Induction of putative stratified epithelial progenitor cells in vitro from mouse-induced pluripotent stem cells

Abstract

Induced pluripotent stem (iPS) cells generally exhibit a normal karyotype, are transcriptionally and epigenetically similar to embryonic stem (ES) cells, and maintain the potential to differentiate into derivatives of all germ layers. Recently, the use of different types of cell or tissue derived from iPS cells for transplantation has become a possibility. However, the differentiation of epithelial lineages from iPS cells has not yet been demonstrated. We attempted to establish a culture technique for the induction of epithelial progenitors from mouse iPS cells. Mouse iPS cells were cultured on dishes coated with type IV collagen in keratinocyte culture medium (KCM) supplemented with or without bone morphogenic protein-4 (BMP-4) or combined with pretreatment of retinoic acid (RA) and BMP-4 in the undifferentiated state. Markers for undifferentiated cells (Oct3/4, Nanog) and for differentiation (p63, cytokeratin14) were analyzed by immunofluorescence staining and real-time RT-PCR. Putative epithelial progenitors were successfully induced in vitro from iPS cells. These progenitors expressed p63, a transcription factor necessary for maintenance of regenerative epithelia and cytokeratin 14 constitutively present in the basal layer of stratified epithelia. Enhancement of putative epithelial progenitor commitment was observed when cultured in KCM with BMP-4 following pretreatment of RA and BMP-4. The differentiation efficiency of putative epithelial progenitors from iPS cell cultures was similar to that of ES cell cultures. This report is the first to demonstrate in vitro differentiation of iPS cells into putative epithelial progenitors. These iPS-derived putative epithelial progenitors provide a powerful tool for understanding the mechanisms of epithelial lineage differentiation.