Functional comparison of distinct Brachyury+ states in a renal differentiation assay.

Zhou, Jing, Plagge, Antonius ORCID: 0000-0001-6592-1343 and Murray, Patricia (2018) Functional comparison of distinct Brachyury+ states in a renal differentiation assay. Biology open, 7 (5). bio031799-.

Text BiO manuscript postprint.docx - Author Accepted Manuscript Download (11MB)

Abstract

Mesodermal populations can be generated in vitro from mouse embryonic stem cells (mESCs) using three-dimensional (3-D) aggregates called embryoid bodies or two-dimensional (2-D) monolayer culture systems. Here, we investigated whether Brachyury-expressing mesodermal cells generated using 3-D or 2-D culture systems are equivalent, or instead, have different properties. Using a Brachyury-GFP/E2-Crimson reporter mESC line, we isolated Brachyury-GFP+ mesoderm cells using flow-activated cell sorting and compared their gene expression profiles and ex vivo differentiation patterns. Quantitative RT-PCR analysis showed significant up-regulation of Cdx2, Foxf1 and Hoxb1 in the Brachyury-GFP+ cells isolated from the 3-D system compared with those isolated from the 2-D system. Furthermore, using an ex vivo mouse kidney rudiment assay, we found that irrespective of their source, Brachyury-GFP+ cells failed to integrate into developing nephrons, which are derived from the intermediate mesoderm. However, Brachyury-GFP+ cells isolated under 3-D conditions appeared to differentiate into endothelial-like cells within the kidney rudiments, whereas the Brachyury-GFP+ isolated from the 2-D conditions only did so to a limited degree. The high expression of Foxf1 in the 3-D Brachyury-GFP+ cells combined with their tendency to differentiate into endothelial-like cells suggests these mesodermal cells may represent lateral plate mesoderm.

Item Type:	Article
Uncontrolled Keywords:	mouse embryonic stem cells, embryoid bodies, Brachyury, mesodermal differentiation, ex vivo culture, kidney rudiments
Depositing User:	Symplectic Admin
Date Deposited:	20 Apr 2018 10:34
Last Modified:	19 Jan 2023 06:35
DOI:	10.1242/bio.031799
Related URLs:	Author Publisher
URI:	https://livrepository.liverpool.ac.uk/id/eprint/3020404