Self-organized intestinal epithelial monolayers in crypt and villus-like domains show effective barrier function
Self-organized intestinal epithelial monolayers in crypt and villus-like domains show effective barrier function
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DOI：
10.1038/s41598-019-46497-x
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发表时间：
2019-07-12
影响因子：
4.6
通讯作者：
Martinez, Elena
中科院分区：
文献类型：
--
作者：
Altay, Gizem;Larranaga, Enara;Martinez, Elena
由于肠类器官在结构和功能水平上与体内组织相似，因此已成为研究肠道生物学的强大体外工具。然而，它们的球形几何形状阻止了进入上皮的顶端侧，使得它们不适合为扁平细胞单层设计的标准功能测定。在这里，我们描述了一种形成上皮单层的简单方法，该方法概括了体内样细胞类型的组成和组织，并且适用于功能性组织屏障测定。在我们的方法中，上皮单层扩散是由基质硬度驱动的，而组织屏障功能是通过基底外侧输送富含干细胞生态位和肌成纤维细胞衍生因子的培养基来实现的。这些单层含有主要的肠上皮细胞类型，组织成增殖的隐窝样结构域和分化的绒毛样区域，与体内细胞分布非常相似。作为一个独特的特征，这些上皮单层形成功能性上皮屏障，具有可触及的顶端表面和生理相关的跨上皮电阻值。我们的技术提供了一种最新的新颖的肠上皮培养方法，以组织培养形式提供类似体内的细胞组成和分布，与高通量药物吸收或微生物-上皮相互作用研究兼容。
Intestinal organoids have emerged as a powerful in vitro tool for studying intestinal biology due to their resemblance to in vivo tissue at the structural and functional levels. However, their sphere-like geometry prevents access to the apical side of the epithelium, making them unsuitable for standard functional assays designed for flat cell monolayers. Here, we describe a simple method for the formation of epithelial monolayers that recapitulates the in vivo-like cell type composition and organization and that is suitable for functional tissue barrier assays. In our approach, epithelial monolayer spreading is driven by the substrate stiffness, while tissue barrier function is achieved by the basolateral delivery of medium enriched with stem cell niche and myofibroblast-derived factors. These monolayers contain major intestinal epithelial cell types organized into proliferating crypt-like domains and differentiated villus-like regions, closely resembling the in vivo cell distribution. As a unique characteristic, these epithelial monolayers form functional epithelial barriers with an accessible apical surface and physiologically relevant transepithelial electrical resistance values. Our technology offers an up-to-date and novel culture method for intestinal epithelium, providing an in vivo-like cell composition and distribution in a tissue culture format compatible with high-throughput drug absorption or microbe-epithelium interaction studies.