Location and phenotype of human adult keratinocyte stem cells of the skin

The location and identity of interfollicular epidermal stem cells of adult human skin remain undefined. Based on our previous work in both adult murine and neonatal human foreskin, we demonstrate that cell surface levels of the alpha(6) integrin and the transferrin receptor (CD71) are valid markers for resolving a putative stem cell, transit amplifying and differentiating compartment in adult human skin by flow cytometry. Specifically, epidermal cells expressing high levels of alpha(6) integrin and low levels of the transferrin receptor CD71 (phenotype alpha(6)(bri)CD71(dim)) exhibit several stem cell characteristics, comprising a minor population (2%-5%) of the K14(bri) fraction, enriched for quiescent and small blast-like cells with high clonogenic capacity, lacking the differentiation marker K10. Conversely, the majority of K14(bri) K10(neg) epidermal cells express high levels of CD71 (phenotype alpha(6)(bri)CD71(bri)), and represent the actively cycling fraction of keratinocytes displaying greater cell size due to an increase in cytoplasmic area, consistent with their being transient amplifying cells. The alpha(6)(bri)CD71(bri) population exhibited intermediate clonogenic capacity. A third population of K14(dim) but K10 positive epidermal cells could be identified by their low levels of alpha(6) integrin expression (i.e. alpha(6)(dim) cells), representing the differentiation compartment; predictably, this subpopulation exhibited poor clonogenic efficiency. Flow cytometric analysis for the hair follicle bulge region (stem cell) marker K15 revealed preferential expression of this keratin in alpha(6)(bri) cells (i.e., both stem and transient amplifying fractions), but not the alpha(6)(dim) population. Given that K15 positive cells could only be detected in the deep rete ridges of adult skin in situ, we conclude that stem and transient amplifying cells reside in this location, while differentiating (K15 negative) cells are found in the shallow rete ridges.