Basal-type lumenogenesis in extraembryonic endoderm stem cells models the early visceral endoderm

Cultured rat primitive extraembryonic endoderm (pXEN) cells easily form free-floating multicellular vesicles de novo, exemplifying a poorly studied type of morphogenesis. Here, we reveal the underlying mechanism and the identity of the vesicles. We resolve the morphogenesis into vacuolization, vesiculation and maturation, and define the molecular characteristics and requirements of each step. Vacuolization is fueled by macropinocytosis and occurs by default if not blocked by high cell density or matrix proteins. Fine-tuned cell-cell contact then forms nascent three-cell vesicles with vacuole-derived lumina. In maturation, the vesicles complete epithelialization, expand via mitosis and continued fluid uptake, and differentiate further. The mature vesicles consist of a simple squamous epithelium with an apical-outside/basal-inside polarity that we trace back to the single cell stage. The polarity and gene expression pattern of the vesicles are similar to those of the early visceral endoderm. pXEN cells provide a useful in vitro model for study of matrix-independent, basal-type lumenogenesis and the physiology of the visceral endoderm.This article has an associated First Person interview with the first author of the paper.