Department of Prosthodontics and Periodontics, Piracicaba Dental School, University of Campinas, Piracicaba, São Paulo 13414-018, Brazil.
School of Dentistry, Institute of Clinical Sciences, Institute of Cancer and Genomic Sciences, University of Birmingham, Birmingham B5 7EG, UK.
Cells. 2022 Mar 26;11(7):1126. doi: 10.3390/cells11071126.
Periodontal ligament stem cells (PDLCs) can be used as a valuable source in cell therapies to regenerate bone tissue. However, the potential therapeutic outcomes are unpredictable due to PDLCs' heterogeneity regarding the capacity for osteoblast differentiation and mineral nodules production. Here, we identify epigenetic (DNA (hydroxy)methylation), chromatin (ATAC-seq) and transcriptional (RNA-seq) differences between PDLCs presenting with low (l) and high (h) osteogenic potential. The primary cell populations were investigated at basal state (cultured in DMEM) and after 10 days of osteogenic stimulation (OM). At a basal state, the expression of transcription factors (TFs) and the presence of gene regulatory regions related to osteogenesis were detected in h-PDLCs in contrast to neuronal differentiation prevalent in l-PDLCs. These differences were also observed under stimulated conditions, with genes and biological processes associated with osteoblast phenotype activated more in h-PDLCs. Importantly, even after the induction, l-PDLCs showed hypermethylation and low expression of genes related to bone development. Furthermore, the analysis of TFs motifs combined with TFs expression suggested the relevance of SP1, SP7 and DLX4 regulation in h-PDLCs, while motifs for SIX and OLIG2 TFs were uniquely enriched in l-PDLCs. Additional analysis including a second l-PDLC population indicated that the high expression of , and TFs could be predictive of low osteogenic commitment. In summary, several biological processes related to osteoblast commitment were activated in h-PDLCs from the onset, while l-PDLCs showed delay in the activation of the osteoblastic program, restricted by the persistent methylation of gene related to bone development. These processes are pre-determined by distinguishable epigenetic and transcriptional patterns, the recognition of which could help in selection of PDLCs with pre-osteoblastic phenotype.
牙周膜干细胞（PDLCs）可作为细胞治疗中再生骨组织的有价值的来源。然而，由于 PDLCs 在成骨细胞分化和矿化结节生成能力方面存在异质性，其潜在的治疗效果是不可预测的。在这里，我们鉴定了具有低（l）和高（h）成骨能力的 PDLCs 之间在表观遗传（DNA（羟）甲基化）、染色质（ATAC-seq）和转录（RNA-seq）方面的差异。在基础状态（在 DMEM 中培养）和 10 天成骨刺激（OM）后，研究了原代细胞群体。在基础状态下，与 l-PDLCs 中普遍存在的神经元分化相反，在 h-PDLCs 中检测到转录因子（TFs）的表达和与成骨相关的基因调控区域的存在。在刺激条件下也观察到这些差异，与成骨细胞表型相关的基因和生物学过程在 h-PDLCs 中更活跃。重要的是，即使在诱导后，l-PDLCs 也表现出与骨发育相关基因的高甲基化和低表达。此外，结合 TFs 表达分析 TFs 基序表明 SP1、SP7 和 DLX4 调节在 h-PDLCs 中的重要性，而 SIX 和 OLIG2 TFs 的基序仅在 l-PDLCs 中富集。包括第二个 l-PDLC 群体的额外分析表明，高表达 、 和 TFs 可能预示着低成骨能力。总之，在 h-PDLCs 中，与成骨细胞定向相关的几个生物学过程从一开始就被激活，而 l-PDLCs 显示出成骨细胞程序激活的延迟，这受到与骨发育相关基因的持续甲基化的限制。这些过程由可区分的表观遗传和转录模式预先确定，识别这些模式有助于选择具有前成骨表型的 PDLCs。
