Early-age-dependent selective decrease of differentiation potential of hair-follicle-associated pluripotent (HAP) stem cells to beating cardiac-muscle cells

We have previously discovered nestin-expressing hair-follicle-associated pluripotent (HAP) stem cells and have shown that they can differentiate to neurons, glia, and many other cell types. HAP stem cells can be used for nerve and spinal cord repair. We have recently shown the HAP stem cells can differentiate to beating heart-muscle cells and tissue sheets of beating heart-muscle cells. In the present study, we determined the efficiency of HAP stem cells from mouse vibrissa hair follicles of various ages to differentiate to beating heart-muscle cells. We observed that the whiskers located near the ear were more efficient to differentiate to cardiac-muscle cells compared to whiskers located near the nose. Differentiation to cardiac-muscle cells from HAP stem cells in cultured whiskers in 4-week-old mice was significantly greater than in 10-, 20-, and 40-week-old mice. There was a strong decrease in differentiation potential of HAP stem cells to cardiac-muscle cells by 10 weeks of age. In contrast, the differentiation potential of HAP stem cells to other cell types did not decrease with age. The possibility of rejuvenation of HAP stem cells to differentiate at high efficiency to cardiac-muscle cells is discussed.     

Implanted hair-follicle-associated pluripotent (HAP) stem cells encapsulated in polyvinylidene fluoride membrane cylinders promote effective recovery of peripheral nerve injury

Hair follicle-associated-pluripotent (HAP) stem cells are located in the bulge area of the hair follicle, express the stem-cell marker, nestin, and have been shown to differentiate to nerve cells, glial cells, keratinocytes, smooth muscle cells, cardiac muscle cells, and melanocytes. Transplanted HAP stem cells promote the recovery of peripheral nerve and spinal cord injuries and have the potential for heart regeneration as well. In the present study, we implanted mouse green fluorescent protein (GFP)-expressing HAP stem-cell spheres encapsulated in polyvinylidene fluoride (PVDF)-membrane cylinders into the severed sciatic nerve of immunocompetent and immunocompromised (nude) mice. Eight weeks after implantation, immunofluorescence staining showed that the HAP stem cells differentiated into neurons and glial cells. Fluorescence microscopy showed that the HAP stem cell hair spheres promoted rejoining of the sciatic nerve of both immunocompetent and immunodeficient mice. Hematoxylin and eosin (H&E) staining showed that the severed scatic nerves had regenerated. Quantitative walking analysis showed that the transplanted mice recovered the ability to walk normally. HAP stem cells are readily accessible from everyone, do not form tumors, and can be cryopreserved without loss of differentiation potential. These results suggest that HAP stem cells may have greater potential than iPS or ES cells for regenerative medicine.     

Direct transplantation of uncultured hair‐follicle pluripotent stem (hfPS) cells promotes the recovery of peripheral nerve injury

We previously showed that the stem cell marker nestin is expressed in hair follicle stem cells which suggested their pluripotency. We subsequently showed that the nestin‐expressing hair‐follicle pluripotent stem (hfPS) cells can differentiate in culture to neurons, glial cells, keratinocytes, and other cell types and can promote regeneration of peripheral nerve and spinal cord injuries upon injection to the injured nerve or spinal cord. The location of the hfPS cells has been termed the hfPS cell area (hfPSCA). Previously, hfPS cells were cultured for 1–2 months before transplantation to the injured nerve or spinal cord which would not be optimal for clinical application of these cells for nerve or spinal cord repair, since the patient should be treated soon after injury. In the present study, we addressed this issue by directly using the upper part of the hair follicle containing the hfPSCA, without culture, for injection into the severed sciatic nerve in mice. After injection of hfPSCA, the implanted hfPS cells grew and promoted joining of the severed nerve. The transplanted hfPS cells differentiated mostly to glial cells forming myelin sheaths, which promoted axonal growth and functional recovery of the severed nerve. These results suggest that the direct transplantation of the uncultured upper part of the hair follicle containing the hfPSA is an important method to promote the recovery of peripheral nerve injuries and has significant clinical potential. J. Cell. Biochem. 110: 272–277, 2010. © 2010 Wiley‐Liss, Inc.     

