Comparison of responses of melanocyte lineages from p75(+) and p75(-) human scalp-derived neural crest stem cells under phototherapy.

Phototherapy is an effective therapeutic option in the treatment of vitiligo; however, responses varied among the different types. The underlying mechanism has scarcely been investigated. To investigate and compare the effects of phototherapy on the mutation of melanocyte lineage differentiated from human scalp-derived neural crest stem cells (HS-NCSCs) with p75 neurotrophin receptor expression positive and p75 neurotrophin receptor expression negative group in vitro, the HS-NCSCs were isolated from fetal scalp tissue, which is identified by immunofluorescent staining. The p75(+) and p75(-) cells from HS-NCSCs were isolated by magnetic cell sorting, respectively. The embryonic neural crest stem cell biomarkers were detected by RT-PCR. Narrow-band UVB (NB-UVB) was used to irradiate the cells. Cell proliferation was evaluated by cell count. Tyrosinase, Tyrp1, and Tyrp2 gene expression were measured by quantitative RT-PCR. Tyrosinase and GRCR protein levels were investigated by Western blot analysis. The electrophoretic strip showed that Sox2, Oct4, Sox10, and Nestin of p75(+) HS-NCSCs were brighter than the p75(-) HS-NCSCs. After the same dose radiation with NB-UVB, the cell proliferation of p75(+) group showed less inhibitory rate compared with the p75(-) HS-NCSCs. The tyrosinase mRNA and protein expression of differentiated melanocytes increased significantly in the group of p75(+) HS-NCSCs compared with the p75(-) group. The melanocytic mutation of p75(+) HS-NCSCs increased significantly compared with the p75(-) HS-NCSCs under NB-UVB, which indicated there were more melanocyte precursors in the differentiated cells from p75(+) HS-NCSCs. This may provide new insights for the different repigmentation efficacy of segmental and non-segmental vitiligo.

Inhibition of Shh Signaling through MAPK Activation Controls Chemotherapy-Induced Alopecia.

Chemotherapy-induced hair loss (alopecia) (CIA) remains a major unsolved problem in clinical oncology. CIA is often considered to be a consequence of the antimitotic and apoptosis-promoting properties of chemotherapy drugs acting on rapidly proliferating hair matrix keratinocytes. Here, we show that in a mouse model of CIA, the downregulation of Shh signaling in the hair matrix is a critical early event. Inhibition of Shh signaling recapitulated key morphological and functional features of CIA, whereas recombinant Shh protein partially rescued hair loss. Phosphoproteomics analysis revealed that activation of the MAPK pathway is a key upstream event, which can be further manipulated to rescue CIA. Finally, in organ-cultured human scalp hair follicles as well as in patients undergoing chemotherapy, reduced expression of SHH gene correlates with chemotherapy-induced hair follicle damage or the degree of CIA, respectively. Our work revealed that Shh signaling is an evolutionarily conserved key target in CIA pathobiology. Specifically targeting the intrafollicular MAPK-Shh axis may provide a promising strategy to manage CIA.

Pre-aggregation of scalp progenitor dermal and epidermal stem cells activates the WNT pathway and promotes hair follicle formation in in vitro and in vivo systems.

BACKGROUND: Billions of dollars are invested annually by pharmaceutical companies in search of new options for treating hair loss conditions; nevertheless, the challenge remains. One major limitation to hair follicle research is the lack of effective and efficient drug screening systems using human cells. Organoids, three-dimensional in vitro structures derived from stem cells, provide new opportunities for studying organ development, tissue regeneration, and disease pathogenesis. The present study focuses on the formation of human hair follicle organoids. METHODS: Scalp-derived dermal progenitor cells mixed with foreskin-derived epidermal stem cells at a 2:1 ratio aggregated in suspension to form hair follicle-like organoids, which were confirmed by immunostaining of hair follicle markers and by molecular dye labeling assays to analyze dermal and epidermal cell organization in those organoids. The hair-forming potential of organoids was examined using an in vivo transplantation assay. RESULTS: Pre-aggregation of dermal and epidermal cells enhanced hair follicle formation in vivo. In vitro pre-aggregation initiated the interactions of epidermal and dermal progenitor cells resulting in activation of the WNT pathway and the formation of pear-shape structures, named type I aggregates. Cell-tracing analysis showed that the dermal and epidermal cells self-assembled into distinct epidermal and dermal compartments. Histologically, the type I aggregates expressed early hair follicle markers, suggesting the hair peg-like phase of hair follicle morphogenesis. The addition of recombinant WNT3a protein to the medium enhanced the formation of these aggregates, and the Wnt effect could be blocked by the WNT inhibitor, IWP2. CONCLUSIONS: In summary, our system supports the rapid formation of a large number of hair follicle organoids (type I aggregates). This system provides a platform for studying epithelial-mesenchymal interactions, for assessing inductive hair stem cells and for screening compounds that support hair follicle regeneration.

