Efficient Generation of Functional Epithelial and Epidermal Cells from Human Pluripotent Stem Cells Under Defined Conditions

Abstract

Human pluripotent stem cells (hPSCs) have an unparalleled potential to generate limitless quantities of any somatic cell type. However, current methods for producing populations of various somatic cell types from hPSCs are generally not standardized and typically incorporate undefined cell culture components often resulting in variable differentiation efficiencies and poor reproducibility. To address this, we have developed a defined approach for generating epithelial progenitor and epidermal cells from hPSCs. In doing so, we have identified an optimal starting cell density to maximize yield and maintain high purity of K18+/p63+ simple epithelial progenitors. In addition, we have shown that the use of synthetic, defined substrates in lieu of Matrigel and gelatin can successfully facilitate efficient epithelial differentiation, maintaining a high (>75%) purity of K14+/p63+ keratinocyte progenitor cells and at a two to threefold higher yield than a previously reported undefined differentiation method. These K14+/p63+ cells also exhibited a higher expansion potential compared to cells generated using an undefined differentiation protocol and were able to terminally differentiate and recapitulate an epidermal tissue architecture in vitro. In summary, we have demonstrated the production of populations of functional epithelial and epidermal cells from multiple hPSC lines using a new, completely defined differentiation strategy.