Multipotent nestin-expressing stem cells capable of forming neurons are located in the upper,middle and lower part of the vibrissa hair follicle

We have previously demonstrated that the neural stem-cell marker nestin is expressed in hair follicle stem cells. Nestin-expressing cells were initially identified in the hair follicle bulge area (BA) using a transgenic mouse model in which the nestin promoter drives the green fluorescent protein (ND-GFP). The hair-follicle ND-GFP-expressing cells are keratin 15-negative and CD34-positive and could differentiate to neurons, glia, keratinocytes, smooth muscle cells and melanocytes in vitro. Subsequently, we showed that the nestin-expressing stem cells could affect nerve and spinal cord regeneration after injection in mouse models. In the present study, we separated the mouse vibrissa hair follicle into three parts (upper, middle and lower). Each part of the follicle was cultured separately in DMEM-F12 containing B-27 and 1% methylcellulose supplemented with basic FGF. After 2 mo, the nestin-expressing cells from each of the separated parts of the hair follicle proliferated and formed spheres. Upon transfer of the spheres to RPMI 1640 medium containing 10% FBS, the nestin-expressing cells in the spheres differentiated to neurons, as well as glia, keratinocytes, smooth muscle cells and melanocytes. The differentiated cells were produced by spheres which formed from nestin-expressing cells from all segments of the hair follicle. However, the differentiation potential is greatest in the upper part of the follicle. This result is consistent with trafficking of nestin-expressing cells throughout the hair follicle from the bulge area to the dermal papilla that we previously observed. The nestin-expressing cells from the upper part of the follicle produced spheres in very large amounts, which in turn differentiated to neurons and other cell types. The results of the present study demonstrate that multipotent, nestin-expressing stem cells are present throughout the hair follicle and that the upper part of the follicle can produce the stem cells in large amounts that could be used for nerve and spinal cord repair.     

Multipotent nestin-expressing stem cells capable of forming neurons are located in the upper,middle and lower part of the vibrissa hair follicle

We have previously demonstrated that the neural stem-cell marker nestin is expressed in hair follicle stem cells. Nestin-expressing cells were initially identified in the hair follicle bulge area (BA) using a transgenic mouse model in which the nestin promoter drives the green fluorescent protein (ND-GFP). The hair-follicle ND-GFP-expressing cells are keratin 15-negative and CD34-positive and could differentiate to neurons, glia, keratinocytes, smooth muscle cells and melanocytes in vitro. Subsequently, we showed that the nestin-expressing stem cells could affect nerve and spinal cord regeneration after injection in mouse models. In the present study, we separated the mouse vibrissa hair follicle into three parts (upper, middle and lower). Each part of the follicle was cultured separately in DMEM-F12 containing B-27 and 1% methylcellulose supplemented with basic FGF. After 2 mo, the nestin-expressing cells from each of the separated parts of the hair follicle proliferated and formed spheres. Upon transfer of the spheres to RPMI 1640 medium containing 10% FBS, the nestin-expressing cells in the spheres differentiated to neurons, as well as glia, keratinocytes, smooth muscle cells and melanocytes. The differentiated cells were produced by spheres which formed from nestin-expressing cells from all segments of the hair follicle. However, the differentiation potential is greatest in the upper part of the follicle. This result is consistent with trafficking of nestin-expressing cells throughout the hair follicle from the bulge area to the dermal papilla that we previously observed. The nestin-expressing cells from the upper part of the follicle produced spheres in very large amounts, which in turn differentiated to neurons and other cell types. The results of the present study demonstrate that multipotent, nestin-expressing stem cells are present throughout the hair follicle and that the upper part of the follicle can produce the stem cells in large amounts that could be used for nerve and spinal cord repair.     

Isoproterenol directs hair follicle-associated pluripotent (HAP) stem cells to differentiate in vitro to cardiac muscle cells which can be induced to form beating heart-muscle tissue sheets

Nestin-expressing hair-follicle-associated pluripotent (HAP) stem cells are located in the bulge area of the follicle. Previous studies have shown that HAP stem cells can differentiate to neurons, glia, keratinocytes, smooth muscle cells, and melanocytes in vitro. HAP stem cells effected nerve and spinal cord regeneration in mouse models. Recently, we demonstrated that HAP stem cells differentiated to beating cardiac muscle cells. The differentiation potential to cardiac muscle cells was greatest in the upper part of the follicle. The beat rate of the cardiac muscle cells was stimulated by isoproterenol. In the present study, we observed that isoproterenol directs HAP stem cells to differentiate to cardiac muscle cells in large numbers in culture compared to HAP stem cells not supplemented with isoproterenol. The addition of activin A, bone morphogenetic protein 4, and basic fibroblast growth factor, along with isoproternal, induced the cardiac muscle cells to form tissue sheets of beating heart muscle cells. These results demonstrate that HAP stem cells have great potential to form beating cardiac muscle cells in tissue sheets.     