The peptide AC 2 isolated from Bacillus-treated Trapa japonica fruit extract rescues DHT (dihydrotestosterone)-treated human dermal papilla cells and mediates mTORC1 signaling for autophagy and apoptosis suppression.

The Trapa japonica fruit is a natural plant growing in ponds with its roots in the mud. It has long been used as a home remedy for many diseases; however, a major problem with this kind of natural extract is the multicomponents-multitargets for diseases. Such problems make it difficult to identify the mechanism of action. Another problem is quality control and consistency. The aim of this research was to isolate a single bioactive compound (peptide) derived from the Trapa japonica fruit. The research was conducted with various experimental techniques, such as fermentation and liquid chromatography, to isolate a peptide. We isolated the AC 2 peptide from Trapa japonica fruit and found it to be promising on human dermal papilla cells. Dihydrotestosterone (DHT) stresses human dermal papilla cells and is a major cause of hair loss resulting from hormones and environmental factors. The purpose of this research was to develop an understanding of the mechanism by which the AC 2 peptide rescues dihydrotestosterone (DHT)-treated human dermal papilla cells. We explored the effects of the AC 2 peptide on the cell biological functions of human dermal papilla cells (HDPs). HDPs were treated with the AC 2 peptide and DHT. Then, a cytotoxicity assay, flow cytometry, Western blot, immunoprecipitation, and 3D cell culture for immunohistochemistry were conducted to investigate the mTORC1 pathway and suppression of autophagy and apoptosis. In addition, we also synthesized the AC2 peptide as an alternative to the expensive and difficult isolation and purification procedures and confirmed its potential in biomedical applications. We also validated the effects of the synthetic AC2 peptide as well as the isolated and purified AC2 peptide and established their similarity. Although extensive research has been carried out on natural extracts, few single studies have isolated and separated a bioactive peptide (single compound).

VEGFR-2 Is in a State of Activation in Hair Follicles, Sebaceous Glands, Eccrine Sweat Glands, and Epidermis from Human Scalp: An In Situ Immunohistochemistry Study of Phosphorylated VEGFR-2.

BACKGROUND Recent research reports that VEGFR-2 is expressed in the whole hair follicle, sebaceous glands, eccrine sweat glands, and epidermis. However, phosphorylated VEGFR-2 was not found, and it could not be ascertained whether the activated form of VEGFR-2 actually participates in the biological control of epidermal appendages. In this study we aimed to determine whether the VEGFR-2 pathway is directly involved in the daily regulation of epidermal appendages biology. MATERIAL AND METHODS In this study, we investigated the expression of phosphorylation of VEGFR-2 by immunohistochemical analysis in the epidermis and epidermal appendages in normal human scalp skin. RESULTS Immunohistochemical analysis revealed phosphorylation of VEGFR-2 in a whole hair follicle, mainly in the infundibulum basal layer, hair cortex, and medulla in the isthmus, and matrix in the hair bulb. Phosphorylated VEGFR-2 also was found in the sebaceous glands, eccrine sweat glands, and epidermis. CONCLUSIONS Therefore, we suggest that VEGFR-2 activation is involved in routine regulation of human epidermal appendages.

Chondrogenic differentiation of human scalp adipose-derived stem cells in Polycaprolactone scaffold and using Freeze Thaw Freeze method.

