Poole Nina M, Rajan Anubama, Maresso Anthony W
Department of Molecular Virology and Microbiology, Baylor College of Medicine, Houston, Texas.
Curr Protoc Microbiol. 2018 Aug;50(1):e55. doi: 10.1002/cpmc.55. Epub 2018 May 17.
Adherence, invasion, and translocation to and through the intestinal epithelium are important drivers of disease for many enteric bacteria. However, most work has been limited to transformed intestinal cell lines or murine models that often do not faithfully recapitulate key elements associated with human disease. The recent technological advances in organotypic tissue and cell culture are providing unparalleled access to systems with human physiology and complexity. Human intestinal enteroids (HIEs), derived from patient biopsy or surgical specimens of intestinal tissues, are organotypic cultures now being adapted to the study of enteric infections. HIEs are comprised of the dominant cell types of the human gastrointestinal epithelium, can be grown in two- or three-dimensional structures, form a crypt-villus axis with defined apical and basolateral compartments, and undergo physiologic responses to many different stimuli. Here, we describe a series of protocols that encompass the use of human enteroids for the measurement of the adherence, invasion, and translocation of E. coli to and through the intestinal epithelium. We also outline the steps needed to grow and prepare enteroids for this purpose and highlight some common problems to troubleshoot. © 2018 by John Wiley & Sons, Inc.
对于许多肠道细菌而言，黏附、侵袭以及向肠上皮细胞的转移和穿过肠上皮细胞是引发疾病的重要因素。然而，大多数研究工作局限于转化的肠细胞系或小鼠模型，这些模型往往无法如实地重现与人类疾病相关的关键要素。近期在器官型组织和细胞培养方面的技术进步，为研究具有人类生理特征和复杂性的系统提供了前所未有的便利。源自患者肠道组织活检或手术标本的人肠道类器官（HIEs）是一种器官型培养物，目前正被应用于肠道感染的研究。HIEs由人胃肠道上皮的主要细胞类型组成，可以在二维或三维结构中生长，形成具有明确顶端和基底外侧区室的隐窝 - 绒毛轴，并对许多不同刺激产生生理反应。在此，我们描述了一系列方案，包括使用人肠道类器官来测量大肠杆菌对肠上皮细胞的黏附、侵袭以及穿过肠上皮细胞的转移情况。我们还概述了为此目的培养和制备肠道类器官所需的步骤，并突出了一些常见问题及解决方法。© 2018 约翰威立国际出版公司
