Outer Root Sheath Of Hair Follicles Research Articles

In vitro hair growth-promoting effect of Lgr5-binding octapeptide in human primary hair cells.

Hair loss occurs due to various biological and environmental causes, which can have psychosocial consequences. The Wnt/β-catenin signaling is well-known for its role in hair growth and regeneration, as it induces the proliferation and differentiation of hair cells. When the leucine-rich G protein-coupled receptor 5 (Lgr5) interacts with the R-spondins, the frizzled receptor (FZD), a Wnt receptor, becomes stabilized, resulting in an increased β-catenin activity. We investigated whether the octapeptide that binds to Lgr5 enhances proliferation and differentiation of human primary hair cells through the activation of Wnt/β-catenin signaling. The binding affinity of the octapeptide to Lgr5 was evaluated using surface plasmon resonance (SPR). We confirmed changes in proliferation and related factors like β-catenin activation and growth factors (GFs) expression in human hair follicle dermal papilla cells (HHFDPCs). Additionally, we observed the proliferation and the expression of differentiation markers in human hair follicle outer root sheath cells (HHFORSCs), human hair follicle germinal matrix cells (HHFGMCs), and human hair follicle stem cells (HHFSCs). We used three-dimensional HHFDPC spheroid culture treated with dihydrotestosterone (DHT) to create invitro conditions that mimic androgenetic alopecia, and we studied the effects of octapeptide on Wnt expression and HHFSC differentiation. The binding of the octapeptide to Lgr5 was confirmed using SPR analysis. In HHFDPCs, treatment with octapeptide resulted in a concentration-dependent increase in proliferation. We also observed increased nuclear translocation of β-catenin and increased expression of its downstream targets. HHFDPCs treated with octapeptide exhibited increased expression of growth factors and phosphorylation of Akt and ERK. In addition, we confirmed that octapeptide increased proliferation and induced differentiation in HHFORSCs, HHFGMCs, and HHFSCs. Under the HHFDPC spheroid culture conditions, we found that octapeptide restored the inhibition of Wnt-5a and Wnt-10b expressions by DHT. In HHFSCs treated with HHFDPC spheroid culture media, we observed that octapeptide recovered the inhibition of differentiation by DHT. We found that octapeptides activated the Wnt/β-catenin signaling and induced the proliferation and differentiation of human primary hair cells by acting as an exogenous ligand for Lgr5. In addition, octapeptides recovered inhibited hair regeneration characters by DHT in androgenetic alopecia-mimic invitro model. These findings suggest that octapeptides may be a promising therapeutic option for treating hair loss.

Mechanical stimulation of human hair follicle outer root sheath cultures activates adjacent sensory neurons.

Mechanical stimuli, such as stroking or pressing on the skin, activate mechanoreceptors transmitting information to the sensory nervous system and brain. It is well accepted that deflection of the hair fiber that occurs with a light breeze or touch directly activates the sensory neurons surrounding the hair follicle, facilitating transmission of mechanical information. Here, we hypothesized that hair follicle outer root sheath cells act as transducers of mechanical stimuli to sensory neurons surrounding the hair follicle. Using electrochemical analysis on human hair follicle preparations in vitro, we were able to show that outer root sheath cells release ATP and the neurotransmitters serotonin and histamine in response to mechanical stimulation. Using calcium imaging combined with pharmacology in a coculture of outer root sheath cells with sensory neurons, we found that the release of these three molecules from hair follicle cells leads to activation of sensory neurons.

Mesenchymal Stem Cells Isolated from Equine Hair Follicles Using a Method of Air-Liquid Interface

Equine mesenchymal stem cells (MSC) of various origins have been identified in horses, including MSCs from the bone marrow and adipose tissue. However, these stem cell sources are highly invasive in sampling, which thereby limits their clinical application in equine veterinary medicine. This study presents a novel method using an air-liquid interface to isolate stem cells from the hair follicle outer root sheath of the equine forehead skin. These stem cells cultured herewith showed high proliferation and asumed MSC phenotype by expressing MSC positive biomarkers (CD29, CD44 CD90) while not expressing negative markers (CD14, CD34 and CD45). They were capable of differentiating towards chondrogenic, osteogenic and adipogenic lineages, which was comparable with MSCs from adipose tissue. Due to their proliferative phenotype in vitro, MSC-like profile and differentiation capacities, we named them equine mesenchymal stem cells from the hair follicle outer root sheath (eMSCORS). eMSCORS present a promising alternative stem cell source for the equine veterinary medicine.Graphical abstract

