Hamano Sayuri, Yamashita Diaki, Hasegawa Daigaku, Sugii Hideki, Itoyama Tomohiro, Maeda Hidefumi
Department of Endodontology and Operative Dentistry, Faculty of Dental Science, Kyushu University, Fukuoka, Japan.
Department of Endodontology and Operative Dentistry, Faculty of Dental Science, Kyushu University, OBT Research Center, Faculty of Dental Science, Kyushu University, Fukuoka, Japan.
Stem Cells Dev. 2024 May;33(9-10):228-238. doi: 10.1089/scd.2024.0013. Epub 2024 Apr 13.
Periodontal tissue regeneration is important for preserving teeth. Periodontal ligament stem cells (PDLSCs) are useful in periodontal tissue regeneration; however, tooth extraction is required to obtain these cells. Therefore, we focused on induced pluripotent stem (iPS) cells and established a method to obtain PDLSC-like cells from iPS cells. Specifically, we first differentiated iPS cells into neural crest-like cells (iNCs). Next, we obtained PDLSC-like cells (iPDLSCs) by culturing iNCs on extracellular matrix (ECM) derived from human primary periodontal ligament cells (HPDLCs). This differentiation method suggested that ECM derived from HPDLCs is important for iPDLSC differentiation. Thus, we aimed to identify the PDLSC-inducing factor present in HPDLC-derived ECM in this study. We first performed comprehensive analyses of HPDLC genes and identified fibrillin-2 (), an ECM-related factor. Furthermore, to clarify the effect of FBN2 on iPDLSC differentiation, we cultured iNCs using ECM derived from HPDLCs with knocked down. As a result, expression of PDL-related markers was reduced in iNCs cultured on ECM derived from HPDLCs transfected with FBN2 siRNA (iNC-siFBN2) compared with iPDLSCs. Furthermore, the expression of CD105 (a mesenchymal stem cell marker), proliferation ability, and multipotency of iNC-siFBN2 were lower compared with iPDLSCs. Next, we cultured iNCs on FBN2 recombinant protein; however, expression of PDL-related markers did not increase compared with iPDLSC. The present results suggest the critical involvement of FBN2 in inducing iPDLSCs from iNCs when in fact it does not promote iPDLSC differantiation. Therefore, we need to elucidate the entire HPDLC-ECMs, responsible for iPDLSCs induction.
牙周组织再生对于保留牙齿至关重要。牙周膜干细胞（PDLSCs）在牙周组织再生中很有用；然而，需要拔牙才能获得这些细胞。因此，我们专注于诱导多能干细胞（iPS细胞），并建立了一种从iPS细胞中获得类PDLSC细胞的方法。具体而言，我们首先将iPS细胞分化为神经嵴样细胞（iNCs）。接下来，我们通过在源自人原代牙周膜细胞（HPDLCs）的细胞外基质（ECM）上培养iNCs来获得类PDLSC细胞（iPDLSCs）。这种分化方法表明，源自HPDLCs的ECM对iPDLSC分化很重要。因此，在本研究中，我们旨在鉴定HPDLC衍生的ECM中存在的PDLSC诱导因子。我们首先对HPDLC基因进行了全面分析，并鉴定出一种ECM相关因子原纤蛋白-2（FBN2）。此外，为了阐明FBN2对iPDLSC分化的影响，我们使用敲低FBN2的HPDLC衍生的ECM培养iNCs。结果，与iPDLSCs相比，在转染了FBN2 siRNA的HPDLC衍生的ECM（iNC-siFBN2）上培养的iNCs中，牙周相关标志物的表达降低。此外，与iPDLSCs相比，iNC-siFBN2的CD105（间充质干细胞标志物）表达、增殖能力和多能性较低。接下来，我们在FBN2重组蛋白上培养iNCs；然而，与iPDLSC相比，牙周相关标志物的表达并未增加。目前的结果表明，FBN2在从iNCs诱导iPDLSCs过程中起关键作用，而实际上它并不促进iPDLSC分化。因此，我们需要阐明负责诱导iPDLSCs的整个HPDLC-ECMs。
