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Mouse Embryonic Stem Cell–Derived Embryoid Bodies Generate Progenitors That Integrate Long Term into Renal Proximal Tubules In Vivo

Cécile Vigneau^*,, Katalin Polgar^*, Gary Striker^*, Justine Elliott^*, Deborah Hyink^*, Odile Weber, Hans-Joerg Fehling, Gordon Keller, Christopher Burrow^* and Patricia Wilson^*

Departments of ^* Medicine and Gene and Cell Medicine, Mount Sinai School of Medicine, New York, New York; Department of Medicine, University Pierre and Marie Curie, Paris, France; and Department of Immunology, University of Ulm, Ulm, Germany

Address correspondence to: Dr. Patricia Wilson, Mount Sinai School of Medicine, Division of Nephrology, Department of Medicine, 1425 Madison Avenue, New York, NY 10029. Phone: 212-659-9383; Fax: 212-849-2434; E-mail: pat.wilson{at}mssm.edu

Received for publication October 2, 2006. Accepted for publication March 18, 2007.

The metanephric kidney is a mesodermal organ that develops as a result of reciprocal interactions between the ureteric bud and the blastema. The generation of embryonic stem (ES) cell–derived progenitors offers potential for regenerative therapies but is often limited by development of tumor formation. Because brachyury (T) denotes mesoderm specification, a mouse ES cell line with green fluorescence protein (GFP) knocked into the functional T locus as well as lacZ in the ROSA26 locus (LacZ/T/GFP) was used in cell selection and lineage tracing. In the absence of leukemia inhibitory factor, mouse ES cells give rise to embryoid bodies that can differentiate into mesoderm. Culture conditions were optimized (4 d, 10 ng/ml Activin-A) to generate maximal numbers of renal progenitor populations identified by expression of the specific combination of renal markers cadherin-11, WT-1, Pax-2, and Wnt-4. LacZ/T/GFP+ cells were further enriched by FACS selection. Five days after injection of LacZ/T/GFP+ cells into embryonic kidney explants in organ culture, -galactosidase immunohistochemistry showed incorporation into blastemal cells of the nephrogenic zone. After a single injection into developing live newborn mouse kidneys, co-localization studies showed that the LacZ/T/GFP+ cells were stably integrated into proximal tubules with normal morphology and normal polarization of alkaline phosphatase and aquaporin-1 for 7 mo, without teratoma formation. It is concluded that defined differentiation of ES cells into embryoid bodies with Activin-A and selection for T expression provides a means to isolate and purify renal proximal tubular progenitor cells with the potential for safe use in regenerative therapies.

Copyright © 2008 by the American Society of Nephrology. Online ISSN: 1533-3450 Print ISSN: 1046-6673