Laboratory of Molecular Genetics, Dental Research Institute, School of Dentistry, Seoul National University, Seoul, Korea.
Stem Cells Dev. 2024 Apr;33(7-8):189-199. doi: 10.1089/scd.2023.0220. Epub 2024 Mar 21.
Research on tooth regeneration using human-induced pluripotent stem cells (hiPSCs) is valuable for autologous dental regeneration. Acquiring mesenchymal and epithelial cells as a resource for dental regeneration is necessary because mesenchymal-epithelial interactions play an essential role in dental development. We reported the establishment of hiPSCs-derived dental epithelial-like cell (EPI-iPSCs), but hiPSCs-derived dental mesenchymal stem cells (MSCs) have not yet been reported. This study was conducted to establish hiPSCs-derived MSCs and to differentiate them into dental cells with EPI-iPSCs. Considering that dental MSCs are derived from the neural crest, hiPSCs were induced to differentiate into MSCs through neural crest formation to acquire the properties of dental MSCs. To differentiate hiPSCs into MSCs through neural crest formation, established hiPSCs were cultured and differentiated with PA6 stromal cells and differentiated hiPSCs formed neurospheres on ultralow-attachment plates. Neurospheres were differentiated into MSCs in serum-supplemented medium. Neural crest-mediated MSCs (NC-MSCs) continuously showed typical MSC morphology and expressed MSC markers. After 8 days of odontogenic induction, the expression levels of odontogenic/mineralization-related genes and dentin sialophosphoprotein (DSPP) proteins were increased in the NC-MSCs alone group in the absence of coculturing with dental epithelial cells. The NC-MSCs and EPI-iPSCs coculture groups showed high expression levels of amelogenesis/odontogenic/mineralization-related genes and DSPP proteins. Furthermore, the NC-MSCs and EPI-iPSCs coculture group yielded calcium deposits earlier than the NC-MSCs alone group. These results indicated that established NC-MSCs from hiPSCs have dental differentiation capacity with dental epithelial cells. In addition, it was confirmed that hiPSCs-derived dental stem cells could be a novel cell source for autologous dental regeneration.
利用人诱导多能干细胞（hiPSCs）进行牙齿再生的研究对于自体牙再生是有价值的。获得间充质和上皮细胞作为牙再生的资源是必要的，因为间充质-上皮相互作用在牙齿发育中起着至关重要的作用。我们已经报道了 hiPSCs 衍生的牙上皮样细胞（EPI-iPSCs）的建立，但尚未报道 hiPSCs 衍生的牙间充质干细胞（MSCs）。本研究旨在建立 hiPSCs 衍生的 MSCs，并将其分化为与 EPI-iPSCs 共培养的牙细胞。考虑到牙 MSCs 来源于神经嵴，我们通过诱导 hiPSCs 分化为 MSCs 来获得牙 MSCs 的特性。为了通过神经嵴形成将 hiPSCs 分化为 MSCs，我们用 PA6 基质细胞培养和分化建立的 hiPSCs，并在超低附着平板上分化 hiPSCs 形成神经球。在含血清的培养基中，神经球分化为 MSCs。神经嵴介导的 MSCs（NC-MSCs）持续表现出典型的 MSC 形态，并表达 MSC 标志物。在牙向诱导 8 天后，在没有与牙上皮细胞共培养的情况下，NC-MSCs 单独组的牙向/矿化相关基因和牙本质涎磷蛋白（DSPP）蛋白的表达水平增加。NC-MSCs 和 EPI-iPSCs 共培养组表现出高表达的成釉/牙向/矿化相关基因和 DSPP 蛋白。此外，NC-MSCs 和 EPI-iPSCs 共培养组比 NC-MSCs 单独组更早产生钙沉积物。这些结果表明，从 hiPSCs 建立的 NC-MSCs 具有与牙上皮细胞共培养的牙向分化能力。此外，证实了 hiPSCs 衍生的牙干细胞可以作为自体牙再生的新型细胞来源。
