Inventors: Elaine Fuchs, Michael Rendi
Publication date: 2008-09-04
Abstract:
The present invention relates to dermal papilla-specific markers and a method for isolating dermal papilla cells using the same. Dermal papilla cells isolated in accordance with the method of the invention and treated with BMP6 are useful for promoting hair growth.
Claims:
A method for isolating a dermal papilla cell comprising obtaining a population of cells from a sample and sorting the population of cells based on the absence of CD34, CD45 and CD117 and the presence of a selected dermal papilla cell marker expressed by each cell, so that a dermal papilla cell is isolated.
2. A cell isolated according to the method of claim 1.
3. An isolated dermal papilla cell wherein said cell lacks CD34, CD45 and CD117 and expresses a selected dermal papilla cell marker.
4. A method for promoting hair growth comprising contacting dermal papilla cells with an effective amount of BMP6 so that hair growth is promoted when said dermal papilla cells are contacted with skin cells.
Background:
During embryogenesis, hair follicle formation is dependent upon a series of reciprocal interactions between the single-layered epithelium and a dermal cell condensate. This specialized cluster of mesenchymal cells becomes enveloped by the epithelial (matrix) cells at the base of the developing follicle, and postnatally, they persist as the dermal papilla (Hardy (1992)Trends Genet. 8:55-61; Schmidt-Ullrich and Paus (2005) Bioessays 27:247-261).
The architecture and biology of the mature follicle is complex. At the base, and in close association with the dermal papilla, matrix cells are transiently proliferative and maintain a relatively undifferentiated status. As matrix cells progress upward, they differentiate into the hair shaft (cortex and medulla) and the channel or inner root sheath that surrounds the hair. The inner root sheath is then encased by an outer root sheath contiguous with the epidermis. The entire structure is enclosed by a basement membrane composed of extracellular matrix proteins that separate the skin epithelium from dermis and dermal papilla. A small number of follicle melanocytes reside just above this membrane in the epithelial compartment of the hair bulb.
When matrix cells exhaust their proliferative capacity, the hair stops growing, and the lower epithelial part of the follicle enters a destructive phase (catagen). As the epithelium shrinks, the basement membrane and dermal papilla move upward. Following a resting period (telogen), epithelial stem cells at the base of the remaining hair follicle (the bulge) receive signals from the now adjacent dermal papilla and re-enter a growth phase (anagen) to regenerate the follicle and produce a new hair.
Genetic engineering has recently enabled the isolation of epithelial stem cells within the bulge (Tumbar, et al. (2004) Science 303:359-363; Morris, et al. (2004) Nat. Biotechnol. 22:411-417). When exposed to skin dermis, the descendants of a single epithelial stem cell can give rise to epidermis, follicles and sebaceous glands when engrafted onto the backs of Nude mice lacking hair (Blanpain, et al. (2004) Cell 118:635-648). It has long been recognized that the critical dermal cells in this process are the dermal papilla (Hardy (1992) supra). In contrast to dermal (3T3) skin fibroblasts, which only permit epidermal repair in this assay, microdissected rat whisker dermal papilla cells induce hair growth (Jahoda, et al. (1984) Nature 311:560-562; Lichti, et al. (1993) J. Invest. Dermatol. 101:124S-129S). In vitro, the dermal papilla cells lose this ability. Co-culturing dermal papilla either with epidermal keratinocytes (Inamatsu, et al. (1998) J. Invest. Dermatol. 111:767-775) or with embryonic fibroblasts expressing a Wnt3a, but not a Sonic hedgehog transgene (Kishimoto, et al. (2000) Genes Dev. 14:1181-11), prolongs their potential. However, more recent studies suggest that Wnt3a on its own is not sufficient and that additional as yet unidentified factors are necessary for maintaining dermal papilla activity (Shimizu and Morgan (2004) J. Invest. Dermatol. 122:239-245).
A knowledge of the genes expressed by the dermal papilla and its neighbors would be of value in sifting through the complex mechanisms by which dermal papilla cells maintain their remarkable inductive function while in the niche and lose them outside of it. Most known dermal papilla markers have been found fortuitously. The relative inaccessibility of dermal papilla cells and/or their loss of potential in vitro have posed technical hurdles in isolating pure populations of these cells. Thus, although microarray and cDNA library analyses have been conducted on microdissected and/or cultured whisker dermal papillae (Sleeman, et al. (2000) Genomics 69:214-224; O'Shaughnessy, et al. (2004) Exp. Dermatol. 13:621-629; O'Shaughnessy, et al. (2004b) J. Invest. Dermatol. 123:613-621), the array data have yielded only a handful of the known dermal papilla markers, making it difficult to evaluate the potential significance of unexpectedly expressed genes from these arrays.
A set of putative dermal papilla markers emerged when it was noted that cell aggregates cultured from whole skin dermis bear a resemblance to neurospheres cultured from neural crest cells (Fernandes, et al. (2004) Nat. Cell Biol. 6:1082-1093). Subsequent in vivo studies revealed that neural crest markers localized in the vicinity of dermal papilla, raising speculation those neural progenitor cells in the skin are derived from dermal papilla. These analogies were complicated by the close proximity of melanocytes (neural crest derived) and dermal papilla in the follicle, and by the fact that the parallels were largely drawn from identifying SKP markers in rodent whiskers. In contrast to other body sites, the entire head mesenchyme develops embryologically from neural crest (Le Douarin and Dupin (1993) J. Neurobiol. 24:146-161). These potential caveats aside, the existence of a population of multipotent neuroprogenitor cells in adult follicles would place the dermal papilla squarely at the center of significant clinical relevance. Accordingly, needed in the art are suitable markers for the identification and isolation of such cells. The present invention meets this long-felt need.
Summary:
The present invention is a method for isolating a dermal papilla cell. The method involves the steps of obtaining a population of cells from a sample and sorting the population of cells based on the absence of CD34, CD45 and CD117 and the presence of a selected dermal papilla cell marker expressed by each cell. Isolated dermal papilla cells are lacking CD34, CD45 and CD117 and expressing a selected dermal papilla cell marker are also provided.
The present invention is also a method for promoting hair growth by contacting dermal papilla cells with an effective amount of BMP6 so that hair growth is promoted when said dermal papilla cells are contacted with skin cells. |
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