Human adipose tissue has been identified as a viable alternative source for mesenchymal stem cells. SADSCs were isolated from human scalp biopsy and then were characterized by Flow cytometry. SADSCS expressed CD90, CD44, and CD105 but did not express CD45 surface marker. Growth factors were used for chondrogenesis induction. Histology and immunohistology methods and gene expression by real-time PCR 14 days after induced cells have shown the feature of chondrocytes in their morphology and extracellular matrix in both inducing patterns of combination and cycling induction. Moreover, the expression of gene markers of chondrogenesis for example collagen type II aggrecan and SOX9 has shown by real-time PCR assay. Then, SADSCs were seeded alone on polycaprolatone (PCL) and with Freeze thaw Freeze (PCL+FTF) scaffolds and SADSCs differentiated toward the chondrogenic lineage and chondrogenesis induction were evaluated using scanning electron microcopy (SEM) and MTT assay. Our results showed that SADSCs were also similar to the other adipose-derived stem cells. Using TGF-beta3 and BMP-6 were effective for chondrogenesis induction. Therefore using of TGF-beta3 and BMP-6 growth factors may be the important key for in vitro chondrogenesis induction. The bio-composite of PCL+FTF nanofibrous scaffolds enhance the chondroblast differentiation and proliferation compared to PCL scaffolds .Therefore, our model will make it possible to study the mechanism of transition from chondroblast to chondrocyte.

The walking dead: sequential nuclear and organelle destruction during hair development.

BACKGROUND: Transition of hair shaft keratinocytes from actively respiring, nucleated cells to structural cells devoid of nucleus and cytoplasm is key to hair production. This form of cell 'death', or cornification, requires cellular organelle removal to allow the cytoplasm to become packed with keratin filament bundles that further require cross-linking to create a strong hair fibre. Although these processes are well described in epidermal keratinocytes, there is a lack of understanding of such mechanisms, specifically in the hair follicle. OBJECTIVES: To gain insights into cornification mechanisms within the hair follicle and thus improve our understanding of normal hair physiology. METHODS: Scalp biopsies and hair-pluck samples were obtained from healthy human donors and analysed microscopically after immunohistochemical staining. RESULTS: A focal point of respiratory activity was evident in keratogenous zone cells within the hair shaft, which also exhibited nuclear damage. Nuclear degradation occurred via both caspase-dependent and caspase-independent pathways. Conversely, mitophagy was driven by Bnip3L and restricted to the boundary of the keratogenous zone at Adamson's Fringe. CONCLUSIONS: We propose a model of stepwise living-dead transition within the first 1 mm of hair formation, whereby fully functional, nucleated cells first consolidate required functions by degrading nuclear DNA, yet continue to respire and provide the source of reactive oxygen species required for keratin cross-linking. Finally, as the cells become packed with keratin bundles, Bnip3L expression triggers mitophagy to rid the cells of the last remaining 'living' characteristic, thus completing the march from 'living' to 'dead' within the hair follicle.

Knockout of p16INK4a promotes aggregative growth of dermal papilla cells.

OBJECTIVE: Dermal papilla cells (DPCs) are located in the hair follicles and play an important role in hair growth. These cells have the ability to induce hair follicle formation when they display aggregative behavior. DPCs derived from the androgenetic alopecia (AGA) area undergo premature senescence in vitro, associated with p16INK4a expression. The aim of the current study was to investigate the expression of p16INK4a in aggregative and non-aggregative DPCs and the effect of p16INK4a down-regulation in these cells by adenovirus-mediated RNA interference (RNAi). METHOD: DPCs were isolated and cultured from healthy human scalp. p16INK4a gene and protein were detected in aggregative and non-aggregative cells. Expression of p16INK4a in DPCs was silenced by infection with rAd5-CDKN1A-1p2shRNA. Cell fate was monitored after infection. The growth of cells was measured by MTT assay. Cell cycle was evaluated by flow cytometry (FCM). RESULTS: DPCs were isolated by digestion and showed aggregative behavior for six passages. The expression of p16INK4a showed a clear upward trend in non-aggregative cells when compared with aggregative group. p16INK4a expression was silenced by rAd5-CDKN1A-1p2shRNA (p<0.05). The p16INK4a-silenced cells grew more rapidly and exhibited a trend towards aggregative growth. There was an increase in the proportion of cells in G1 phase, while those in S phase were reduced after p16INK4a gene silencing (p<0.05). CONCLUSION: Our results suggest that p16INK4a plays an important role in the premature senescence and aggregative behavior of DPCs. These observations can lead to novel therapeutic strategies for treatment of AGA.

