Stem Cell and Tissue Engineering Laboratory, Department of Orthopaedics, West Virginia University, Morgantown, West Virginia 26506-9196, USA.
Tissue Eng Part A. 2013 May;19(9-10):1144-54. doi: 10.1089/ten.TEA.2012.0351. Epub 2013 Jan 15.
Cartilage defects have a limited ability to self-heal. Stem cell treatment is a promising approach; however, replicative senescence is a challenge to acquiring large-quantity and high-quality stem cells for cartilage regeneration. Synovium-derived stem cells (SDSCs) are a tissue-specific stem cell for cartilage regeneration. Our recent findings suggest that decellularized stem cell matrix (DSCM) can rejuvenate expanded SDSCs in cell proliferation and chondrogenic potential. In this study, we were investigating (1) whether transforming growth factor (TGF)-β1 and TGF-β3 played a similar role in chondrogenic induction of SDSCs after expansion on either DSCM or plastic flasks (plastic), and (2) whether DSCM-expanded SDSCs had an enhanced capacity in repairing partial-thickness cartilage defects in a minipig model. SDSCs were isolated from synovium in two 3-month-old pigs and DSCM was prepared using SDSCs. Passage 2 SDSCs were expanded on either DSCM or plastic for one passage. The expanded cells were evaluated for cell morphology, chondrogenic capacity, and related mechanisms. TGF-β1 and TGF-β3 were compared for their role in chondrogenesis of SDSCs after expansion on either DSCM or plastic. The chondrogenic induction medium without TGF-β served as a control. In 13 minipigs, we intraarticularly injected DSCM- or plastic-expanded SDSCs or saline into knee partial-thickness cartilage defects and assessed their repair using histology and immunohistochemistry. We found DSCM-expanded SDSCs were small, had a fibroblast-like shape, and grew quickly in a three-dimensional format with concomitant up-regulation of phosphocyclin D1 and TGF-β receptor II. Plastic-expanded SDSCs exhibited higher mRNA levels of chondrogenic markers when incubated with TGF-β3, while DSCM-expanded SDSCs displayed comparable chondrogenic potential when treated with either TGF-β isotype. In the minipig model, DSCM-expanded SDSCs were better than plastic-expanded SDSCs in enhancing collagen II and sulfated glycosaminoglycan expression in repair of partial-thickness cartilage defects, but both groups were superior to the saline control group. Our observations suggested that DSCM is a promising cell expansion system that can promote cell proliferation and enhance expanded cell chondrogenic potential in vitro and in vivo. Our approach could lead to a tissue-specific cell expansion system providing large-quantity and high-quality stem cells for the treatment of cartilage defects.
软骨缺陷的自我修复能力有限。干细胞治疗是一种很有前途的方法；然而，复制性衰老对获取大量高质量的干细胞用于软骨再生是一个挑战。滑膜来源的干细胞（SDSCs）是一种用于软骨再生的组织特异性干细胞。我们最近的研究结果表明，去细胞干细胞基质（DSCM）可以恢复在 DSCM 或塑料培养瓶上扩增后的 SDSCs 的细胞增殖和软骨生成潜能。在这项研究中，我们正在研究：（1）转化生长因子（TGF）-β1 和 TGF-β3 在 DSCM 或塑料培养瓶上扩增后的 SDSCs 软骨诱导中是否发挥相似的作用；（2）DSCM 扩增后的 SDSCs 是否在小型猪模型中修复部分厚度软骨缺陷方面具有增强的能力。SDSCs 从小猪滑膜中分离，每只 3 月龄的猪分离出 2 个滑膜，并使用 SDSCs 制备 DSCM。第 2 代 SDSCs 在 DSCM 或塑料上扩增 1 代。评估扩增细胞的细胞形态、软骨生成能力和相关机制。比较 TGF-β1 和 TGF-β3 在 DSCM 或塑料上扩增后的 SDSCs 软骨生成中的作用。不含 TGF-β 的软骨形成诱导培养基作为对照。在 13 只小型猪中，我们将 DSCM 或塑料扩增后的 SDSCs 或生理盐水关节内注射到膝关节部分厚度软骨缺陷中，并通过组织学和免疫组织化学评估其修复情况。我们发现，DSCM 扩增后的 SDSCs 体积小，呈成纤维细胞样，在三维环境中快速生长，同时磷酸环化酶 D1 和 TGF-β 受体 II 的表达上调。当用 TGF-β3 孵育时，塑料扩增后的 SDSCs 表现出更高的软骨生成标志物 mRNA 水平，而当用任何一种 TGF-β 同工型处理时，DSCM 扩增后的 SDSCs 均显示出相当的软骨生成潜能。在小型猪模型中，DSCM 扩增后的 SDSCs 在增强胶原蛋白 II 和硫酸化糖胺聚糖的表达方面优于塑料扩增后的 SDSCs 修复部分厚度软骨缺陷，但两组均优于生理盐水对照组。我们的观察结果表明，DSCM 是一种很有前途的细胞扩增系统，可促进细胞增殖并增强体外和体内扩增细胞的软骨生成潜能。我们的方法可以为软骨缺陷的治疗提供大量高质量的组织特异性细胞扩增系统。