SIRT1 downregulation provokes immune-inflammatory responses in hair follicle outer root sheath cells and may contribute to development of alopecia areata

BackgroundSilent information regulator 1 (SIRT1), a type III histone deacetylase, is involved in various cutaneous and systemic autoimmune diseases including systemic lupus erythematosus, rheumatoid arthritis, and psoriasis. However, little is known about the role of SIRT1 in the development of alopecia areata (AA). ObjectivesThis study investigated whether SIRT1 regulates the hair follicle immune system and is involved in AA pathogenesis. MethodsSIRT1 expression in human scalp tissue was analyzed using immunohistochemical staining, qPCR, and western blotting. The regulatory effect of SIRT1 was evaluated after stimulation with the double-stranded RNA mimic polyinosinic:polycytidylic acid (poly I:C) in hair follicle outer root sheath (ORS) cells and C3H/HeJ mice. ResultsSIRT1 expression was significantly reduced in the AA scalp compared to the normal scalp. SIRT1 inhibition upregulated MHC class I polypeptide-related sequence A and UL16 binding protein 3 in hair follicle ORS cells. SIRT1 inhibition also promoted the production of Th1 cytokines (IFN-γ and TNF-α), IFN-inducible chemokines (CXCL9 and CXCL10), and T cell migration in ORS cells. Conversely, SIRT1 activation suppressed the autoreactive inflammatory responses. The counteractive effect of the immune response by SIRT1 was mediated through the deacetylation of NF-κB and phosphorylation of STAT3. ConclusionSIRT1 downregulation induces immune-inflammatory responses in hair follicle ORS cells and may contribute to AA development.

IL-33 Contributes to the Pathological Changes of Hair Follicles in Psoriasis: A Potential Target for Psoriatic Alopecia.

IL-33 is constitutively expressed in skin tissues. Alopecia, a T cells-driven disorder of the hair follicles (HFs), is a common complication in the development of psoriasis. However, the role of IL-33 in psoriatic alopecia remains uncovered. Here, we investigated the roles of IL-33 in inducing pathological changes of hair follicles in psoriasis. Clinical samples and imiquimod (IMQ)-induced psoriatic mice samples were used to investigate the pathological changes and T-cell infiltration of HFs. By using immunohistochemistry staining, the distribution and expression alteration of IL-33 in HFs were determined. Next, by using IL-33 and ST2 knockout mice, we investigated the role of IL-33/ST2 axis in the pathological changes of HFs in psoriasis. Meanwhile, recombinant IL-33 protein was subcutaneous injected to confirm its effect. Finally, RNA sequencing was used to clarify the genes and signaling pathways that involved in this process. Differentially expressed genes were further verified by RT-PCR in cultured HFs in vitro. We found that the pathological changes of HFs and T cells infiltration in imiquimod-induced psoriatic mice were similar to that in psoriasis patients. The IL-33 positive keratinocytes in the outer root sheath of HFs were increased in both psoriasis patients and psoriatic model mice compared with the controls. By using gene knockout mice, we found that the pathological changes and T cell infiltration were attenuated in IL-33-/- and ST2-/- psoriatic model mice. In addition, subcutaneous injection of recombinant IL-33 exacerbated the pathological changes of HFs and T cell infiltration. RNA sequencing and RT-RCR revealed that IL-33 upregulated the transcription of genes related to keratinocytes proliferation and T lymphocytes chemotaxis. Our study identifies that IL-33 promotes the pathological changes of HFs in psoriasis, which contributes to psoriatic alopecia. Inhibition of IL-33 may be a potential therapeutic approach for psoriatic alopecia.

The regulation mechanism of different hair types in inner Mongolia cashmere goat based on PI3K-AKT pathway and FGF21.

