Identification and Clonal Characterisation of a Progenitor Cell Sub-Population in Normal Human Articular Cartilage

BackgroundArticular cartilage displays a poor repair capacity. The aim of cell-based therapies for cartilage defects is to repair damaged joint surfaces with a functional replacement tissue. Currently, chondrocytes removed from a healthy region of the cartilage are used but they are unable to retain their phenotype in expanded culture. The resulting repair tissue is fibrocartilaginous rather than hyaline, potentially compromising long-term repair. Mesenchymal stem cells, particularly bone marrow stromal cells (BMSC), are of interest for cartilage repair due to their inherent replicative potential. However, chondrocyte differentiated BMSCs display an endochondral phenotype, that is, can terminally differentiate and form a calcified matrix, leading to failure in long-term defect repair. Here, we investigate the isolation and characterisation of a human cartilage progenitor population that is resident within permanent adult articular cartilage.
Methods and FindingsHuman articular cartilage samples were digested and clonal populations isolated using a differential adhesion assay to fibronectin. Clonal cell lines were expanded in growth media to high population doublings and karyotype analysis performed. We present data to show that this cell population demonstrates a restricted differential potential during chondrogenic induction in a 3D pellet culture system. Furthermore, evidence of high telomerase activity and maintenance of telomere length, characteristic of a mesenchymal stem cell population, were observed in this clonal cell population. Lastly, as proof of principle, we carried out a pilot repair study in a goat in vivo model demonstrating the ability of goat cartilage progenitors to form a cartilage-like repair tissue in a chondral defect.
ConclusionsIn conclusion, we propose that we have identified and characterised a novel cartilage progenitor population resident in human articular cartilage which will greatly benefit future cell-based cartilage repair therapies due to its ability to maintain chondrogenicity upon extensive expansion unlike full-depth chondrocytes that lose this ability at only seven population doublings.

【背景】关节软骨（Articular cartilage）修复能力极为有限。针对软骨缺损的细胞治疗策略，其核心目标是借助具备生理功能的替代组织修复受损的关节面。当前临床常用的治疗方案是从健康软骨区域分离获取软骨细胞（chondrocytes），但这类细胞在扩增培养过程中无法维持其固有表型（phenotype），最终形成的修复组织为纤维软骨而非透明软骨（hyaline cartilage），可能对长期修复效果造成不利影响。间充质干细胞（Mesenchymal stem cells），尤其是骨髓基质细胞（bone marrow stromal cells, BMSC），因其天然具备的复制潜能，成为软骨修复领域的研究热点。然而，经软骨细胞诱导分化的BMSC会呈现软骨内骨化表型（endochondral phenotype），即能够发生终末分化并形成钙化基质，最终导致软骨缺损的长期修复失败。本研究聚焦于分离并鉴定定位于成熟成人关节软骨内的人类软骨祖细胞群（cartilage progenitor population）。

【方法与结果】我们对人类关节软骨样本进行酶消化处理，并通过针对纤连蛋白（fibronectin）的差异黏附实验分离得到克隆细胞群。将克隆细胞系在生长培养基中扩增至较高的群体倍增（population doublings）次数，并开展核型分析（karyotype analysis）。本研究的数据表明，该细胞群在三维微团培养系统（3D pellet culture system）中进行软骨诱导分化时，表现出受限的分化潜能。此外，该克隆细胞群还表现出高端粒酶（telomerase）活性及端粒（telomere）长度维持能力，这正是间充质干细胞群的典型特性。最后，作为原理验证实验，我们在山羊体内模型中开展了初步修复研究，结果证实山羊软骨祖细胞能够在软骨缺损部位形成类似正常软骨的修复组织。

【结论】综上，我们成功鉴定并表征了一种定位于人类关节软骨内的新型软骨祖细胞群。与仅经历7次群体倍增便丧失成软骨能力的全层软骨细胞不同，该细胞群在大规模扩增后仍可维持其成软骨特性，因此将极大推动未来基于细胞的软骨修复治疗研究的发展。