Immortalization and characterization of human dental mesenchymal cells
Immortalization and characterization of human dental mesenchymal cells
复制标题
人牙间充质细胞的永生化和表征。
DOI：
10.1016/j.jdent.2015.02.008
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发表时间：
2015-05-01
影响因子：
4.4
通讯作者：
Lin, Yao
中科院分区：
文献类型：
--
作者：
Huang, Yide;Yang, Yun;Lin, Yao
目的：由于人胚胎样本的稀有性和原代人牙间充质细胞增殖能力有限，建立永生化人牙间充质细胞系对于研究人牙本质形成过程中牙间充质细胞的分化和信号通路具有重要价值。方法：本研究从妊娠19周的人磨牙牙胚中分离出牙间充质细胞，并用pSV40进行永生化。然后通过 96 孔板稀释选择单细胞集落。使用免疫荧光显微镜、RT-PCR 和蛋白质印迹对永生化细胞系进行表征，以表达 SV40 大 T 抗原和牙齿发育过程中间充质阶段特异的 5 个基因。使用脂肪形成和钙化诱导比较永生化细胞和原代细胞的分化和矿化活性。结果：永生化牙齿间充质细胞系表现出更高的增殖率，表达多种牙齿特异性标记物，包括Msx1、Pax9、Lhx6、Barx1和Runx2，并保持分化和形成矿化结节的能力。结论：我们的结果表明，永生化人间充质细胞系保留了与原代细胞相似的特征。人牙间充质细胞，可用于研究人牙间充质细胞分化的机制和参与人牙发育的信号通路。 (C) 2015 Elsevier Ltd. 保留所有权利。
Objectives: Due to the rarity of human embryonic samples and limited proliferating capability of primary human dental mesenchymal cells, it is valuable to create an immortalized human dental mesenchymal cell line for studying dental mesenchymal cell differentiation and signalling pathways during detinogenesis in humans.Methods: In this study, dental mesenchymal cells from human molar tooth germs at 19-week gestation were isolated and immortalized with pSV40. Single cell colonies were then selected by 96-well plate dilution. The immortalized cell line was characterized using immmunofluorescent microscopy, RT-PCR and Western blot for the expression of SV40 large T antigen and five genes specific for the mesenchymal stage during tooth development. The differentiation and mineralization activities of the immortalized and primary cells were compared using adipogenic and calcifying induction.Results: The immortalized dental mesenchymal cell line displayed a higher proliferation rate, expressed several tooth-specific markers including Msx1, Pax9, Lhx6, Barx1, and Runx2, and maintained the ability to differentiate and form mineralized nodules.Conclusions: Our results demonstrated that the immortalized human mesenchymal cell line retained the characteristics similar to primary human dental mesenchymal cells and can be used for studying the mechanisms of human dental mesenchymal cell differentiation and signalling pathways involved in human odontogenesis. (C) 2015 Elsevier Ltd. All rights reserved.