Department of Cariology, Restorative Sciences and Endodontics, University of Michigan School of Dentistry, Ann Arbor, MI, USA.
Department of Biomaterials Science, Osaka University Graduate School of Dentistry, Suita City, Osaka, Japan.
J Dent Res. 2020 Apr;99(4):437-445. doi: 10.1177/0022034520902458. Epub 2020 Feb 6.
It is known that dental pulp stem cells (DPSCs) can be induced to differentiate into vasculogenic endothelial (VE) cells. However, the process that results in sprouting and anastomosis of DPSC-derived vessels remains unclear. Here, we performed studies to understand the mechanisms underpinning the anastomosis of the host vasculature with blood vessels generated by DPSCs (a model for mesenchymal stem cells). VE-cadherin-silenced primary human DPSCs seeded in tooth slice/scaffolds and transplanted into the subcutaneous space of immunodeficient mice generated fewer functional blood vessels (i.e., anastomosed with the host vasculature) than control DPSCs transduced with scrambled sequences. Both VE-cadherin-silenced and mitogen-activated protein kinase kinase 1 (MEK1)-silenced cells showed a decrease in the number of capillary sprouts in vitro. Interestingly, DPSC stably transduced with a VE-cadherin reporter demonstrated that vascular endothelial growth factor (VEGF) induces VE-cadherin expression in sprouting DPSCs undergoing anastomosis, but not in quiescent DPSCs. To begin to understand the mechanisms regulating VE-cadherin, we stably silenced MEK1 and observed that VEGF was no longer able to induce VE-cadherin expression and capillary sprout formation. Notably ERG, a transcriptional factor downstream from MEK/ERK, binds to the promoter region of VE-cadherin (chip assay) and is induced by VEGF in DPSCs. Collectively, these data defined a signaling pathway triggered by VEGF that results in phosphorylation of MEK1/ERK and activation of ERG leading to expression of VE-cadherin, which is required for anastomosis of DPSC-derived blood vessels. In conclusion, these results unveiled a signaling pathway that enables the generation of functional blood vessels upon vasculogenic differentiation of DPSCs.
已知牙髓干细胞（DPSCs）可被诱导分化为血管生成内皮（VE）细胞。然而，导致 DPSCs 衍生血管发芽和吻合的过程尚不清楚。在这里，我们进行了研究，以了解支持 DPSCs 衍生血管（间充质干细胞模型）与宿主血管吻合的机制。与转导乱序序列的对照 DPSCs 相比，接种在牙切片/支架中并移植到免疫缺陷小鼠皮下空间的 VE-钙粘蛋白沉默的原代人 DPSCs 产生的功能性血管（即与宿主血管吻合）较少。沉默 VE-钙粘蛋白和丝裂原活化蛋白激酶激酶 1（MEK1）的细胞在体外均显示出毛细血管芽数目的减少。有趣的是，稳定转导 VE-钙粘蛋白报告基因的 DPSC 表明血管内皮生长因子（VEGF）诱导正在吻合的发芽 DPSCs 中 VE-钙粘蛋白的表达，但不诱导静止的 DPSCs。为了开始了解调节 VE-钙粘蛋白的机制，我们稳定沉默了 MEK1，并观察到 VEGF 不再能够诱导 VE-钙粘蛋白表达和毛细血管芽形成。值得注意的是，MEK/ERK 下游的转录因子 ERG 与 VE-钙粘蛋白的启动子区域结合（芯片分析），并在 DPSCs 中被 VEGF 诱导。总之，这些数据定义了一个由 VEGF 触发的信号通路，导致 MEK1/ERK 的磷酸化和 ERG 的激活，从而导致 VE-钙粘蛋白的表达，这是 DPSCs 衍生血管吻合所必需的。总之，这些结果揭示了一个信号通路，该通路使 DPSCs 血管生成分化后能够产生功能性血管。
