Functional Characterization of Quiescent Keratinocyte Stem Cells and Their Progeny Reveals a Hierarchical Organization in Human Skin Epidermis

Although homeostatic renewal of human skin epidermis is achieved by the combined activity of quiescent stem cells (SCs) and their actively cycling progeny, whether these two populations are equipotent in their capacity to regenerate tissue has not been determined in biological assays that mimic lifelong renewal. Using fluorescence activated cell separation strategy validated previously by us, human epidermis was fractionated into three distinct subsets: that is, alpha(bri)(6)CD71(dim), alpha(bri)(6)CD71(bri), and alpha(dim)(6) with characteristics of keratinocyte stem, transient amplifying, and early differentiating cells, respectively. The global gene expression profile of these fractions was determined by microarray, confirming that the alpha(bri)(6)CD71(dim) subset was quiescent, the alpha(bri)(6)CD71(bri) was actively cycling, and the alpha(dim)(6) subset expressed markers of differentiation. More importantly, functional evaluation of these populations in an in vivo model for tissue reconstitution at limiting cell dilutions revealed that the quiescent alpha(bri)(6)CD71(dim) fraction was the most potent proliferative and tissue regenerative population of the epidermis, capable of long-term (LT) epidermal renewal from as little as 100 cells for up to 10 weeks. In contrast, the cycling alpha(bri)(6)CD71(bri) fraction was the first to initiate tissue reconstitution, although this was not sustained in the LT, while differentiating alpha(dim)(6) cells possessed the lowest demonstrable tissue regenerative capacity. Our data suggest that in human skin, the epidermal proliferative compartment is not composed of equipotent cells, but rather is organized in a functionally hierarchical manner with the most potent quiescent SCs at its apex (i.e., alpha(bri)(6)CD71(dim)) followed by cycling progenitors (i.e., alpha(bri)(6)CD71(bri)) and finally early differentiating keratinocytes (i.e., alpha(dim)(6)). STEM CELLS 2011; 29:1256-1268