Basement membrane assembly and differentiation of cultured corneal cells: importance of culture environment and endothelial cell interaction.
Basement membrane assembly and differentiation of cultured corneal cells: importance of culture environment and endothelial cell interaction.
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DOI：
10.1006/excr.1994.1300
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发表时间：
1994-10
影响因子：
3.7
通讯作者：
J. Zieske;Valerie S. Mason;M. Wasson;Susan F. Meunier;C. Nolte;N. Fukai;Björn Olsen;N. Parenteau
中科院分区：
文献类型：
--
作者：
J. Zieske;Valerie S. Mason;M. Wasson;Susan F. Meunier;C. Nolte;N. Fukai;Björn Olsen;N. Parenteau
采用三维角膜组织结构检测培养环境和内皮细胞相互作用对上皮细胞分化和基底膜组装的影响。兔角膜上皮细胞在兔基质成纤维细胞上进行培养，在胶原基质中添加或不添加永生化的小鼠角膜内皮细胞(村崎、柴田、井上、大岛、奥尔森和尼诺米亚)。(1992)欧元。J.生物化学。207、895-902)。培养物是在水下或在干燥或潮湿的界面上生长的。使用透射式电子显微镜和间接免疫荧光显微镜观察基底膜、锚定纤维和半桥粒组装的层粘蛋白、II型胶原和α6整合素。角蛋白3(K3)和α-烯醇化酶抗体分别标记分化和未分化的角膜上皮细胞。当三种细胞都在潮湿的界面上培养时，在将培养物提升到气液界面(气升式)2wk后，观察到半桥粒、锚定纤维和连续的基底膜。阿尔法烯醇化酶和K3的分布与角膜边缘区域的模式相同。没有内皮细胞层的空运组织结构仅显示层粘连蛋白和III型胶原在上皮-基质交界处的分布有限。α-6整合素分布于基底层细胞的整个质膜上，上皮分化不完全，在基底层和2~3层超基底层细胞中可见α-烯醇化酶。无内皮细胞的液体培养细胞不表达分化标志物或基底膜成分。这些数据表明，内皮细胞的相互作用显著提高了上皮基底膜组装的数量和质量，并且上皮分化受到组织、液体和空气之间的界面类型的影响。
A three-dimensional corneal tissue construct was used to examine the effect of culture environment and endothelial cell interaction on epithelial differentiation and basement membrane assembly. Rabbit corneal epithelial cells were cultured over rabbit stromal fibroblasts in a collagen matrix with or without an underlying layer of immortalized mouse corneal endothelial cells (Muragaki, Shiota, Inoue, Ooshima, Olsen, and Ninomiya. (1992) Eur. J. Biochem. 207, 895-902). The cultures were grown submerged or at a dry or moist interface. Basement membrane, anchoring fibril, and hemidesmosome assembly was monitored using transmission electron microscopy as well as indirect immunofluorescence microscopy of laminin, type VII collagen, and alpha 6 integrin. Antibodies against keratin 3 (K3) and alpha-enolase marked differentiated and undifferentiated corneal epithelial cells, respectively. When all three cell types were cultured at a moist interface, hemidesmosomes, anchoring fibrils, and a continuous basement membrane were observed 2 wk after lifting the cultures to an air-liquid interface (air-lift). The distribution of alpha-enolase and K3 was identical to patterns seen in the limbal region of the cornea. Air-lifted tissue constructs lacking the endothelial cell layer showed only limited distribution of laminin and type VII collagen at the epithelial-matrix junction. alpha 6 Integrin was present along the entire plasma membrane of the basal cells; epithelial differentiation was not complete as alpha-enolase was seen in basal and two to three layers of suprabasal cells. Submerged cultures without endothelial cells did not express differentiation markers or basement membrane components. These data indicate that endothelial cell interaction dramatically enhances the amount and quality of epithelial basement membrane assembly and that epithelial differentiation is influenced by the type of interface between tissue, liquid, and air.