Trinity Centre for Bioengineering, Trinity Biomedical Sciences Institute, Trinity College Dublin, Dublin, Ireland.
Tissue Eng Part C Methods. 2013 Feb;19(2):117-27. doi: 10.1089/ten.TEC.2012.0142. Epub 2012 Sep 17.
Human infrapatellar fat pad contains a source of mesenchymal stem cells (FPSCs) that potentially offer a novel population for the treatment of damaged or diseased articular cartilage. Existing cartilage repair strategies such as microfracture harness the presence of a low-oxygen microenvironment, fibrin clot formation at sites of microfracture, and elevations in growth factors in the damaged joint milieu. Bearing this in mind, the objective of this study was to determine the chondrogenic potential of diseased human FPSCs in a model system that recapitulates some of these features. In the first phase of the study, the role of transforming growth factor beta-3 (TGF-β3) and fibroblast growth factor-2 (FGF-2), in addition to an altered oxygen-tension environment, on the colony-forming unit-fibroblast (CFU-F) capacity and growth kinetics of human FPSCs during monolayer expansion was evaluated. The subsequent chondrogenic capacity of these cells was quantified in both normoxic (20%) and low- (5%) oxygen conditions. Expansion in FGF-2 was shown to reduce CFU-F numbers, but simultaneously increase both the colony size and the cell yield compared to standard expansion conditions. Supplementation with both FGF-2 and TGF-β3 significantly reduced cell-doubling time. Expansion in FGF-2, followed by differentiation at 5% oxygen tension, was observed to synergistically enhance subsequent sulfated glycosaminoglycan (sGAG) accumulation after chondrogenic induction. FPSCs expanded in FGF-2 were then encapsulated in either agarose or fibrin hydrogels in an attempt to engineer cartilaginous grafts. sGAG synthesis was higher in fibrin constructs, and was further enhanced by differentiation at 5% oxygen tension, accumulating 2.7% (ww) sGAG after 42 days in culture. These results indicate that FPSCs, a readily accessible cell population, form cartilage in an in vitro environment that recapitulates several key biological features of cartilage repair during microfracture and also point toward the potential utility of such cells when combined with fibrin hydrogel scaffolds.
人髌下脂肪垫含有间充质干细胞（FPSCs）的来源，这些细胞可能为治疗受损或患病的关节软骨提供一种新的细胞来源。现有的软骨修复策略，如微骨折，利用了低氧微环境、微骨折部位纤维蛋白凝块的形成以及受损关节环境中生长因子的升高。考虑到这一点，本研究的目的是在一个模型系统中确定患病人类 FPSCs 的软骨生成潜力，该模型系统再现了其中的一些特征。在研究的第一阶段，评估了转化生长因子β-3（TGF-β3）和成纤维细胞生长因子-2（FGF-2）的作用，以及改变的氧张力环境对人类 FPSCs 在单层扩增过程中的集落形成单位成纤维细胞（CFU-F）能力和生长动力学的影响。随后在常氧（20%）和低氧（5%）条件下，对这些细胞的随后软骨生成能力进行了量化。与标准扩增条件相比，FGF-2 的扩增显示出降低 CFU-F 数量，但同时增加集落大小和细胞产量。同时补充 FGF-2 和 TGF-β3 可显著缩短细胞倍增时间。在 FGF-2 扩增后，在 5%氧张力下进行分化，观察到协同增强随后软骨诱导后的硫酸化糖胺聚糖（sGAG）积累。然后将在 FGF-2 中扩增的 FPSCs 包埋在琼脂糖或纤维蛋白水凝胶中，试图构建软骨移植物。纤维蛋白构建体中的 sGAG 合成更高，在 5%氧张力下分化进一步增强，在培养 42 天后积累 2.7%（ww）sGAG。这些结果表明，FPSCs 是一种易于获得的细胞群体，在体外环境中形成软骨，该环境再现了微骨折过程中软骨修复的几个关键生物学特征，并且当与纤维蛋白水凝胶支架结合使用时，这些细胞具有潜在的用途。
