Local and systemic mechanisms that control the hair follicle stem cell niche.

Hair follicles are essential appendages of the mammalian skin, as hair performs vital functions of protection, thermoregulation and sensation. Hair follicles harbour exceptional regenerative abilities as they contain multiple somatic stem cell populations such as hair follicle stem cells (HFSCs) and melanocyte stem cells. Surrounding the stem cells and their progeny, diverse groups of cells and extracellular matrix proteins are organized to form a microenvironment (called 'niche') that serves to promote and maintain the optimal functioning of these stem cell populations. Recent studies have shed light on the intricate nature of the HFSC niche and its crucial role in regulating hair follicle regeneration. In this Review, we describe how the niche serves as a signalling hub, communicating, deciphering and integrating both local signals within the skin and systemic inputs from the body and environment to modulate HFSC activity. We delve into the recent advancements in identifying the cellular and molecular nature of the niche, providing a holistic perspective on its essential functions in hair follicle morphogenesis, regeneration and ageing.

Organ-level quorum sensing directs regeneration in hair stem cell populations.

Coordinated organ behavior is crucial for an effective response to environmental stimuli. By studying regeneration of hair follicles in response to patterned hair plucking, we demonstrate that organ-level quorum sensing allows coordinated responses to skin injury. Plucking hair at different densities leads to a regeneration of up to five times more neighboring, unplucked resting hairs, indicating activation of a collective decision-making process. Through data modeling, the range of the quorum signal was estimated to be on the order of 1 mm, greater than expected for a diffusible molecular cue. Molecular and genetic analysis uncovered a two-step mechanism, where release of CCL2 from injured hairs leads to recruitment of TNF-α-secreting macrophages, which accumulate and signal to both plucked and unplucked follicles. By coupling immune response with regeneration, this mechanism allows skin to respond predictively to distress, disregarding mild injury, while meeting stronger injury with full-scale cooperative activation of stem cells.

The cellular and molecular basis of direction selectivity of Aδ-LTMRs.

The perception of touch, including the direction of stimulus movement across the skin, begins with activation of low-threshold mechanosensory neurons (LTMRs) that innervate the skin. Here, we show that murine Aδ-LTMRs are preferentially tuned to deflection of body hairs in the caudal-to-rostral direction. This tuning property is explained by the finding that Aδ-LTMR lanceolate endings around hair follicles are polarized; they are concentrated on the caudal (downward) side of each hair follicle. The neurotrophic factor BDNF is synthesized in epithelial cells on the caudal, but not rostral, side of hair follicles, in close proximity to Aδ-LTMR lanceolate endings, which express TrkB. Moreover, ablation of BDNF in hair follicle epithelial cells disrupts polarization of Aδ-LTMR lanceolate endings and results in randomization of Aδ-LTMR responses to hair deflection. Thus, BDNF-TrkB signaling directs polarization of Aδ-LTMR lanceolate endings, which underlies direction-selective responsiveness of Aδ-LTMRs to hair deflection.

Signalling by senescent melanocytes hyperactivates hair growth.

Niche signals maintain stem cells in a prolonged quiescence or transiently activate them for proper regeneration1. Altering balanced niche signalling can lead to regenerative disorders. Melanocytic skin nevi in human often display excessive hair growth, suggesting hair stem cell hyperactivity. Here, using genetic mouse models of nevi2,3, we show that dermal clusters of senescent melanocytes drive epithelial hair stem cells to exit quiescence and change their transcriptome and composition, potently enhancing hair renewal. Nevus melanocytes activate a distinct secretome, enriched for signalling factors. Osteopontin, the leading nevus signalling factor, is both necessary and sufficient to induce hair growth. Injection of osteopontin or its genetic overexpression is sufficient to induce robust hair growth in mice, whereas germline and conditional deletions of either osteopontin or CD44, its cognate receptor on epithelial hair cells, rescue enhanced hair growth induced by dermal nevus melanocytes. Osteopontin is overexpressed in human hairy nevi, and it stimulates new growth of human hair follicles. Although broad accumulation of senescent cells, such as upon ageing or genotoxic stress, is detrimental for the regenerative capacity of tissue4, we show that signalling by senescent cell clusters can potently enhance the activity of adjacent intact stem cells and stimulate tissue renewal. This finding identifies senescent cells and their secretome as an attractive therapeutic target in regenerative disorders.

