Fetal Mesenchymal Stromal Cells Differentiating towards Chondrocytes Acquire a Gene Expression Profile Resembling Human Growth Plate Cartilage

We used human fetal bone marrow-derived mesenchymal stromal cells (hfMSCs) differentiating towards chondrocytes as an alternative model for the human growth plate (GP). Our aims were to study gene expression patterns associated with chondrogenic differentiation to assess whether chondrocytes derived from hfMSCs are a suitable model for studying the development and maturation of the GP. hfMSCs efficiently formed hyaline cartilage in a pellet culture in the presence of TGFβ3 and BMP6. Microarray and principal component analysis were applied to study gene expression profiles during chondrogenic differentiation. A set of 232 genes was found to correlate with in vitro cartilage formation. Several identified genes are known to be involved in cartilage formation and validate the robustness of the differentiating hfMSC model. KEGG pathway analysis using the 232 genes revealed 9 significant signaling pathways correlated with cartilage formation. To determine the progression of growth plate cartilage formation, we compared the gene expression profile of differentiating hfMSCs with previously established expression profiles of epiphyseal GP cartilage. As differentiation towards chondrocytes proceeds, hfMSCs gradually obtain a gene expression profile resembling epiphyseal GP cartilage. We visualized the differences in gene expression profiles as protein interaction clusters and identified many protein clusters that are activated during the early chondrogenic differentiation of hfMSCs showing the potential of this system to study GP development.

本研究以向软骨细胞分化的人胎儿骨髓来源间充质基质细胞（human fetal bone marrow-derived mesenchymal stromal cells，hfMSCs）作为人类生长板（growth plate，GP）的替代模型，旨在探究与软骨分化过程相关的基因表达模式，以评估由hfMSCs分化得到的软骨细胞是否可作为研究GP发育与成熟的适宜模型。在转化生长因子β3（TGFβ3）与骨形态发生蛋白6（BMP6）存在的条件下，hfMSCs可在团块培养中高效形成透明软骨。本研究采用基因芯片（microarray）与主成分分析（principal component analysis）分析软骨分化过程中的基因表达谱，发现共有232个基因与体外软骨形成过程相关；其中多个已鉴定基因已被证实参与软骨形成，验证了该分化中hfMSCs模型的稳健性。针对这232个基因开展的KEGG通路分析（KEGG pathway analysis）显示，共有9条显著信号通路与软骨形成过程相关。为明确生长板软骨形成的进程，本研究将分化中的hfMSCs的基因表达谱与已发表的骨骺GP软骨表达谱进行比对，结果显示随着软骨分化进程推进，hfMSCs的基因表达谱逐渐趋近于骨骺GP软骨的表达谱。本研究将基因表达谱的差异以蛋白质相互作用簇的形式进行可视化，鉴定出多个在hfMSCs早期软骨分化过程中被激活的蛋白质簇，证实了该系统用于研究GP发育的潜力。