There is genetic diversity of hair types in the Inner Mongolia cashmere goat population. Previous studies have found that fibroblast growth factor 21 (FGF21) and PI3K-AKT signal pathways may be related to different hair types in Inner Mongolia cashmere goats. Therefore, the purpose of this study was to explore the effects of the PI3K-AKT signal pathway on different hair types, the expression of mRNA and protein expression sites of FGF21 in the hair follicles of cashmere goats with different hair types, so as to lay a foundation for understanding the molecular mechanism of different hair types and the role of skin hair follicle development. In this experiment, the skin tissues of long hair type (LHG) and short hair type (SHG) of Inner Mongolia cashmere goat were collected in three key periods of secondary hair follicle growth, namely, anagen (September), catagen (December), and telogen (March). The relative expression of FGF21 and PI3K-AKT signal pathway candidate gene mRNA in different periods and different hair types was detected by real-time fluorescence quantitative technique (qRT-PCR), and the expression site of FGF21 protein was located by immunohistochemical technique. Through qRT-PCR, it was found that the relative expression of FGF21, FGFR1, AKT3, BRCA1, PKN3, SPP1, and GNG4 was significantly different between LHG and SHG. The expression of FGF21 in the skin of LHG was significantly higher than that of SHG in the three periods. Through immunohistochemical test, it was found that FGF21 protein was mainly expressed in primary hair follicle connective tissue sheath, primary hair follicle outer root sheath, secondary hair follicle outer root sheath, and sebaceous glands. It was also found that the expression of LHG skin tissue in the outer root sheath of primary hair follicles was higher than that of SHG in three periods. In summary, it is suggested that the PI3K-AKT signal pathway may play an important role in the formation of different hair types in Inner Mongolia cashmere goats.

Laminin N-terminus α31 expression during development is lethal and causes widespread tissue-specific defects in a transgenic mouse model.

Laminins (LMs) are essential components of all basement membranes where they regulate an extensive array of tissue functions. Alternative splicing from the laminin α3 gene produces a non‐laminin but netrin‐like protein, Laminin N terminus α31 (LaNt α31). LaNt α31 is widely expressed in intact tissue and is upregulated in epithelial cancers and during wound healing. In vitro functional studies have shown that LaNt α31 can influence numerous aspects of epithelial cell behavior via modifying matrix organization, suggesting a new model of laminin auto‐regulation. However, the function of this protein has not been established in vivo. Here, a mouse transgenic line was generated using the ubiquitin C promoter to drive inducible expression of LaNt α31. When expression was induced at embryonic day 15.5, LaNt α31 transgenic animals were not viable at birth, exhibiting localized regions of erythema. Histologically, the most striking defect was widespread evidence of extravascular bleeding across multiple tissues. Additionally, LaNt α31 transgene expressing animals exhibited kidney epithelial detachment, tubular dilation, disruption of the epidermal basal cell layer and of the hair follicle outer root sheath, and ~50% reduction of cell numbers in the liver, associated with depletion of hematopoietic erythrocytic foci. These findings provide the first in vivo evidence that LaNt α31 can influence tissue morphogenesis.

Mesenchymal Stem Cells From Mouse Hair Follicles Reduce Hypertrophic Scarring in a Murine Wound Healing Model

Wound healing of acute full-thickness injuries and chronic non-healing ulcers leads to delayed wound closure, prolonged recovery period and hypertrophic scarring, generating a demand for an autologous cell therapy and a relevant pre-clinical research models for wound healing. In this study, an immunocompetent model for wound healing was employed using a syngeneic murine cell line of mesenchymal stem cells cultured from the mouse whisker hair follicle outer root sheath (named moMSCORS). moMSCORS were isolated using an air-liquid interface method, expanded in vitro and characterized according to the MSC definition criteria - cell viability, in vitro proliferation, MSC phenotype and multi-lineage differentiations. Moreover, upon applying moMSCORS in an in vivo full-thickness wound model in the syngeneic C57BL/6 mice, the treated wounds displayed different morphology to that of the untreated wound beds. Quantitative evaluation of angiogenesis, granulation and wound closure involving clinical scoring and software-based quantification indicated a lower degree of inflammation in the treated wounds. Histological staining of treated wounds by the means of H&E, Alcian Blue, PicroSirius Red and αSMA immune labelling showed lower cellularity, less collagen filaments as well as thinner dermal and epidermal layers compared with the untreated wounds, indicating a general reduction of hypertrophic scars. The decreased inflammation, accelerated wound closure and non-hypertrophic scarring, which were facilitated by moMSCORS, hereby address a common problem of hypertrophic scars and non-physiological tissue properties upon wound closure, and additionally offer an in vivo model for the autologous cell-based wound healing.Graphic

Osteogenic Potential of Mesenchymal Stem Cells from Adipose Tissue, Bone Marrow and Hair Follicle Outer Root Sheath in a 3D Crosslinked Gelatin-Based Hydrogel.

