Characterisation of DNA methylation during mesenchymal stem cell chondrogenesis

Regulation of transcription occurs in a cell type specific manner orchestrated by epigenetic mechanisms including DNA methylation. Methylation changes may also play a key role in lineage specification during stem cell differentiation. To further our understanding of epigenetic regulation in chondrocytes we characterised DNA methylation changes during chondrogenesis of mesenchymal stem cells (MSCs) by Infinium 450K methylation array. Significant DNA hypomethylation was identified during chondrogenesic differentiation including changes at many key cartilage gene loci. Integration with chondrogenesis gene expression data revealed an enrichment of significant CpGs in upregulated genes, while characterisation of significant CpG loci indicated their predominant localisation to enhancer regions. Comparison with methylation profiles of other tissues, including healthy and diseased adult cartilage, identified chondrocyte-specific regions of hypomethylation and the overlap with differentially methylated CpGs in osteoarthritis. Taken together we have associated DNA methylation levels with the chondrocyte phenotype. The consequences of which has potential to improve cartilage generation for tissue engineering purposes and also to provide context for observed methylation changes in cartilage diseases such as osteoarthritis Bisulphite converted DNA from the 12 samples were hybridised to the Illumina Infinium HumanMethylation450 BeadChip array.

转录调控以细胞类型特异性的方式进行，由包括DNA甲基化在内的表观遗传机制统筹调控。甲基化修饰变化在干细胞分化过程中的谱系特化阶段亦可发挥关键作用。为深化对软骨细胞表观遗传调控机制的认知，本研究采用Infinium 450K甲基化芯片（Infinium 450K methylation array），对间充质干细胞（Mesenchymal Stem Cells, MSCs）软骨分化过程中的DNA甲基化变化开展表征分析。研究在软骨分化阶段检测到显著的DNA低甲基化现象，涉及诸多关键软骨相关基因位点的甲基化改变。整合软骨分化基因表达数据进行分析后发现，显著差异的CpG位点在上调基因中显著富集；对显著差异CpG位点的定位分析显示，其主要富集于增强子区域。将本研究的甲基化谱与其他组织（包括健康及病变成人软骨）的甲基化谱进行比对后，鉴定出软骨细胞特异性的低甲基化区域，同时发现该区域与骨关节炎患者中存在差异甲基化的CpG位点存在重叠。综上，本研究将DNA甲基化水平与软骨细胞表型建立了关联。该研究成果既可用于优化组织工程领域的软骨生成技术，也可为软骨疾病（如骨关节炎）中观测到的甲基化变化提供理论依据。本研究将12份样本的亚硫酸氢盐转化DNA与Illumina Infinium HumanMethylation450 BeadChip芯片进行杂交实验。