Formation of Bovine Placental Trophoblast Spheroids
Formation of Bovine Placental Trophoblast Spheroids
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DOI：
10.1159/000320544
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发表时间：
2011-01-01
影响因子：
2.7
通讯作者：
Pfarrer, C.
中科院分区：
文献类型：
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作者：
Haeger, J-D;Hambruch, N.;Pfarrer, C.
引言：在这项研究中，我们的目的是形成球体与牛胎盘滋养层细胞系F3。球体是三维培养模型，可用于进行多功能的体外和体内实验。材料和方法：采用悬滴法，25%甲基纤维素和基质胶制备球形体。通过光学显微镜和透射（TEM）和扫描电子显微镜（SEM）以及免疫组织化学（埃兹蛋白、波形蛋白、细胞角蛋白、胎盘催乳素）对F3球体进行形态学表征。用荧光染料钙黄绿素和乙锭同型二聚体通过免疫荧光显微镜测定球形F3细胞的活力。结果：加入基质胶后，F3细胞悬滴法只能形成球状体。滋养层球体被细胞外基质界定并完全覆盖（光学显微镜/TEM/SEM）。形成球状体的细胞不能相互区分（光学显微镜）。外球形层由具有顶极的细胞组成，顶极具有指向外部的微绒毛（光学显微镜/TEM）。所有的球形F3细胞表达ezrin，波形蛋白和细胞角蛋白，但不胎盘催乳素。球状体核心含有变性细胞，而外缘的F3细胞是活的（TEM/免疫荧光显微镜）。讨论：我们建立了牛胎盘滋养层细胞系F3的三维球体模型。该培养模型对牛滋养层细胞的体外分化研究具有一定的参考价值。版权所有（C）2010 S. Karger AG，巴塞尔
Introduction: In this study, we aimed to form spheroids with the bovine placental trophoblast cell line F3. Spheroids are 3-dimensional culture models which can be used to conduct versatile in vitro and in vivo experiments. Materials and Methods:The spheroids were generated using the hanging drop technique, 25% methocel and matrigel. The F3 spheroids were characterized morphologically by light microscopy and transmission (TEM) and scanning electron microscopy (SEM) and immunohistochemistry (ezrin, vimentin, cytokeratin, placental lactogen). The fluorescent dyes calcein and ethidium homodimer were used to determine the viability of the spheroidal F3 cells by immunofluorescence microscopy. Results: The cell line F3 only formed spheroids by the hanging drop technique when matrigel was added. The trophoblast spheroids were delimited and fully covered by extracellular matrix (light microscopy/TEM/SEM). Cells contributing to spheroids could not be discriminated from each other (light microscopy). The outer spheroidal layer consisted of cells which possessed an apical pole with microvilli that were directed to the outside (light nnicroscopy/TEM). All of the spheroidal F3 cells expressed ezrin, vimentin and cytokeratin, but not placental lactogen. The spheroid core contained degenerating cells whilst the F3 cells of the outer rim were viable (TEM/immunofluorescence microscopy). Discussion: We have established a 3-dimensional spheroid model for the bovine placental trophoblast cell line F3. The developed culture model might prove valuable for future in vitro studies on the differentiation of bovine trophoblast cells. Copyright (C) 2010 S. Karger AG, Basel