FOXN1GFP/w Reporter hESCs Enable Identification of Integrin-β4, HLA-DR, and EpCAM as Markers of Human PSC-Derived FOXN1+ Thymic Epithelial Progenitors

Summary 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 FOXN1GFP/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.
Highlights • FOXN1-GFP reporter hESC lines were generated • KGF promotes the proliferation of FOXN1-GFP+ cells • FOXN1-GFP+ cells express TEC-associated genes • ITGB4, HLA-DR, and EpCAM can be used to purify FOXN1+ TEC progenitors (219)
Stanley and colleagues generated human embryonic stem cell (hESC) reporter lines in which sequences encoding GFP were inserted into the FOXN1 locus, enabling the isolation of viable thymic progenitors from in vitro differentiation cultures. These reporter lines were used to identify integrin-β4, HLA-DR, and EpCAM as markers of human pluripotent stem cell-derived FOXN1+ thymic epithelial progenitors.