5-Aza-2′-deoxycytidine treatment induces skeletal myogenic differentiation of mouse dental pulp stem cells
5-Aza-2′-deoxycytidine treatment induces skeletal myogenic differentiation of mouse dental pulp stem cells
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DOI：
10.1016/j.archoralbio.2010.03.003
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发表时间：
2010-05-01
影响因子：
3
通讯作者：
Sonoda, Yoshiaki
中科院分区：
文献类型：
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作者：
Nakatsuka, Ryusuke;Nozaki, Tadashige;Sonoda, Yoshiaki
目的：牙髓组织干细胞被认为具有与间充质干细胞（MSCs）相似的分化潜能。本体外研究的目的是检测小鼠牙髓干细胞（DPSCs）的分化潜力，并开发合适的分化试验系统，用于这些细胞的骨骼肌分化。方法：从G57/BL6小鼠下颌骨切片中提取牙髓，培养分离贴壁牙髓细胞。这些细胞在成骨或成脂诱导培养基中培养，诱导成骨和成脂分化。另一方面，研究了这些细胞在不同条件下的分化潜力，如无血清培养基、Myod1过表达或5-Aza-2'-脱氧胞苷（5-Aza）处理DNA去甲基化。采用RT-PCR检测肌肉特异性转录因子表达，采用相差显微镜和免疫荧光染色分别检测肌管形成和肌球蛋白重链表达。结果：黏附牙髓细胞表现出增殖能力，并表现出成骨和成脂分化。虽然在无血清条件下未检测到Myod1 mRNA的表达和肌管的形成，但Myod1的强制表达上调了Myogenin和Pax7 mRNA的表达。然而，肌管形成尚未得到证实。有趣的是，在5-Aza处理这些细胞后，观察到肌球蛋白重链表达和肌管形成。结论：小鼠DPSCs具有msc样分化潜能。5-Aza处理诱导的DNA去甲基化导致小鼠DPSCs骨骼肌分化，提示DNA去甲基化可能触发小鼠DPSCs的这种差异诱导。(C) 2010 Elsevier Ltd.版权所有。
Objective: Tissue stem cells in dental pulp are assumed to possess differentiation potentials similar to mesenchymal stem cells (MSCs). The aim of this in vitro study is to examine the differentiation potentials of mouse dental pulp stem cells (DPSCs) and develop the appropriate differentiation assay systems for skeletal myogenic differentiation of these cells.Methods: Dental pulps were extracted from mandible sections of G57/BL6 mice, and adherent dental pulp cells were isolated in culture. These cells were cultured in osteogenic or adipogenic induction medium to induce osteogenic and adipogenic differentiation. On the other hand, the skeletal myogenic differentiation potential of these cells was investigated using different conditions, such as serum-free medium, Myod1 overexpression, or 5-Aza-2'-deoxycytidine (5-Aza) treatment for DNA demethylation. Muscle-specific transcriptional factor expression was evaluated by RT-PCR, and myotube formation and myosin heavy chain expression were evaluated by phase-contrast microscopy and immunofluorescence staining, respectively.Results: The adherent dental pulp cells exhibited a proliferative capacity and they showed osteogenic and adipogenic differentiation as seen in previous studies. Although the expression of Myod1 mRNA and myotube formation was not detected in serum-free conditions, the forced expression of Myod1 up-regulated the expression of Myogenin and Pax7 mRNA. However, myotube formation was not confirmed. Interestingly, myosin heavy chain expression and myotube formation were observed following 5-Aza treatment of these cells.Conclusions: These results demonstrated that mouse DPSCs possess MSC-like differentiation potential. DNA demethylation induced by 5-Aza treatment resulted in the skeletal muscle differentiation in mouse DPSCs, suggesting that DNA demethylation might trigger this differential induction of mouse DPSCs. (C) 2010 Elsevier Ltd. All rights reserved.