions
Cell therapy: A therapeutic alternative to treat focal cartilage lesions
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DOI：
10.1016/j.transproceed.2005.09.122
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发表时间：
2005-11-01
影响因子：
0.9
通讯作者：
Blanco, FJ
中科院分区：
文献类型：
--
作者：
Gimeno, MJ;Maneiro, E;Blanco, FJ
背景人骨髓间充质干细胞（MSC）存在于大多数组织基质中，当损伤或损伤发生时参与其再生。本研究的目的是研究骨关节炎（OA）患者滑膜中具有多能特性的细胞的存在以及这些细胞分化为软骨细胞的能力。用胶原酶消化OA患者的滑膜（n = 8）。用DMEM、20%FBS和FGFb 10 ng/mL培养分离的细胞。来自第二次传代培养的细胞用于进行表型表征实验（用11种单克隆抗体进行流式细胞术分析）和软骨形成分化实验（在软骨形成培养基中培养的微团）。通过以下技术定量软骨细胞外基质成分来评估细胞的成软骨分化：番红O、甲苯胺蓝和阿尔新蓝染色检测蛋白多糖和免疫组织化学检测I型和II型胶原。流式细胞术分析显示，在我们的群体中，超过90%的细胞对MSC标志物呈阳性：CD 29（95%），CD 44（90%），CD 73（95%），CD 90（98%）。细胞的造血标志物（CD 11b、CD 34和CD 45）呈阴性。此外，细胞对多能标记物如CD 117（c-kit）（98%）、CD 166（74%）和STRO-1（88%）以及静止卫星细胞如PAX-7（35%）显示阳性染色。微粒分析显示，这些细胞与TGF β-3培养2周和3周刺激蛋白聚糖和II型胶原蛋白的合成。这两种分子都是透明关节软骨的特征。在这项工作中，我们证明了在OA患者的滑膜组织中存在具有MSC特征的细胞群。由于MSC参与成人组织的修复过程，这些细胞可能在OA发病机制和治疗中发挥重要作用。
Background. Human mesenchymal stem cells (MSCs) are present in most of the tissue matrix, taking part in their regeneration when injury or damage occurs. The aim of this study was to investigate the presence of cells with pluripotential characteristics in synovial membranes from osteoarthritic (OA) patients and the capacity of these cells to differentiate to chondrocytes.Methods. Synovial membranes (n = 8) from OA patients were digested with collagenase. Isolated cells were cultured with DMEM, 20% FBS, and FGFb10 ng/mL. Cells from second subculture were used to carry out phenotypic characterization experiments (flow cytometry analysis with 11 monoclonal antibodies) and chondrogenic differentiation experiments(micropellet cultured in chondrogenic medium). Chondrogenic differentiation of cells was assessment by quantification of cartilage extracellular matrix components by following techniques: Safranin O, Toluidine Blue, and Alcian Blue stains to detect proteoglycans and immunohistochemistry to detect type I and II collagen.Results. Flow cytometry analyses showed that in our population more than 90% of cells were positive for MSC markers: CD29 (95%), CD44 (90%), CD73 (95%), CD90 (98%). Cells were negative for hematopoietic markers (CD11b, CD34, and CD45). Furthermore, cells showed positive stain to multipotent markers such as CD117 (c-kit) (98%), CD166 (74%), and STRO-1 (88%) and to quiescent satellite cells like PAX-7 (35%). The micropellet analyses showed that the culture of these cells with TGFbeta-3 for 2 and 3 weeks stimulates proteoglycan and collagen type II synthesis. Both molecules are characteristic of hyaline articular cartilage.Conclusion. In this work, we demonstrate the presence of a cellular population with MSC characteristics in synovial tissue from OA patients. As MSC takes part in reparative processes of adult tissues, these cells could play an important role in OA pathogenesis and treatment.