Human hair follicle pluripotent stem (hfPS) cells promote regeneration of peripheral‐nerve injury: An advantageous alternative to ES and iPS cells

The optimal source of stem cells for regenerative medicine is a major question. Embryonic stem (ES) cells have shown promise for pluripotency but have ethical issues and potential to form teratomas. Pluripotent stem cells have been produced from skin cells by either viral‐, plasmid‐ or transposon‐mediated gene transfer. These stem cells have been termed induced pluripotent stem cells or iPS cells. iPS cells may also have malignant potential and are inefficiently produced. Embryonic stem cells may not be suited for individualized therapy, since they can undergo immunologic rejection. To address these fundamental problems, our group is developing hair follicle pluripotent stem (hfPS) cells. Our previous studies have shown that mouse hfPS cells can differentiate to neurons, glial cells in vitro, and other cell types, and can promote nerve and spinal cord regeneration in vivo. hfPS cells are located above the hair follicle bulge in what we have termed the hfPS cell area (hfPSA) and are nestin positive and keratin 15 (K‐15) negative. Human hfPS cells can also differentiate into neurons, glia, keratinocytes, smooth muscle cells, and melanocytes in vitro. In the present study, human hfPS cells were transplanted in the severed sciatic nerve of the mouse where they differentiated into glial fibrillary‐acidic‐protein (GFAP)‐positive Schwann cells and promoted the recovery of pre‐existing axons, leading to nerve generation. The regenerated nerve recovered function and, upon electrical stimulation, contracted the gastrocnemius muscle. The hfPS cells can be readily isolated from the human scalp, thereby providing an accessible, autologous and safe source of stem cells for regenerative medicine that have important advantages over ES or iPS cells. J. Cell. Biochem. 107: 1016–1020, 2009. © 2009 Wiley‐Liss, Inc.     

Hair follicle-associated-pluripotent (HAP) stem cells

Various types of stem cells reside in the skin, including keratinocyte progenitor cells, melanocyte progenitor cells, skin-derived precursors (SKPs), and nestin-expressing hair follicle-associated-pluripotent (HAP) stem cells. HAP stem cells, located in the bulge area of the hair follicle, have been shown to differentiate to nerve cells, glial cells, keratinocytes, smooth muscle cells, cardiac muscle cells, and melanocytes. HAP stem cells are positive for the stem-cell marker CD34, as well as K15-negative, suggesting their relatively undifferentiated state. Therefore, HAP stem cells may be the most primitive stem cells in the skin. Moreover, HAP stem cells can regenerate the epidermis and at least parts of the hair follicle. These results suggest that HAP stem cells may be the origin of other stem cells in the skin. Transplanted HAP stem cells promote the recovery of peripheral-nerve and spinal-cord injuries and have the potential for heart regeneration as well. HAP stem cells are readily accessible from everyone, do not form tumors, and can be cryopreserved without loss of differentiation potential. These results suggest that HAP stem cells may have greater potential than iPS or ES cells for regenerative medicine.     

Comparison of label-free and GFP multiphoton imaging of hair follicle-associated pluripotent (HAP) stem cells in mouse whiskers

Hair-follicle-associated pluripotent (HAP) stem cells can differentiate into many cell types, including neurons and heart muscle cells, and have been shown to repair peripheral nerves and the spinal cord in mice. HAP stem cells can be obtained from each individual patient for regenerative medicine which overcomes problems with immune rejection. Previously, we have demonstrated that genetically-encoded protein markers such as GFP in transgenic mice can be used to visualize HAP stem cells in vivo by multiphoton tomography. Detection and visualization of stem cells in vivo without exogenous labels such as GFP would be important for human application. In the present report, we demonstrate label-free visualization of hair follicle stem cells in mouse whiskers by multiphoton tomography due to the intrinsic fluorophores such as NAD(P)H/flavins. We compared multiphoton tomography of GFP-labeled HAP stem cells and unlabeled stem cells in isolated mouse whiskers. We show that observation of HAP stem cells by label-free multiphoton tomography is comparable to detection using GFP-labeled stem cells. The results described here have important implications for detection and isolation of human HAP stem cells for regenerative medicine.     