Knockout of p16INK4a promotes aggregative growth of dermal papilla cells

Summary Objective: Dermal papilla cells (DPCs) are located in the hair follicles and play an important role in hair growth. These cells have the ability to induce hair follicle formation when they display aggregative behavior. DPCs derived from the androgenetic alopecia (AGA) area undergo premature senescence in vitro, associated with p16INK4a expression. The aim of the current study was to investigate the expression of p16INK4a in aggregative and non-aggregative DPCs and the effect of p16INK4a down-regulation in these cells by adenovirus-mediated RNA interference (RNAi). Method: DPCs were isolated and cultured from healthy human scalp. p16INK4a gene and protein were detected in aggregative and non-aggregative cells. Expression of p16INK4a in DPCs was silenced by infection with rAd5-CDKN1A-1p2shRNA. Cell fate was monitored after infection. The growth of cells was measured by MTT assay. Cell cycle was evaluated by flow cytometry (FCM). Results: DPCs were isolated by digestion and showed aggregative behavior for six passages. The expression of p16INK4a showed a clear upward trend in non-aggregative cells when compared with aggregative group. p16INK4a expression was silenced by rAd5-CDKN1A-1p2shRNA (p<0.05). The p16INK4a-silenced cells grew more rapidly and exhibited a trend towards aggregative growth. There was an increase in the proportion of cells in G1 phase, while those in S phase were reduced after p16INK4a gene silencing (p<0.05). Conclusion: Our results suggest that p16INK4a plays an important role in the premature senescence and aggregative behavior of DPCs. These observations can lead to novel therapeutic strategies for treatment of AGA.

Muse Cells Derived from Dermal Tissues Can Differentiate into Melanocytes.

The objective of the authors has been to obtain multilineage-differentiating stress-enduring cells (Muse cells) from primary cultures of dermal fibroblasts, identify their pluripotency, and detect their ability to differentiate into melanocytes. The distribution of SSEA-3-positive cells in human scalp skin was assessed by immunohistochemistry, and the distribution of Oct4, Sox2, Nanog, and SSEA-3-positive cells was determined by immunofluorescence staining. The expression levels of Sox2, Oct4, hKlf4, and Nanog mRNAs and proteins in Muse cells were determined by reverse transcription polymerase chain reaction (RT-PCR) analyses and Western blots, respectively. These Muse cells differentiated into melanocytes in differentiation medium. The SSEA-3-positive cells were scattered in the basement membrane zone and the dermis, with comparatively more in the sebaceous glands, vascular and sweat glands, as well as the outer root sheath of hair follicles, the dermal papillae, and the hair bulbs. Muse cells, which have the ability to self-renew, were obtained from scalp dermal fibroblasts by flow cytometry sorting with an anti-SSEA-3 antibody. The results of RT-PCR, Western blot, and immunofluorescence staining showed that the expression levels of Oct4, Nanog, Sox2, and Klf4 mRNAs and proteins in Muse cells were significantly different from their parental dermal fibroblasts. Muse cells differentiated into melanocytes when cultured in melanocyte differentiation medium, and the Muse cell-derived melanocytes expressed the melanocyte-specific marker HMB45. Muse cells could be obtained by flow cytometry from primary cultures of scalp dermal fibroblasts, which possessed the ability of pluripotency and self-renewal, and could differentiate into melanocytes in vitro.

Sox9 facilitates proliferation, differentiation and lipogenesis in primary cultured human sebocytes.

BACKGROUND: The transcription factor Sox9 is pivotal in the morphogenesis of hair follicles, but its role in sebocytes is poorly understood. OBJECTIVE: We investigated the effect of Sox9 on human sebocyte proliferation, differentiation and lipogenesis. METHODS: Sox9 expression was detected by immunohistochemistry in normal skin and acne lesion. Primary cultured human sebocytes were transfected with adenovirus expressing GFP-Sox9 or Sox9 microRNA. Sox9 and peroxisome proliferator-activated receptor (PPAR)γ expression in sebocytes was detected by quantitative real-time PCR, Western blot and immunocytofluorescence; cell proliferation was measured by MTS and [3H]-thymidine incorporation assays; cell cycle distribution and apoptosis were evaluated by propidium iodide staining-based flow cytometry; and intracellular lipid levels were assessed by Oil Red O stain. RESULTS: Sox9 immunostaining was increased in mature sebocytes of acne lesion compared with normal skin. Expression of Sox9 mRNA and protein and PPARγ protein was elevated with cell confluent levels in sebocytes. Sox9 overexpression enhanced proliferation, differentiation, proportion of S and G2/M cells, lipogenesis and PPARγ expression in sebocytes, while Sox9 silencing caused inhibition of differentiation, lipogenesis and PPARγ expression, and increase of G1 and sub-G1 (apoptotic) cell fraction. The suppression of Sox9 knockdown on sebocyte growth was observed using [3H]-thymidine incorporation but not MTS assay. CONCLUSION: These results demonstrate that Sox9 can reinforce sebocyte proliferation, differentiation and lipogenesis. The G1/S transition arrest and apoptotic induction might contribute to inhibitory effect of Sox9 silencing on sebocyte proliferation.

