Department of Endodontics, Beijing Stomatological Hospital, School of Stomatology, Capital Medical University , Beijing, China.
Beijing Key Laboratory of Tooth Regeneration and Function Reconstruction, Beijing Stomatological Hospital, School of Stomatology, Capital Medical University , Beijing, China.
Connect Tissue Res. 2020 Sep;61(5):498-508. doi: 10.1080/03008207.2019.1620224. Epub 2019 May 28.
Periodontal ligament mesenchymal stem cells (PDLSCs) are important for periodontal tissue regeneration, but how these cells are regulated remains unclear. PRDM (PRDI-BF1 and RIZ homology domain containing) genes play key roles in cell proliferation and differentiation. The present study aimed to investigate the role of one PRDM gene, PRDM9, in the proliferation, migration and chemotaxis potential of PDLSCs.
Cell proliferation was examined on the basis of the cell doubling time, cell counting kit-8 (CCK8) assays, and flow cytometry analysis of the cell cycle. Gene expression was detected by Western blotting and real-time RT-PCR. Scratch migration and Transwell chemotaxis assays were used to analyse cell migration and chemotaxis abilities. Microarray analysis and ChIP assays were used to examine the downstream genes of PRDM9 and the corresponding mechanism.
The results showed that knock-down of PRDM9 enhanced cell proliferation by promoting cell cycle progression and rapid transition from the G1 to S phase via downregulation of p21 and p27 and upregulation of cyclin E. Additionally, depletion of PRDM9 increased the migration and chemotaxis potential of PDLSCs. Microarray results showed that 13 genes, including , and , were upregulated and 34 genes, including PIP, were downregulated after the depletion of PRDM9. Furthermore, we observed that the depletion of PRDM9 promoted the transcription of IGFBP5 by increasing H3K4me3 methylation in the IGFBP5 promoter.
These discoveries indicated that depletion of PRDM9 increased the cell proliferation, migration and chemotaxis potential of PDLSCs and revealed important downstream genes.
牙周膜间充质干细胞（PDLSCs）对牙周组织再生很重要，但这些细胞是如何被调控的尚不清楚。PRDM（PRDI-BF1 和 RIZ 同源结构域）基因在细胞增殖和分化中起关键作用。本研究旨在探讨 PRDM 基因 PRDM9 对 PDLSCs 增殖、迁移和趋化潜能的作用。
根据细胞倍增时间、细胞计数试剂盒-8（CCK8）检测和细胞周期流式细胞术分析，检测细胞增殖。通过 Western blot 和实时 RT-PCR 检测基因表达。划痕迁移和 Transwell 趋化实验分析细胞迁移和趋化能力。微阵列分析和 ChIP 实验用于检测 PRDM9 的下游基因及其相应机制。
结果表明，敲低 PRDM9 通过下调 p21 和 p27 以及上调细胞周期蛋白 E 促进细胞周期进程和快速从 G1 期向 S 期转变，从而增强细胞增殖。此外，PRDM9 的耗竭增加了 PDLSCs 的迁移和趋化潜能。微阵列结果显示，PRDM9 耗竭后，包括 、 和 在内的 13 个基因上调，而包括 PIP 在内的 34 个基因下调。此外，我们观察到 PRDM9 的耗竭通过增加 IGFBP5 启动子中的 H3K4me3 甲基化来促进 IGFBP5 的转录。
这些发现表明，PRDM9 的耗竭增加了 PDLSCs 的细胞增殖、迁移和趋化潜能，并揭示了重要的下游基因。