Multipotent hair follicle stem cells promote repair of spinal cord injury and recovery of walking function

The mouse hair follicle is an easily accessible source of actively growing, pluripotent adult stem cells. C57BL transgenic mice, labeled with the fluorescent protein GFP, afforded follicle stem cells whose fate could be followed when transferred to recipient animals. These cells appear to be relatively undifferentiated since they are positive for the stem cell markers nestin and CD34 but negative for the keratinocyte marker keratin 15. These hair follicle stem cells can differentiate into neurons, glia, keratinocytes, smooth muscle cells, and melanocytes in vitro. Implanting hair follicle stem cells into the gap region of severed sciatic or tibial nerves greatly enhanced the rate of nerve regeneration and restoration of nerve function. The transplanted follicle cells transdifferentiated mostly into Schwann cells, which are known to support neuron regrowth. The treated mice regained the ability to walk essentially normally. In the present study, we severed the thoracic spinal chord of C57BL/6 immunocompetent mice and transplanted GFP-expressing hair follicle stem cells to the injury site. Most of the transplanted cells also differentiated into Schwann cells that apparently facilitated repair of the severed spinal cord. The rejoined spinal cord reestablished extensive hind-limb locomotor performance. These results suggest that hair follicle stem cells can promote the recovery of spinal cord injury. Thus, hair follicle stem cells provide an effective accessible, autologous source of stem cells for the promising treatment of peripheral nerve and spinal cord injury.     

Long-Term Extensive Ectopic Hair Growth on the Spinal Cord of Mice from Transplanted Whisker Follicles

We have previously demonstrated that hair follicles contain nestin-expressing pluripotent stem cells that can effect nerve and spinal cord repair upon transplantation. In the present study, isolated whisker follicles from nestin-driven green fluorescent protein (ND-GFP) mice were histocultured on Gelfoam for 3 weeks for the purpose of transplantation to the spinal cord to heal an induced injury. The hair shaft was cut off from Gelfoam-histocultured whisker follicles, and the remaining part of the whisker follicles containing GFP-nestin expressing pluripotent stem cells were transplanted into the injured spinal cord of nude mice, along with the Gelfoam. After 90 days, the mice were sacrificed and the spinal cord lesion was observed to have healed. ND-GFP expression was intense at the healed area of the spinal cord, as observed by fluorescence microscopy, demonstrating that the hair follicle stem cells were involved in healing the spinal cord. Unexpectedly, the transplanted whisker follicles sprouted out remarkably long hair shafts in the spinal cord during the 90 days after transplantation of Gelfoam whisker histocultures to the injured spine. The pigmented hair fibers, grown from the transplanted whisker histocultures, curved and enclosed the spinal cord. The unanticipated results demonstrate the great potential of hair growth after transplantation of Gelfoam hair follicle histocultures, even at an ectopic site.     

Extensive Hair Shaft Growth after Mouse Whisker Follicle Isolation,Cryopreservation and Transplantation in Nude Mice

We previously demonstrated that whole hair follicles could be cryopreserved to maintain their stem-cells differentation potential. In the present study, we demonstrated that cryopreserved mouse whisker hair follicles maintain their hair growth potential. DMSO better cryopreserved mouse whisker follicles compared to glycerol. Cryopreserved hair follicles also maintained the hair follicle-associated-pluripotent (HAP) stem cells, evidenced by P75^NTR expression. Subcutaneous transplantation of DMSO-cryopreserved hair follicles in nude mice resulted in extensive hair fiber growth over 8 weeks, indicating the functional recovery of hair shaft growth of cryopreserved hair follicles.     

The bulge area is the major hair follicle source of nestin-expressing pluripotent stem cells which can repair the spinal cord compared to the dermal papilla

Nestin has been shown to be expressed in the hair follicle, both in the bulge area (BA) as well as the dermal papilla (DP). Nestin-expressing stem cells of both the BA and DP have been previously shown to be pluripotent and be able to form neurons and other non-follicle cell types. The nestin-expressing pluripotent stem cells from the DP have been termed skin precursor or SKP cells. The objective of the present study was to determine the major source of nestin-expressing pluripotent stem cells in the hair follicle and to compare the ability of the nestin-expressing pluripotent stem cells from the BA and DP to repair spinal cord injury. Transgenic mice in which the nestin promoter drives GFP (ND-GFP) were used in order to observe nestin expression in the BA and DP. Nestin-expressing DP cells were found in early and middle anagen. The BA had nestin expression throughout the hair cycle and to a greater extent than the DP. The cells from both regions had very long processes extending from them as shown by two-photon confocal microscopy. Nestin-expressing stem cells from both areas differentiated into neuronal cells at high frequency in vitro. Both nestin-expressing DP and BA cells differentiated into neuronal and glial cells after transplantation to the injured spinal cord and enhanced injury repair and locomotor recovery within four weeks. Nestin-expressing pluripotent stem cells from both the BA and DP have potential for spinal cord regeneration, with the BA being the greater and more constant source.     