The arrector pili muscle, the bridge between the follicular stem cell niche and the interfollicular epidermis.

Proximally, the arrector pili muscle (APM) attaches to the follicular stem cell niche in the bulge, but its distal properties are comparatively unclear. In this work, a novel method employing an F-actin probe, phalloidin, was employed to visualize the APM anatomy. Phalloidin staining of the APM was validated by comparison with conventional antibodies/stains and by generating three-dimensional reconstructions. The proximal attachment of the APM to the bulge in 8 patients with androgenic alopecia was studied using Masson's trichrome stain. Phalloidin visualized extensive branching of the APM. The distal end of the human APM exhibits a unique "C"-shaped structure connecting to the dermal-epidermal junction. The proximal APM attachment was observed to be lost or extremely miniaturized in androgenic alopecia. The unique shape, location, and attachment sites of the APM suggest a significant role for this muscle in maintaining follicular integrity. Proximally, the APM encircles the follicular unit and only attaches to the primary hair follicle in the bulge; this attachment is lost in irreversible hair loss. The APM exhibits an arborized morphology as it ascends toward the epidermis, and anchors to the basement membrane.

Strong Components of Epigenetic Memory in Cultured Human Fibroblasts Related to Site of Origin and Donor Age.

Differentiating pluripotent cells from fibroblast progenitors is a potentially transformative tool in personalized medicine. We previously identified relatively greater success culturing dura-derived fibroblasts than scalp-derived fibroblasts from postmortem tissue. We hypothesized that these differences in culture success were related to epigenetic differences between the cultured fibroblasts by sampling location, and therefore generated genome-wide DNA methylation and transcriptome data on 11 intrinsically matched pairs of dural and scalp fibroblasts from donors across the lifespan (infant to 85 years). While these cultured fibroblasts were several generations removed from the primary tissue and morphologically indistinguishable, we found widespread epigenetic differences by sampling location at the single CpG (N = 101,989), region (N = 697), "block" (N = 243), and global spatial scales suggesting a strong epigenetic memory of original fibroblast location. Furthermore, many of these epigenetic differences manifested in the transcriptome, particularly at the region-level. We further identified 7,265 CpGs and 11 regions showing significant epigenetic memory related to the age of the donor, as well as an overall increased epigenetic variability, preferentially in scalp-derived fibroblasts-83% of loci were more variable in scalp, hypothesized to result from cumulative exposure to environmental stimuli in the primary tissue. By integrating publicly available DNA methylation datasets on individual cell populations in blood and brain, we identified significantly increased inter-individual variability in our scalp- and other skin-derived fibroblasts on a similar scale as epigenetic differences between different lineages of blood cells. Lastly, these epigenetic differences did not appear to be driven by somatic mutation--while we identified 64 probable de-novo variants across the 11 subjects, there was no association between mutation burden and age of the donor (p = 0.71). These results depict a strong component of epigenetic memory in cell culture from primary tissue, even after several generations of daughter cells, related to cell state and donor age.

