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A 3D culture model study monitoring differentiation of dental epithelial cells into ameloblast-like cells

机译：一种3D培养模型，将牙齿上皮细胞的分化分化为含有类血管细胞样细胞

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摘要

The present study was designed to investigate whether a three dimensional (3D) culture of the rat incisor-derived dental epithelial cell line SF2 enhances its differentiation into ameloblast-like cells. SF2 cells were incubated in a laboratory-fabricated culture device to form spheroids in the presence or absence of differentiation supplements. The size of the spheroids was monitored microscopically. Differentiation was analyzed by measuring mRNA expression or the localization of amelogenin, ameloblastin, and enamel protease KLK-4 by immunofluorescence staining. The mineralization of the spheroids was monitored by Von Kossa staining and by the hardness test using a micro force sensor. The cell morphology was observed by transmission electron microscopy. Apoptosis was also assessed. The results indicated that the 3D culture allowed the SF2 cells to form homogenous spheroids within one day that exhibited a significant increase in matrix protein mRNA compared to the two dimensional culture at 2 days of culture. Ameloblastin and KLK-4 expression was detected throughout the incubation period (up to 8 days), but amelogenin expression decreased markedly, suggesting increased KLK-4 enzymatic activity. Both medium preparations increased mineralization, hardness, and apoptosis, although the differentiation medium tended to have a stronger effect. Ultrastructural analysis indicated the presence of intracellular and extracellular secretory granules, which could represent the secretion of matrix proteins. Overall our results suggest that the spheroid culture could be a useful model for analyzing ameloblast differentiation.
机译：设计本研究以研究大鼠切除衍生的牙科上皮细胞系SF2的三维（3D）培养是否增强其分化为类似于Ameloblast的细胞。将SF2细胞在实验室制造的培养装置中温育，以在存在或不存在分化补充剂中形成球状体。显微镜监测球状体的尺寸。通过免疫荧光染色测量mRNA表达或氨基脲蛋白，氨基脲蛋白酶KLK-4的mRNA表达或定位来分析分化。通过von kossa染色和使用微力传感器的硬度测试监测球状体的矿化。通过透射电子显微镜观察细胞形态。还评估了细胞凋亡。结果表明，3D培养物使SF2细胞在一天内形成均匀的球状体，其在培养2天的二维培养中表现出基质蛋白mRNA的显着增加。在整个培养期（最多8天）的情况下检测到亚利亚细胞素和KLK-4表达，但Amelogenin表达显着降低，表明KLK-4酶活性增加。虽然分化介质往往具有更强的效果，但两种中等制剂都增加了矿化，硬度和凋亡。超微结构分析表明存在细胞内和细胞外分泌颗粒，其可以代表基质蛋白的分泌。背景之利，我们的结果表明，球形培养物可以是分析Ameloblast分化的有用模型。

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  • 来源
    《RSC Advances》 |2016年第67期|共10页
  • 作者

    Tadaki Mayu; Anada Takahisa; Shiwaku Yukari; Nakamura Takashi; Nakamura Masanori; Kojima Masaru; Arai Tatsuo; Fukumoto Satoshi; Suzuki Osamu;

  • 作者单位

    Tohoku Univ Grad Sch Dent Div Pediat Dent Sendai Miyagi Japan;

    Tohoku Univ Grad Sch Dent Div Craniofacial Funct Engn Aoba Ku 4-1 Seiryo Machi Sendai Miyagi 9808575 Japan;

    Tohoku Univ Grad Sch Dent Div Craniofacial Funct Engn Aoba Ku 4-1 Seiryo Machi Sendai Miyagi 9808575 Japan;

    Tohoku Univ Grad Sch Dent Div Mol Pharmacol &

    Cell Biophys Sendai Miyagi Japan;

    Showa Univ Sch Dent Oral Anat &

    Dev Biol Tokyo Japan;

    Osaka Univ Grad Sch Engn Sci Dept Syst Innovat Toyonaka Osaka Japan;

    Osaka Univ Grad Sch Engn Sci Dept Syst Innovat Toyonaka Osaka Japan;

    Tohoku Univ Grad Sch Dent Div Pediat Dent Sendai Miyagi Japan;

    Tohoku Univ Grad Sch Dent Div Craniofacial Funct Engn Aoba Ku 4-1 Seiryo Machi Sendai Miyagi 9808575 Japan;

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