Bone transplantation is regarded as the preferred therapy to treat a variety of bone defects. Autologous bone tissue is often lacking at the source, and the mesenchymal stem cells (MSCs) responsible for bone repair mechanisms are extracted by invasive procedures. This study explores the potential of autologous mesenchymal stem cells derived from the hair follicle outer root sheath (MSCORS). We demonstrated that MSCORS have a remarkable capacity to differentiate in vitro towards the osteogenic lineage. Indeed, when combined with a novel gelatin-based hydrogel called Osteogel, they provided additional osteoinductive cues in vitro that may pave the way for future application in bone regeneration. MSCORS were also compared to MSCs from adipose tissue (ADMSC) and bone marrow (BMMSC) in a 3D Osteogel model. We analyzed gel plasticity, cell phenotype, cell viability, and differentiation capacity towards the osteogenic lineage by measuring alkaline phosphatase (ALP) activity, calcium deposition, and specific gene expression. The novel injectable hydrogel filled an irregularly shaped lesion in a porcine wound model displaying high plasticity. MSCORS in Osteogel showed a higher osteo-commitment in terms of calcium deposition and expression dynamics of OCN, BMP2, and PPARG when compared to ADMSC and BMMSC, whilst displaying comparable cell viability and ALP activity. In conclusion, autologous MSCORS combined with our novel gelatin-based hydrogel displayed a high capacity for differentiation towards the osteogenic lineage and are acquired by non-invasive procedures, therefore qualifying as a suitable and expandable novel approach in the field of bone regeneration therapy.

The Angiogenic Potential of Mesenchymal Stem Cells from the Hair Follicle Outer Root Sheath.

Neovascularization is regarded as a pre-requisite in successful tissue grafting of both hard and soft tissues alike. This study considers mesenchymal stem cells from hair follicle outer root sheath (MSCORS) as powerful tools with a neat angiogenic potential that could in the future have wide scopes of neo-angiogenesis and tissue engineering. Autologous MSCORS were obtained ex vivo by non-invasive plucking of hair and they were differentiated in vitro into both endothelial cells and vascular smooth muscle cells (SMCs), two crucial cellular components of vascular grafts. Assessment was carried out by immunostaining, confocal laser-scanning microscopy, gene expression analysis (qRT-PCR), quantitative analysis of anastomotic network parameters, and cumulative length quantification of immunostained α-smooth muscle actin-containing stress fibers (α -SMA). In comparison to adipose mesenchymal stem cells, MSCORS exhibited a significantly higher differentiation efficiency according to key quantitative criteria and their endothelial derivatives demonstrated a higher angiogenic potential. Furthermore, the cells were capable of depositing their own extracellular matrix in vitro in the form of a membrane-cell sheet, serving as a base for viable co-culture of endothelial cells and SMCs integrated with their autologous matrix. Differentiated MSCORS hereby provided a complex autologous cell-matrix construct that demonstrates vascularization capacity and can serve as a base for personalized repair grafting applications.

The Middle Part of the Plucked Hair Follicle Outer Root Sheath Is Identified as an Area Rich in Lineage-Specific Stem Cell Markers.

