FOXN1 (GFP/w) reporter hESCs enable identification of integrin-β4, HLA-DR, and EpCAM as markers of human PSC-derived FOXN1(+) thymic epithelial progenitors

Chew-Li Soh; Antonietta Giudice; Robert A Jenny; David A Elliott; Tanya Hatzistavrou; Suzanne J Micallef; Korosh Kianizad; Natalie Seach; Juan Carlos Zúñiga-Pflücker; Ann P Chidgey; Alan Trounson; Susan K Nilsson; David N Haylock; Richard L Boyd; Andrew G Elefanty; Edouard G Stanley

doi:10.1016/j.stemcr.2014.04.009

FOXN1 (GFP/w) reporter hESCs enable identification of integrin-β4, HLA-DR, and EpCAM as markers of human PSC-derived FOXN1(+) thymic epithelial progenitors

Stem Cell Reports. 2014 May 22;2(6):925-37. doi: 10.1016/j.stemcr.2014.04.009. eCollection 2014 Jun 3.

Authors

Chew-Li Soh¹, Antonietta Giudice¹, Robert A Jenny¹, David A Elliott², Tanya Hatzistavrou¹, Suzanne J Micallef¹, Korosh Kianizad³, Natalie Seach¹, Juan Carlos Zúñiga-Pflücker³, Ann P Chidgey¹, Alan Trounson⁴, Susan K Nilsson⁵, David N Haylock⁵, Richard L Boyd¹, Andrew G Elefanty⁶, Edouard G Stanley⁷

Affiliations

¹ Department of Anatomy and Developmental Biology, Monash University, Clayton, VIC 3800, Australia.
² Murdoch Childrens Research Institute, The Royal Children's Hospital, Parkville, VIC 3052, Australia.
³ Sunnybrook Research Institute, Department of Immunology, University of Toronto, Toronto, ON M4N 3M5, Canada.
⁴ California Institute for Regenerative Medicine, San Francisco, CA 94107, USA.
⁵ Materials Science and Engineering, Commonwealth Scientific and Industrial Research Organisation, Clayton, VIC 3168, Australia ; Australian Regenerative Medicine Institute, Monash University, Clayton, VIC 3800, Australia.
⁶ Department of Anatomy and Developmental Biology, Monash University, Clayton, VIC 3800, Australia ; Murdoch Childrens Research Institute, The Royal Children's Hospital, Parkville, VIC 3052, Australia.
⁷ Department of Anatomy and Developmental Biology, Monash University, Clayton, VIC 3800, Australia ; Murdoch Childrens Research Institute, The Royal Children's Hospital, Parkville, VIC 3052, Australia ; Department of Paediatrics, University of Melbourne, Parkville, VIC 3050, Australia.

Abstract

Thymic epithelial cells (TECs) play a critical role in T cell maturation and tolerance induction. The generation of TECs from in vitro differentiation of human pluripotent stem cells (PSCs) provides a platform on which to study the mechanisms of this interaction and has implications for immune reconstitution. To facilitate analysis of PSC-derived TECs, we generated hESC reporter lines in which sequences encoding GFP were targeted to FOXN1, a gene required for TEC development. Using this FOXN1 (GFP/w) line as a readout, we developed a reproducible protocol for generating FOXN1-GFP(+) thymic endoderm cells. Transcriptional profiling and flow cytometry identified integrin-β4 (ITGB4, CD104) and HLA-DR as markers that could be used in combination with EpCAM to selectively purify FOXN1(+) TEC progenitors from differentiating cultures of unmanipulated PSCs. Human FOXN1(+) TEC progenitors generated from PSCs facilitate the study of thymus biology and are a valuable resource for future applications in regenerative medicine.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Antigens, Neoplasm / metabolism*
Cell Adhesion Molecules / metabolism*
Cell Differentiation
Cells, Cultured
Epithelial Cell Adhesion Molecule
Epithelial Cells / cytology
Epithelial Cells / metabolism*
Forkhead Transcription Factors / metabolism*
HLA-DR Antigens / metabolism*
Humans
Integrin beta4 / metabolism*
Pluripotent Stem Cells / cytology*
Pluripotent Stem Cells / metabolism
Thymus Gland / cytology*

Substances

Antigens, Neoplasm
Cell Adhesion Molecules
EPCAM protein, human
Epithelial Cell Adhesion Molecule
Forkhead Transcription Factors
HLA-DR Antigens
Integrin beta4
Whn protein