Department of Medical Molecular Biology, University of Lübeck, Lübeck, Germany.
Cytotechnology. 2003 Mar;41(2-3):177-87. doi: 10.1023/A:1024835025011.
Our knowledge of cellular differentiation processes during chondro- and osteogenesis, in particular the complex interaction of differentiation factors, is still limited. We used the model system of embryonic stem (ES) cell differentiation in vitro via cellular aggregates, so called embryoid bodies (EBs), to analyze chondrogenic and osteogenic differentiation. ES cells differentiated into chondrocytes and osteocytes throughout a series of developmental stages resembling cellular differentiation events during skeletal development in vivo. A lineage from pluripotent ES cells via mesenchymal, prechondrogenic cells, chondrocytes and hypertrophicchondrocytes up to osteogenic cells was characterized. Furthermore, we found evidence for another osteogenic lineage, bypassing the chondrogenic stage. Together our results suggest that this in vitro system will be helpful to answer so far unacknowledged questions regarding chondrogenic and osteogenic differentiation. For example, we isolated an as yet unknown cDNA fragment from ES cell-derived chondrocytes, which showed a developmentally regulated expression pattern during EB differentiation. Considering ES cell differentiation as an alternative approach for cellular therapy, we used two different methods to obtain pure chondrocyte cultures from the heterogenous EBs. First, members of the transforming growth factor (TGF)-beta family were applied and found to modulate chondrogenic differentiation but were not effective enough to produce sufficient amounts of chondrocytes. Second, chondrocytes were isolated from EBs by micro-manipulation. These cells initially showed dedifferentiation into fiboblastoid cells in culture, but later redifferentiated into mature chondrocytes. However, a small amount of chondrocytes isolated from EBs transdifferentiated into other mesenchymal cell types, indicating that chondrocytes derived from ES cells posses a distinct differentiation plasticity.
我们对于软骨和成骨细胞分化过程中的细胞分化知识，特别是分化因子的复杂相互作用，仍然有限。我们使用胚胎干细胞（ES）细胞在体外通过细胞聚集体（即胚状体（EBs））分化的模型系统来分析软骨和成骨分化。ES 细胞分化为软骨细胞和骨细胞，经历了一系列类似于体内骨骼发育过程中细胞分化事件的发育阶段。从多能 ES 细胞到间充质细胞、前软骨细胞、软骨细胞和肥大软骨细胞，再到成骨细胞的谱系得到了描述。此外，我们发现了另一种成骨谱系的证据，绕过了软骨生成阶段。我们的研究结果表明，这种体外系统将有助于回答迄今为止尚未得到承认的关于软骨和成骨分化的问题。例如，我们从 ES 细胞来源的软骨细胞中分离出一个未知的 cDNA 片段，该片段在 EB 分化过程中表现出发育调节表达模式。考虑到 ES 细胞分化是细胞治疗的替代方法，我们使用两种不同的方法从异质 EBs 中获得纯软骨细胞培养物。首先，应用转化生长因子（TGF）-β家族成员发现其调节软骨分化，但效果不够，不能产生足够数量的软骨细胞。其次，通过微操作从 EBs 中分离出软骨细胞。这些细胞在培养中最初表现出向成纤维细胞样细胞的去分化，但后来重新分化为成熟的软骨细胞。然而，从 EBs 分离的少量软骨细胞转分化为其他间充质细胞类型，表明 ES 细胞来源的软骨细胞具有独特的分化可塑性。