Hair follicle outer root sheath (ORS) is a putative source of stem cells with therapeutic capacity. ORS contains several multipotent stem cell populations, primarily in the distal compartment of the bulge region. However, the bulge is routinely obtained using invasive isolation methods, which require human scalp tissue ex vivo. Non-invasive sampling has been standardized by means of the plucking procedure, enabling to reproducibly obtain the mid-ORS part. The mid-ORS shows potential for giving rise to multiple stem cell populations in vitro. To demonstrate the phenotypic features of distal, middle, and proximal ORS parts, gene and protein expression profiles were studied in physically separated portions. The mid-part of the ORS showed a comparable or higher NGFR, nestin/NES, CD34, CD73, CD44, CD133, CK5, PAX3, MITF, and PMEL expression on both protein and gene levels, when compared to the distal ORS part. Distinct subpopulations of cells exhibiting small and round morphology were characterized with flow cytometry as simultaneously expressing CD73/CD271, CD49f/CD105, nestin, and not CK10. Potentially, these distinct subpopulations can give rise to cultured neuroectodermal and mesenchymal stem cell populations in vitro. In conclusion, the mid part of the ORS holds the potential for yielding multiple stem cells, in particular mesenchymal stem cells.

Culturing of Melanocytes from the Equine Hair Follicle Outer Root Sheath

Hair follicles harbor a heterogeneous regenerative cell pool and represent a putative low-to-non-invasively available source of stem cells. We previously reported a technology for culturing human melanocytes from the hair follicle outer root sheath (ORS) for autologous pigmentation of tissue engineered skin equivalents. This study translated the ORS technology to horses. We de-veloped a culture of equine melanocytes from the ORS (eMORS) from equine forelock hair follicles cultured by means of an analogue human hair follicle-based in vitro methodology. The procedure was adjusted to equine physiology by addition of equine serum to the culture medium. The hair follicles were isolated by macerating forelock skin rests, enzymatically digested and subjected to air-medium-interface cultivation method. The procedure resulted in differentiated equine melanocytes, which exhibited typical morphology, presence of melanosomes, expression of cytoskeleton proteins vimentin, α-SMA, Sox2, S100ß and tyrosinase as well as tyrosinase activity followed by production of melanin. According to all assessed parameters, eMORS could be ranked as partially melanotic melanocytes. The results of the study offer an experimental base for further insight into hair follicle biology in equine and for comparative studies of hair follicles across different species.

Generation of Epidermal Equivalents from Hair Follicle Melanocytes, Keratinocytes, and Dermal Fibroblasts.

Bench-to-bedside axis of therapeutic product development is currently being oriented towards minimum invasiveness on both ends-not only clinical application but harvesting of the starting biological material as well. This is particularly relevant for Advanced Therapy Medicinal Products and their specific legislative requirements, even more so in skin regeneration. It is precisely the skin equivalents and grafts that benefit from the minimum-to-noninvasive approach to a noteworthy extent, taking in account the sensitive nature of both skin harvesting and grafting.This chapter includes protocols for two separate steps of generating skin equivalent from the cells cultured from hair follicle outer root sheath. The first step is a non-pigmented epidermal equivalent generated from human keratinocytes from the outer root sheath named non-pigmented epidermal graft. The second step consists of co-cultivating human keratinocytes and human melanocytes from the outer root sheath, hereby producing a pigmented epidermal graft.

ERK1/2 Pathway Is Involved in the Enhancement of Fatty Acids from Phaeodactylum tricornutum Extract (PTE) on Hair Follicle Cell Proliferation.

Extractions from Phaeodactylum tricornutum have been widely studied and evaluated to various biological effects. The aim of this study was to investigate the promotional effect of P. tricornutum extract (PTE) on the ERK1/2 signaling pathway involved in hair follicle cell proliferation. In order to illuminate the enhancement of PTE on hair growth by promoting proliferation of hair follicle cells, the activities of human hair follicle outer root sheath cell (HFORSC), human hair follicle germinal matrix cells (HFGMC), and hair epithelial melanocytes (HEM) were observed under PET treatment. Levels of keratins, PKCζ, ERK1/2, and p38 MAPK in hair follicle cells were determined by Western blotting to illustrate the mechanisms of PTE effects on hair growth. Analyzed by GC-MS, the main polyunsaturated fatty acids which were 9.43% of total fatty acids in PTE were linolenic acid, linoleic acid, eicosapentaenoic acid, and docosahexaenoic acid. Melanin content and tyrosinase activity in HEM were measured. The results showed that PTE exhibited remarkable enhancement on cell proliferation. Melanin production was inhibited by PTE treatment, while keratin-14, keratin-15, and keratin-17 levels on hair follicle cells were elevated at different concentrations. The promotions of ERK1/2 and p38 MAPK levels indicated that the ERK1/2 signaling pathway is involved in the proliferation of hair follicle cells. These results are the evidence that PTE potentially deserves further study as a new natural candidate for hair care applications.

