B3.3 Potential of nestin-expressing hair follicle cells to form neurons and blood vessels

Robert M. Hoffman, Lingna Li, Yasuyuki Amoh, Meng Yang

AntiCancer, Inc., 7917 Ostrow Street, San Diego, CA 92111, USA

The intermediate filament protein, nestin, marks progenitor cells of the CNS. Such CNS stem cells are selectively labeled by placing GFP under the control of the nestin regulatory sequences in transgenic mice. During early anagen or growth phase of the hair follicle, nestin-expressing cells, marked by GFP fluorescence in nestin-GFP transgenic mice, appear in the permanent upper hair follicle immediately below the sebaceous glands in the follicle bulge where the hair follicle stem cells are located. The relatively small, oval-shaped, nestin-expressing cells in the bulge area surround the hair shaft. During telogen or resting phase and in early anagen, the GFP-positive cells are mainly in the bulge area. However, in mid- and late-anagen, the GFP-expressing cells are located in the upper outer-root sheath as well as in the bulge area but not in the hair matrix bulb. These observations show that the nestin-expressing cells form the outer-root sheath and possibly other follicle structures. Immunohistochemical staining showed that nestin, GFP, keratin 5/8, and keratin 15 colocalize in the hair follicle bulge cells, outer-root sheath cells, and basal cells of the sebaceous glands. These data indicate that nestin-expressing cells located in the hair follicle bulge are the hair follicle stem cells. The expression of the unique protein, nestin, in both neural stem cells and hair follicle stem cells suggested their possible relation. In vitro experiments have shown that the nestin-expressing follicle stem cells can be converted to neurospheres, which in turn can be converted to neurons. In the nestin-GFP transgenic mice, the hair follicles are linked by a network of nestin-GFP-expressing blood vessels. When a red fluorescent protein (RFP)-expressing B16 melanoma was transplanted to the nestin-GFP mice, nestin-expressing vessels grew from the hair follicles into the developing melanoma.  When fibrissa hair follicles were transplanted from nestin-GFP mice to nude mice, nascent GFP-expressing vessels were observed growing from the transplanted hair follicles in the nude mouse skin. When two nestin-GFP-expressing follicles were transplanted to nude mice, they formed a network of nestin-GFP-expressing blood vessels interconnecting the follicles.  Immunohistochemical staining showed that endothelial-cell-specific markers CD31 as well as von Willebrand factor (vWF) and nestin colocalize in the nestin-GFP-expressing vessels both in the nestin-GFP mice and in nude mice transplanted with nestin-GFP-expressing vibrissa hair follicles.  The data thus suggest hair follicles give rise to blood vessels in the skin. This model enables very early events in skin angiogenesis, including skin-tumor angiogenesis to be visualized. The results suggest a new paradigm of the pluripotency of hair follicle nestin-expressing cells which can form blood vessels as well as much of the hair follicle structure and can be converted to neurons.