Nestin-Expressing Stem Cells Promote Nerve Growth in Long-Term 3-Dimensional Gelfoam?-Supported Histoculture

We have previously reported that hair follicles contain multipotent stem cells which express nestin. The nestin-expressing cells form the hair follicle sensory nerve. In vitro, the nestin-expressing hair follicle cells can differentiate into neurons, Schwann cells, and other cell types. In the present study, the sciatic nerve was excised from transgenic mice in which the nestin promoter drives green fluorescent protein (ND-GFP mice). The ND-GFP cells of the sciatic nerve were also found to be multipotent as the ND-GFP cells in the hair follicle. When the ND-GFP cells in the mouse sciatic nerve cultured on Gelfoam® and were imaged by confocal microscopy, they were observed forming fibers extending the nerve. The fibers consisted of ND-GFP-expressing spindle cells, which co-expressed the neuron marker β-III tubulin, the immature Schwann-cell marker p75^NTR and TrkB which is associated with neurons. The fibers also contain nestin-negative spherical cells expressing GFAP, a Schwann-cell marker. The β-III tubulin-positive fibers had growth cones on their tips expressing F-actin, indicating they are growing axons. When the sciatic nerve from mice ubiquitously expressing red fluorescent protein (RFP) was co-cultured on Gelfoam® with the sciatic nerve from ND-GFP transgenic mice, the interaction of nerves was observed. Proliferating nestin-expressing cells in the injured sciatic nerve were also observed in vivo. Nestin-expressing cells were also observed in posterior nerves but not in the spinal cord itself, when placed in 3-D Gelfoam® culture. The results of the present report suggest a critical function of nestin-expressing cells in peripheral nerve growth and regeneration.     

Embryonic stem cell-derived L1 overexpressing neural aggregates enhance recovery after spinal cord injury in mice

An obstacle to early stem cell transplantation into the acutely injured spinal cord is poor survival of transplanted cells. Transplantation of embryonic stem cells as substrate adherent embryonic stem cell-derived neural aggregates (SENAs) consisting mainly of neurons and radial glial cells has been shown to enhance survival of grafted cells in the injured mouse brain. In the attempt to promote the beneficial function of these SENAs, murine embryonic stem cells constitutively overexpressing the neural cell adhesion molecule L1 which favors axonal growth and survival of grafted and imperiled cells in the inhibitory environment of the adult mammalian central nervous system were differentiated into SENAs and transplanted into the spinal cord three days after compression lesion. Mice transplanted with L1 overexpressing SENAs showed improved locomotor function when compared to mice injected with wild-type SENAs. L1 overexpressing SENAs showed an increased number of surviving cells, enhanced neuronal differentiation and reduced glial differentiation after transplantation when compared to SENAs not engineered to overexpress L1. Furthermore, L1 overexpressing SENAs rescued imperiled host motoneurons and parvalbumin-positive interneurons and increased numbers of catecholaminergic nerve fibers distal to the lesion. In addition to encouraging the use of embryonic stem cells for early therapy after spinal cord injury L1 overexpression in the microenvironment of the lesioned spinal cord is a novel finding in its functions that would make it more attractive for pre-clinical studies in spinal cord regeneration and most likely other diseases of the nervous system.     

The bulge area is the origin of nestin-expressing pluripotent stem cells of the hair follicle

Nestin-expressing pluripotent stem cells have been found both in the bulge area (BA) as well as the dermal papilla (DP). Nestin-expressing stem cells of both the BA and DP have been previously shown to be able to form neurons and other non-follicle cell types. The nestin-expressing stem cells from the DP have been termed skin precursor or SKP cells. Both nestin-expressing DP and BA cells have been previously shown to effect repair of the injured spinal cord and peripheral nerve, with the BA being the greater and more constant source of the stem cells. The BA contains nestin-expressing stem cells throughout the hair cycle, whereas nestin-expressing dermal papillae stem cells were found in early and mid-anagen only. Our previous studies have shown that the nestin-expressing stem cells in the BA and DP have similar morphological features. The cells from both regions have a small body diameter of approximately 7 μm with long extrusions, as shown by 2-photon imaging. In the present study, using 2-photon imaging of whisker follicles from transgenic mice expressing nestin-driven green fluorescent protein (ND-GFP), we demonstrate that the BA is the source of the nestin-expressing stem cells of the hair follicle. The nestin-expressing stem cells migrate from the BA to the DP as well as into the surrounding skin tissues including the epidermis, and during wound healing, suggesting that the BA may be the source of the stem cells of the skin itself.     