Correction to "Pharmacological Activation of Thermo-Transient Receptor Potential Vanilloid 3 Channels Inhibits Hair Growth by Inducing Cell Death of Hair Follicle Outer Root Sheath".

Correction to "Pharmacological Activation of Thermo-Transient Receptor Potential Vanilloid 3 Channels Inhibits Hair Growth by Inducing Cell Death of Hair Follicle Outer Root Sheath".

Treatment of stable vitiligo using cultured and non-cultured autologous melanocytes from hair follicle outer root sheath cell suspension

Background Several surgical treatment methods have been proposed for the treatment of stable vitiligo, but they remain inadequate in terms of patient satisfaction. Objective To assess the efficacy and safety of a modified autologous cultured hair follicle outer root sheath (HF-ORS) cell suspension transplantation in the treatment of stable vitiligo lesions, and to compare it with transplantation of autologous non-cultured HF-ORS cell suspension for the same patient. Patients and methods Twenty-four patients were enrolled in this prospective controlled study. Hairs were epilated, and HF-ORS cell suspensions were prepared using two methods for the same patient. The recipient site was prepared by CO2 laser resurfacing. Assessment of cellular activity and viability included cellular melanin content determined by enzyme-linked immunosorbent assay and measurement of the fold change in premelanosome (Pmel-17) gene expression. The patients were followed-up for 6 months postprocedure. Results Nineteen patients completed the study. The results demonstrated that the cultured method showed superior repigmentation response in 15 (79%) patients, three (16%) responded equally, and one (5%) was more tolerant of the noncultured method. Conclusion Cultured autologous HF-ORS melanocytes show superior repigmentation response, increased melanin content, and upregulated gene expression of Pmel-17 than that of noncultured ones. However, the study also demonstrated that both methods are potential options for the treatment of stable vitiligo.

A Prospective Observational Comparative Study of Novel Autologous Negative Pressure Epidermal Harvesting System (ANPEHS or EHS) and Suction Blister Grafting (SBG) in Treatment of Stable Vitiligo.

Background:Vitiligo surgery has evolved a long way from punch skin grafts to the epidermal cell suspension and latest to the extracted hair follicle outer root sheath cell suspension transplantation (EHFORSCS). In the latest development, the novel technique autologous negative pressure epidermal harvesting system (ANPEHS) seems to be a good addition to the prevailing therapy in vitiligo.Aims and Objectives:The aim of this work was to study and compare novel ANPEHS and suction blister grafting (SBG) in the treatment of stable vitiligo.Materials and Methods:This was a prospective, single-centered, observational, open-labeled comparative study of the rate and final extent of repigmentation in ANPEHS or EHS and SBG in the management of stable vitiligo. The patients in this study were drawn from the outpatient Department of Dermatology of a Tertiary Care Hospital of the Armed Forces from July 1 2015 to December 31, 2016. A total of 40 patients with at least two comparable depigmented patches of 6 months’ stability were recruited. Both procedures were performed in the same patient on the same day. Informed consents were taken from all the patients. Each patient in the study had undergone a detailed clinical, general physical, systemic, and a dermatological examination. Clinical photographs had been taken before and after grafting, monthly for the first 6 months then bimonthly for the next 6 months. Donor site was preferably inner aspect of thigh.Statistical Analysis Used:“Chi-square test” and “statistical significance” (P value) methods.Results:Of total 80 patches, excellent results were seen in 82.9% patches by using the EHS method and 80% excellent results by using the SBG method. Similarly, very good results were seen in 2.9% patches and good results in 5.7% patches by using both methods. 11.4% patches showed poor results by using the SBG method and 8.6% patches showed poor results by using the EHS method.Conclusions:The EHS method is a simple, painless, less time-consuming, expensive but effective technique to produce homogeneous repigmentation without any donor site anesthesia and complication.