Manejo quirúrgico de la aplasia cutis congénita / Surgical management of aplasia cutis congenita

Introducción: La aplasia cutis congénita (ACC) es una malformación congénita rara que afecta sobre todo al cuero cabelludo, aunque puede afectar al pericráneo, el cráneo y la meninges. Las complicaciones pueden llegar a ser fatales, por lo que es necesario un tratamiento oportuno. El tratamiento sigue siendo controvertido, sin encontrar un consenso entre el abordaje conservador y el quirúrgico. El objetivo de este estudio es describir nuestra experiencia en el manejo de la ACC. Material y métodos: Estudio descriptivo retrospectivo de las historias clínicas de los pacientes menores de 14 años con diagnóstico de ACC, atendidos entre el año 2000 y el 2013. Resultados: Veintidós casos de ACC con lesiones que variaban de 1cm (0,79 cm2) a 14cm (153,94 cm2). Dieciocho casos presentaron lesiones en el cuero cabelludo, 3 en extremidades y uno en tronco. Se realizó tratamiento conservador en 9 y quirúrgico en 13 (8 cierres primarios, 2 plastias, 2 injertos cutáneos y un colgajo). Dos pacientes fallecieron por complicaciones de otras patologías no asociadas a la ACC. Conclusiones: La ACC es infrecuente y puede tener un desenlace fatal. Para prevenirla es necesaria una evaluación inicial completa para establecer un tratamiento oportuno. La cirugía es una buena opción terapéutica, sobre todo en defectos con diámetro>4cm (12,6 cm2), ya que disminuye el riesgo de complicaciones mortales (AU)

Introduction: Aplasia cutis congenita (ACC) is a rare congenital malformation that commonly involves the scalp, but can affect pericranium, bone and dura mater. Complications are rare, but can be fatal, so early treatment must be achieved. The treatment remains controversial with no consensus between the conservative and surgical approach. The aim of this study is to describe our experience in the management of ACC. Material and methods: Retrospective review of the medical records of all children up to 14 years diagnosed with ACC and treated between 2000 and 2013. Results: There were a total of 22 cases of ACC with lesions ranging from 1cm (0.79 cm2) to 14cm (153.94 cm2). ACC of the scalp was found in 18 cases, with 3 in extremities and 1 in trunk. Conservative treatment was performed on 9 patients and 13 underwent surgical treatment (8 primary closures, 2 plasties, 2 skin grafts, and 1 skin flap). Two patients died due to complications of other diseases not related with the ACC. Conclusions: ACC is a rare disease that can be fatal. A complete initial assessment to establish early treatment is necessary to prevent this. Surgery should be considered as an initial therapeutic option in defects >4cm (>12.6 cm2) as it prevents the risk of fatal complications (AU)

A precise automatic system for the hair assessment in hair-care diagnosis applications.

BACKGROUND/PURPOSE: One emerging subject in medical image processing is to quantitatively assess the health and the properties of cranial hairs, including density, diameter, length, level of oiliness, and others. This information helps hair specialists with making a more accurate diagnosis and the therapy required. We develop a practical hair counting algorithm. This analytic system calculates the number of hairs on a scalp using a digital microscope camera, providing accurate information for both the hair specialist and the patient. Our proposed hair counting algorithm is substantially more accurate than the Hough-based one, and is robust to curls, oily scalp, noise-corruption, and overlapping hairs, under various levels of illumination. Rather than manually counting the hairs on a person's scalp, the proposed system determines the density, diameter, length, and level of oiliness of the hairs. METHODS: We propose an automated system for counting the amount of hairs in the microscopy images. To reduce the effect of bright spots, we develop a robust morphological algorithm for color to smooth out the color and preserve the fidelity of the hair. Then, we utilize a modified Hough transform algorithm to detect the different hair lengths and to reduce any false detection due to noise. Our proposed system enables us to look at curved hairs as multiple pieces of straight lines. To avoid missing hairs when the thinning process is applied, we use edge information to discover any hidden or overlapping hairs. Finally, we employ a mutually associative regression method to label a group of line segments into a meaningful 'hair'. RESULTS: We demonstrated a novel approach for accurately computing the number of hairs, and successfully solved the three main obstacles in automated hair counting, including (i) oily and moist hairs, (ii) wavy and curly hairs, and (iii) under-estimation of the number of hairs occurs when hairs cross and occlude each other. The framework of this paper can be seen as the first step toward intelligent computer-aided medical image processing for cosmetic treatment applications. CONCLUSIONS: The goal of this study was to develop an automated hair counting system for clinical application using the microscope image from the hairs. The proposed method reduces the frequent errors and variances encountered when hairs are manually counted by human assessors. This clinical intelligent system can diagnose the health condition of a person's hair and can be applied in therapy recommendations by doctors for their patients.

