The repair of wounds is one of the most complex biological processes that occur during human life. After an injury, multiple biological pathways immediately become activated and are synchronized to respond. In human adults, the wound repair process commonly leads to a non-functioning mass of fibrotic tissue known as a scar. By contrast, early in gestation, injured fetal tissues can be completely recreated, without fibrosis, in a process resembling regeneration. Some organisms, however, retain the ability to regenerate tissue throughout adult life. Knowledge gained from studying such organisms might help to unlock latent regenerative pathways in humans, which would change medical practice as much as the introduction of antibiotics did in the twentieth century.

Colony-forming human epidermal cells are heterogeneous in their capacity for sustained growth. Once a clone has been derived from a single cell, its growth potential can be estimated from the colony types resulting from a single plating, and the clone can be assigned to one of three classes. The holoclone has the greatest reproductive capacity: under standard conditions, fewer than 5% of the colonies formed by the cells of a holoclone abort and terminally differentiate. The paraclone contains exclusively cells with a short replicative lifespan (not more than 15 cell generations), after which they uniformly abort and terminally differentiate. The third type of clone, the meroclone, contains a mixture of cells of different growth potential and is a transitional stage between the holoclone and the paraclone. The incidence of the different clonal types is affected by aging, since cells originating from the epidermis of older donors give rise to a lower proportion of holoclones and a higher proportion of paraclones.The epidermis is a stratified squamous epithelium whose differentiated cells are the progeny of proliferative cells located mainly in the basal cell layer. Although many of the basal cells are capable of multiplication (1), few of them are thought to be self-renewing stem cells (2-4). We have recently shown that the clone-forming ability of a human keratinocyte in culture can be estimated from its size: small keratinocytes give rise to clones with high frequency, larger ones do so with lower frequency, and still larger ones, not at all. But once a colony has formed, its growth potential is not specified by the size of the founding cell (5).We describe here a method of analysis that reveals the growth potential of individual clones. We inoculate a single founding cell, and 7 days later we transfer the progeny, while they are still growing exponentially, to indicator dishes, where they are allowed to grow for a further period of 12 days. According to the growth in the indicator dishes, we can classify the original clone. Holoclones (holo = entire) form large rapidly growing colonies; fewer than 5% of the colonies abort and terminally differentiate. Paraclones (para = beyond) are programmed for limited growth and consequently form uniformly small, terminal colonies on the indicator dishes. Meroclones (mero = partial) form two kinds of colonies on the indicator dishes-growing and terminal. The meroclone therefore contains a proportion of cells that have degraded to paraclone-formers. MATERIALS AND METHODSCell Culture. Human epidermal keratinocytes were cultivated as previously described (6), using lethally irradiated supporting 3T3 cells (7). The epidermal growth factor used to promote multiplication (8) was the cloned human polypeptide kindly provided by Chiron (Emeryville, CA) (9). All experiments were carried out with a single batch of serum tested for its ability to support colony formation. The medium was changed every 4 days. Strains AY and YF 19 were derived from foreskin, strain GMA from thigh,...

Stem cells which have the capacity to self-renew and generate differentiated progeny are thought to be maintained in a specific environment known as a niche. The localization of the niche, however, remains largely obscure for most stem-cell systems. Melanocytes (pigment cells) in hair follicles proliferate and differentiate closely coupled to the hair regeneration cycle. Here we report that stem cells of the melanocyte lineage can be identified, using Dct-lacZ transgenic mice, in the lower permanent portion of mouse hair follicles throughout the hair cycle. It is only the population in this region that fulfils the criteria for stem cells, being immature, slow cycling, self-maintaining and fully competent in regenerating progeny on activation at early anagen (the growing phase of hair follicles). Induction of the re-pigmentation process in K14-steel factor transgenic mice demonstrates that a portion of amplifying stem-cell progeny can migrate out from the niche and retain sufficient self-renewing capability to function as stem cells after repopulation into vacant niches. Our data indicate that the niche has a dominant role in the fate determination of melanocyte stem-cell progeny.

The upper region of the outer root sheath of vibrissal follicles of adult mice contains multipotent stem cells that respond to morphogenetic signals to generate multiple hair follicles, sebaceous glands, and epidermis, i.e., all the lineages of the hairy skin. At the time when hair production ceases and when the lower region of the follicle undergoes major structural changes, the lower region contains a significant number of clonogenic keratinocytes, and can then respond to morphogenetic signals. This demonstrates that multipotent stem cells migrate to the root of the follicle to produce whisker growth. Moreover, our results indicate that the clonogenic keratinocytes are closely related, if not identical, to the multipotent stem cells, and that the regulation of whisker growth necessitates a precise control of stem cell trafficking.