CDK4/6 inhibition mitigates stem cell damage in a novel model for taxane‐induced alopecia

Taxanes are a leading cause of severe and often permanent chemotherapy‐induced alopecia. As the underlying pathobiology of taxane chemotherapy‐induced alopecia remains poorly understood, we investigated how paclitaxel and docetaxel damage human scalp hair follicles in a clinically relevant ex vivo organ culture model. Paclitaxel and docetaxel induced massive mitotic defects and apoptosis in transit amplifying hair matrix keratinocytes and within epithelial stem/progenitor cell‐rich outer root sheath compartments, including within Keratin 15+ cell populations, thus implicating direct damage to stem/progenitor cells as an explanation for the severity and permanence of taxane chemotherapy‐induced alopecia. Moreover, by administering the CDK4/6 inhibitor palbociclib, we show that transit amplifying and stem/progenitor cells can be protected from paclitaxel cytotoxicity through G1 arrest, without premature catagen induction and additional hair follicle damage. Thus, the current study elucidates the pathobiology of taxane chemotherapy‐induced alopecia, highlights the paramount importance of epithelial stem/progenitor cell‐protective therapy in taxane‐based oncotherapy, and provides preclinical proof‐of‐principle in a healthy human (mini‐) organ that G1 arrest therapy can limit taxane‐induced tissue damage.

Enhanced Cultivation of Hair Follicle Outer Root Sheath Cells with K + M Culture Medium

Hair follicle stem cells are vital seed cells for hair follicle and tissue engineering skin. However, there is no valid approach to obtain abundant cells within short times in vitro. The purpose of this study was to establish a new medium for culture and expansion of hair follicle outer root sheath (ORS) cells. ORS cells were harvested from human samples and cultured with a medium mixed with keratinocyte serum free medium and mouse embryonic fibroblasts suspension. The proliferation and vitality of ORS cells were evaluated. The cells exhibited typical cobblestone morphology with good adhesion and colony-forming ability. The cells were also characterized through detecting specific markers with immunofluorescent staining and the results showed that partial ORS cells could express hair follicle stem cell markers of CK15, CK19 and FST. At last, ORS cells and fibroblasts were used as seed cells and preliminarily constructed tissue engineering skin successfully. In conclusion, these findings suggested that this new medium could effectively accelerate cell proliferation and maintain the fundamental characteristics of ORS cells.

Pharmacological Activation of Thermo-Transient Receptor Potential Vanilloid 3 Channels Inhibits Hair Growth by Inducing Cell Death of Hair Follicle Outer Root Sheath.

Hair growth starts from hair follicles that reside in dermis, and abnormal hair growth is an early sign of hair follicle disease or systemic illness such as alopecia or hair loss. Therefore, identifying a target critical for dysfunctional hair follicles is fundamental to alleviating dermatologic or systemic diseases with hair abnormalities. The warm temperature-activated Ca2+-permeable transient receptor potential vanilloid 3 (TRPV3) channel protein is abundantly expressed in the skin keratinocytes, and dysfunctional TRPV3 causes human congenital Olmsted syndrome, characterized by skin diseases and alopecia, indicating an important role for TRPV3 in hair follicle development and hair growth. To validate TRPV3 as a therapeutic target, we investigated the impact of pharmacological modulation of TRPV3 on hair growth using a combination of biochemical and cell biology, immunohistochemical, whole-cell patch clamp, RNA interference, and pharmacological approaches. We found that functional TRPV3 channel proteins are highly expressed in hair follicle outer root sheath (ORS) cells as detected by Western blot analysis, immunohistochemical staining, and electrophysiological techniques. Pharmacological activation of TRPV3 by agonist natural carvacrol induces cell death of ORS cells, and topical application of carvacrol to mouse dorsal skin also inhibits hair growth. Conversely, specific inhibition of TRPV3 by inhibitor natural forsythoside B and short-hairpin RNA reverses the cell death induced by carvacrol-mediated TRPV3 activation in human ORS cells. Furthermore, forsythoside B results in a significant reversal of hair growth inhibition induced by agonist carvacrol. Altogether, our findings demonstrate that TRPV3 channel is critical for regulation of hair growth, and inhibition of TRPV3 may represent a promising therapy for hair loss or hair follicle-related skin diseases.