CD133 expression correlates with membrane beta-catenin and E-cadherin loss from human hair follicle placodes during morphogenesis.

Genetic studies suggest that the major events of human hair follicle development are similar to those in mice, but detailed analyses of this process are lacking. In mice, hair follicle placode "budding" is initiated by invagination of Wnt-induced epithelium into the underlying mesenchyme. Modification of adherens junctions (AJs) is clearly required for budding. Snail-mediated downregulation of AJ component E-cadherin is important for placode budding in mice. Beta-catenin, another AJ component, has been more difficult to study owing to its essential functions in Wnt signaling, a prerequisite for hair follicle placode induction. Here, we show that a subset of human invaginating hair placode cells expresses the stem cell marker CD133 during early morphogenesis. CD133 associates with membrane beta-catenin in early placodes, and its continued expression correlates with loss of beta-catenin and E-cadherin from the cell membrane at a time when E-cadherin transcriptional repressors Snail and Slug are not implicated. Stabilization of CD133 via anti-CD133 antibody treatment of human fetal scalp explants depresses beta-catenin and E-cadherin membrane localization. We discuss this unique correlation and suggest a hypothetical model whereby CD133 promotes morphogenesis in early hair follicle placodes through the localized removal of membrane beta-catenin proteins and subsequent AJ dissolution.

Hypothalamic-pituitary-thyroid axis hormones stimulate mitochondrial function and biogenesis in human hair follicles.

Thyroid hormones regulate mitochondrial function. As other hypothalamic-pituitary-thyroid (HPT) axis hormones, i.e., thyrotropin-releasing hormone (TRH) and thyrotropin (TSH), are expressed in human hair follicles (HFs) and regulate mitochondrial function in human epidermis, we investigated in organ-cultured human scalp HFs whether TRH (30 nM), TSH (10 mU ml(-1)), thyroxine (T4) (100 nM), and triiodothyronine (T3) (100 pM) alter intrafollicular mitochondrial energy metabolism. All HPT-axis members increased gene and protein expression of mitochondrial-encoded subunit 1 of cytochrome c oxidase (MTCO1), a subunit of respiratory chain complex IV, mitochondrial transcription factor A (TFAM), and Porin. All hormones also stimulated intrafollicular complex I/IV activity and mitochondrial biogenesis. The TSH effects on MTCO1, TFAM, and porin could be abolished by K1-70, a TSH-receptor antagonist, suggesting a TSH receptor-mediated action. Notably, as measured by calorimetry, T3 and TSH increased follicular heat production, whereas T3/T4 and TRH stimulated ATP production in cultured HF keratinocytes. HPT-axis hormones did not increase reactive oxygen species (ROS) production. Rather, T3 and T4 reduced ROS formation, and all tested HPT-axis hormones increased the transcription of ROS scavengers (catalase, superoxide dismutase 2) in HF keratinocytes. Thus, mitochondrial biology, energy metabolism, and redox state of human HFs are subject to profound (neuro-)endocrine regulation by HPT-axis hormones. The neuroendocrine control of mitochondrial biology in a complex human mini-organ revealed here may be therapeutically exploitable.

A meeting of two chronobiological systems: circadian proteins Period1 and BMAL1 modulate the human hair cycle clock.

The hair follicle (HF) is a continuously remodeled mini organ that cycles between growth (anagen), regression (catagen), and relative quiescence (telogen). As the anagen-to-catagen transformation of microdissected human scalp HFs can be observed in organ culture, it permits the study of the unknown controls of autonomous, rhythmic tissue remodeling of the HF, which intersects developmental, chronobiological, and growth-regulatory mechanisms. The hypothesis that the peripheral clock system is involved in hair cycle control, i.e., the anagen-to-catagen transformation, was tested. Here we show that in the absence of central clock influences, isolated, organ-cultured human HFs show circadian changes in the gene and protein expression of core clock genes (CLOCK, BMAL1, and Period1) and clock-controlled genes (c-Myc, NR1D1, and CDKN1A), with Period1 expression being hair cycle dependent. Knockdown of either BMAL1 or Period1 in human anagen HFs significantly prolonged anagen. This provides evidence that peripheral core clock genes modulate human HF cycling and are an integral component of the human hair cycle clock. Specifically, our study identifies BMAL1 and Period1 as potential therapeutic targets for modulating human hair growth.