Transcriptome profile of highly osteoblastic/cementoblastic periodontal ligament cell clones

Abstract Heterogeneous cell populations of osteo/cementoblastic (O/C) or fibroblastic phenotypes constitute the periodontal dental ligament (PDL). A better understanding of these PDL cell subpopulations is essential to propose regenerative approaches based on a sound biological rationale. Objective Our study aimed to clarify the differential transcriptome profile of PDL cells poised to differentiate into the O/C cell lineage. Methodology To characterize periodontal-derived cells with distinct differentiation capacities, single-cell-derived clones were isolated from adult human PDL progenitor cells and their potential to differentiate into osteo/cementoblastic (O/C) phenotype (C-O clones) or fibroblastic phenotype (C-F clones) was assessed in vitro. The transcriptome profile of the clonal cell lines in standard medium cultivation was evaluated using next-generation sequencing technology (RNA-seq). Over 230 differentially expressed genes (DEG) were identified, in which C-O clones showed a higher number of upregulated genes (193) and 42 downregulated genes. Results The upregulated genes were associated with the Cadherin and Wnt signaling pathways as well as annotated biological processes, including “anatomical structure development” and “cell adhesion.” Both transcriptome and RT-qPCR showed up-regulation of WNT2, WNT16, and WIF1 in C-O clones. Conclusions This comprehensive transcriptomic assessment of human PDL progenitor cells revealed that expression of transcripts related to the biological process “anatomical structure development,” Cadherin signaling, and Wnt signaling can identify PDL cells with a higher potential to commit to the O/C phenotype. A better understanding of these pathways and their function in O/C differentiation will help to improve protocols for periodontal regenerative therapies.

摘要：具有成骨/成牙骨质细胞（osteo/cementoblastic, O/C）或成纤维细胞表型的异质性细胞群构成了牙周膜（periodontal dental ligament, PDL）。深入解析这些牙周膜细胞亚群，对于基于可靠生物学理论提出再生治疗策略至关重要。

研究目的：本研究旨在阐明倾向于分化为成骨/成牙骨质细胞谱系的牙周膜细胞的差异转录组谱。

研究方法：为表征具有不同分化能力的牙周来源细胞，本研究从成人牙周膜祖细胞中分离出单细胞来源克隆，并在体外评估其分化为成骨/成牙骨质细胞表型（C-O克隆）或成纤维细胞表型（C-F克隆）的潜能。采用下一代测序技术（RNA-seq）对标准培养基培养的克隆细胞系的转录组谱进行分析。最终鉴定出超过230个差异表达基因（differentially expressed genes, DEG），其中C-O克隆的上调基因数量更多（193个），下调基因共42个。

研究结果：上调基因富集于钙粘蛋白（Cadherin）信号通路与Wnt信号通路，同时涉及多个注释生物学过程，包括"解剖结构发育"与"细胞黏附"。转录组测序与实时定量聚合酶链反应（RT-qPCR）结果均显示，C-O克隆中WNT2、WNT16及WIF1基因呈现上调表达。

研究结论：本研究对人类牙周膜祖细胞开展的全面转录组学分析表明，与"解剖结构发育"生物学过程、钙粘蛋白信号通路及Wnt信号通路相关的转录本表达特征，可用于识别具有更高定向分化为成骨/成牙骨质细胞表型潜能的牙周膜细胞。深入解析这些信号通路及其在成骨/成牙骨质细胞分化中的功能，将有助于优化牙周再生治疗方案。