Department of Orthopaedics, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200011, China.
Chin Med J (Engl). 2010 Nov;123(21):3040-8.
Synovium-derived stem cells (SDSCs) with higher chondrogenic potential are attracting considerable attention as a cell source for cartilage regeneration. We investigated the effect of bone morphogenetic protein 2 (BMP-2) on transforming growth factor beta3 (TGF-β3)-induced chondrogenesis of SDSCs isolated from human osteoarthritic synovium in a pellet culture system.
The clonogenicity, stem cell marker expression and multi-differentiation potential of isolated SDSCs were determined by colony forming unit assay, flow cytometry and specific staining including alizarin red S, Oil red O and alcian blue staining, respectively. SDSCs pellet was cultured in chondrogenic medium with or without TGF-β3 or/and BMP-2. At day 21, the diameter and the weight of the pellets were measured. Chondrogenic differentiation of SDSCs was evaluated by Safranin O staining, immunohistochemical staining of collagen type II, sulfated glycosaminoglycan (sGAG) synthesis and mRNA expression of collagen type II, aggrecan, SOX9, link-protein, collagen type X and BMP receptor II.
Cells isolated under the optimized culturing density (10(4)/60 cm(2)) showed clonogenicity and multi-differentiation potential. These cells were positive (> 99%) for CD44, CD90, CD105 and negative (< 10%) for CD34 and CD71. SDSCs differentiated to a chondrocytic phenotype in chondrogenic medium containing TGF-β3 with or without BMP-2. Safranin O staining of the extracellular matrix was positive and the expression of collagen type II was detected. Cell pellets treated with TGF-β3 and BMP-2 were larger in diameter and weight, produced more sGAGs, and expressed higher levels of collagen type II and other chondrogenic markers, except COL10A1, than medium with TGF-β3 alone.
SDSCs could be isolated from human osteoarthritic synovium. Supplementation with BMP-2 significantly promoted the in vitro TGF-β3-induced chondrogenic differentiation of SDSCs.
具有更高成软骨潜力的滑膜衍生干细胞（SDSCs）作为软骨再生的细胞来源备受关注。我们在微球体培养系统中研究了骨形态发生蛋白 2（BMP-2）对分离自人骨关节炎滑膜的 SDSCs 在转化生长因子β3（TGF-β3）诱导下的成软骨分化的影响。
通过集落形成单位测定、流式细胞术和茜素红 S、油红 O 和阿利新蓝染色等特异性染色，确定分离的 SDSCs 的克隆形成能力、干细胞标志物表达和多向分化潜能。SDSCs 微球体在含有或不含有 TGF-β3 或/和 BMP-2 的软骨形成培养基中培养。在第 21 天，测量球体的直径和重量。通过番红 O 染色、Ⅱ型胶原免疫组织化学染色、硫酸软骨素聚糖（sGAG）合成以及Ⅱ型胶原、聚集蛋白聚糖、SOX9、连接蛋白、X 型胶原和 BMP 受体 II 的 mRNA 表达评估 SDSCs 的软骨分化。
在优化的培养密度（10(4)/60 cm(2))下分离的细胞具有克隆形成能力和多向分化潜能。这些细胞对 CD44、CD90、CD105 呈阳性（>99%），对 CD34 和 CD71 呈阴性（<10%）。在含有 TGF-β3 的软骨形成培养基中，SDSCs 分化为软骨细胞表型，无论是否存在 BMP-2 均如此。细胞外基质的番红 O 染色呈阳性，且检测到Ⅱ型胶原的表达。与仅用 TGF-β3 处理的细胞相比，用 TGF-β3 和 BMP-2 处理的细胞微球体直径和重量更大，产生的 sGAG 更多，且表达更高水平的Ⅱ型胶原和其他软骨形成标志物，除 COL10A1 外。
可以从人骨关节炎滑膜中分离出 SDSCs。补充 BMP-2 可显著促进 SDSCs 在体外 TGF-β3 诱导下的成软骨分化。
