Monitoring Effects of Membrane Traffic Via Changes in Cell Polarity and Morphogenesis in Three-Dimensional Human Pluripotent Stem Cell Cysts. Methods Mol Biol 2023;2557:83-98
Date
12/14/2022Pubmed ID
36512211Pubmed Central ID
PMC10276343DOI
10.1007/978-1-0716-2639-9_7Scopus ID
2-s2.0-85144231102 (requires institutional sign-in at Scopus site)Abstract
Membrane traffic at the Golgi and endosomes plays many critical roles in the polarization and the morphogenesis of epithelial tissues. Studies into the roles of traffic in morphogenesis in mammals are often complicated by early embryonic lethality of mutations in membrane traffic as well as the inherent difficulty in imaging developing embryos posed by their size and location. Increasingly, human pluripotent stem cell (hPSC)-derived embryo- and organ-like systems (e.g., embryoids, organoids) provide a useful platform to illuminate the requirements of traffic in human embryonic tissue morphogenesis because these in vitro models are highly amenable to fluorescence microscopy and provide the ability to examine the role of essential genes not possible with animal studies. Here, we present a method to generate hPSC-cysts, a 3-D hPSC-based model of human epiblast lumen formation. This system provides unique opportunities to examine the role of membrane traffic during epithelial morphogenesis. We also present methods to process hPSC-cysts for immunofluorescence and staining with commonly used fluorescence labels useful for detecting defects in polarization and morphogenesis caused by defects in membrane traffic.
Author List
Hamed MM, Taniguchi K, Duncan MCAuthor
Kenichiro Taniguchi PhD Assistant Professor in the Cell Biology, Neurobiology and Anatomy department at Medical College of WisconsinMESH terms used to index this publication - Major topics in bold
AnimalsCell Differentiation
Cell Polarity
Cysts
Humans
Mammals
Morphogenesis
Organoids
Pluripotent Stem Cells