Induced pluripotent stem cells from hair follicles as a cellular model for neurodevelopmental disorders

Disease-specific induced pluripotent stem cells (iPSC) allow unprecedented experimental platforms for basic research as well as high-throughput screening. This may be particularly relevant for neuropsychiatric disorders, in which the affected neuronal cells are not accessible. Keratinocytes isolated from hair follicles are an ideal source of patients' cells for reprogramming, due to their non-invasive accessibility and their common neuroectodermal origin with neurons, which can be important for potential epigenetic memory. From a small number of plucked human hair follicles obtained from two healthy donors we reprogrammed keratinocytes to pluripotent iPSC. We further differentiated these hair follicle-derived iPSC to neural progenitors, forebrain neurons and functional dopaminergic neurons.This study shows that human hair follicle-derived iPSC can be differentiated into various neural lineages, suggesting this experimental system as a promising in vitro model to study normal and pathological neural developments, avoiding the invasiveness of commonly used skin biopsies.     

Rapid Genetic Analysis of Epithelial-Mesenchymal Signaling During Hair Regeneration

Hair follicle morphogenesis, a complex process requiring interaction between epithelia-derived keratinocytes and the underlying mesenchyme, is an attractive model system to study organ development and tissue-specific signaling. Although hair follicle development is genetically tractable, fast and reproducible analysis of factors essential for this process remains a challenge. Here we describe a procedure to generate targeted overexpression or shRNA-mediated knockdown of factors using lentivirus in a tissue-specific manner. Using a modified version of a hair regeneration model ^{5, 6, 11}, we can achieve robust gain- or loss-of-function analysis in primary mouse keratinocytes or dermal cells to facilitate study of epithelial-mesenchymal signaling pathways that lead to hair follicle morphogenesis. We describe how to isolate fresh primary mouse keratinocytes and dermal cells, which contain dermal papilla cells and their precursors, deliver lentivirus containing either shRNA or cDNA to one of the cell populations, and combine the cells to generate fully formed hair follicles on the backs of nude mice. This approach allows analysis of tissue-specific factors required to generate hair follicles within three weeks and provides a fast and convenient companion to existing genetic models.     

Real-time confocal imaging of trafficking of nestin-expressing multipotent stem cells in mouse whiskers in long-term 3-D histoculture

We have previously demonstrated that nestin-expressing multipotent hair follicle stem cells are located above the hair follicle bulge and can differentiate into neurons and other cell types in vitro. The nestin-expressing hair follicle stem cells promoted the recovery of pre-existing axons when they were transplanted to the severed sciatic nerve or injured spinal cord. We have also previously demonstrated that the whisker hair follicle contains nestin-expressing stem cells in the dermal papilla (DP) as well as in the bulge area (BA), but that their origin is in the BA. In the present study, we established the technique of long-term Gelfoam? histoculture of whiskers isolated from transgenic mice in which nestin drives green fluorescent protein (ND-GFP). Confocal imaging was used to monitor ND-GFP-expressing stem cells trafficking in real time between the BA and DP to determine the fate of the stem cells. It was observed over a 2-week period that the stem cells trafficked from the BA toward the DP area and extensively grew out onto Gelfoam? forming nerve-like structures. This new method of long-term histoculture of whiskers from ND-GFP mice will enable the extensive study of the behavior of nestin-expressing multipotent stem cells of the hair follicle.     

Aging of hair follicle stem cells and their niches

Hair follicles in the skin undergo cyclic rounds of regeneration, degeneration, and rest throughout life. Stem cells residing in hair follicles play a pivotal role in maintaining tissue homeostasis and hair growth cycles. Research on hair follicle aging and age-related hair loss has demonstrated that a decline in hair follicle stem cell (HFSC) activity with aging can decrease the regeneration capacity of hair follicles. This review summarizes our understanding of how age-associated HFSC intrinsic and extrinsic mechanisms can induce HFSC aging and hair loss. In addition, we discuss approaches developed to attenuate age-associated changes in HFSCs and their niches, thereby promoting hair regrowth.