Lin28 enhances tissue repair by reprogramming cellular metabolism.

Regeneration capacity declines with age, but why juvenile organisms show enhanced tissue repair remains unexplained. Lin28a, a highly conserved RNA-binding protein expressed during embryogenesis, plays roles in development, pluripotency, and metabolism. To determine whether Lin28a might influence tissue repair in adults, we engineered the reactivation of Lin28a expression in several models of tissue injury. Lin28a reactivation improved hair regrowth by promoting anagen in hair follicles and accelerated regrowth of cartilage, bone, and mesenchyme after ear and digit injuries. Lin28a inhibits let-7 microRNA biogenesis; however, let-7 repression was necessary but insufficient to enhance repair. Lin28a bound to and enhanced the translation of mRNAs for several metabolic enzymes, thereby increasing glycolysis and oxidative phosphorylation (OxPhos). Lin28a-mediated enhancement of tissue repair was negated by OxPhos inhibition, whereas a pharmacologically induced increase in OxPhos enhanced repair. Thus, Lin28a enhances tissue repair in some adult tissues by reprogramming cellular bioenergetics. PAPERCLIP:

Mesenchyme governs hair follicle induction.

Tissue interactions are essential for guiding organ development and regeneration. Hair follicle formation relies on inductive signalling between two tissues, the embryonic surface epithelium and the adjacent mesenchyme. Although previous research has highlighted the hair-inducing potential of the mesenchymal component of the hair follicle - the dermal papilla and its precursor, the dermal condensate - the source and nature of the primary inductive signal before dermal condensate formation have remained elusive. Here, we performed epithelial-mesenchymal tissue recombination experiments using hair-forming back skin and glabrous plantar skin from mouse embryos to unveil that the back skin mesenchyme is inductive even before dermal condensate formation. Moreover, the naïve, unpatterned mesenchyme was sufficient to trigger hair follicle formation even in the oral epithelium. Building on previous knowledge, we explored the hair-inductive ability of the Wnt agonist R-spondin 1 and a Bmp receptor inhibitor in embryonic skin explants. Although R-spondin 1 instigated precocious placode-specific transcriptional responses, it was insufficient for hair follicle induction, either alone or in combination with Bmp receptor inhibition. Our findings pave the way for identifying the hair follicle-inducing cue.

Dynamic duo: Cell-extracellular matrix interactions in hair follicle development and regeneration.

Ectodermal organs, such as hair follicles, originate from simple epithelial and mesenchymal sheets through a complex developmental process driven by interactions between these cell types. This process involves dermal condensation, placode formation, bud morphogenesis, and organogenesis, and all of these processes require intricate interactions among various tissues. Recent research has emphasized the crucial role of reciprocal and dynamic interactions between cells and the extracellular matrix (ECM), referred to as the "dynamic duo", in the development of ectodermal organs. These interactions provide spatially and temporally changing biophysical and biochemical cues within tissues. Using the hair follicle as an example, this review highlights two types of cell-ECM adhesion units-focal adhesion-type and hemidesmosome-type adhesion units-that facilitate communication between epithelial and mesenchymal cells. This review further explores how these adhesion units, along with other cell-ECM interactions, evolve during hair follicle development and regeneration, underscoring their importance in guiding both developmental and regenerative processes.

Cyclic growth of dermal papilla and regeneration of follicular mesenchymal components during feather cycling.

How dermis maintains tissue homeostasis in cyclic growth and wounding is a fundamental unsolved question. Here, we study how dermal components of feather follicles undergo physiological (molting) and plucking injury-induced regeneration in chickens. Proliferation analyses reveal quiescent, transient-amplifying (TA) and long-term label-retaining dermal cell (LRDC) states. During the growth phase, LRDCs are activated to make new dermal components with distinct cellular flows. Dermal TA cells, enriched in the proximal follicle, generate both peripheral pulp, which extends distally to expand the epithelial-mesenchymal interactive interface for barb patterning, and central pulp, which provides nutrition. Entering the resting phase, LRDCs, accompanying collar bulge epidermal label-retaining cells, descend to the apical dermal papilla. In the next cycle, these apical dermal papilla LRDCs are re-activated to become new pulp progenitor TA cells. In the growth phase, lower dermal sheath can generate dermal papilla and pulp. Transcriptome analyses identify marker genes and highlight molecular signaling associated with dermal specification. We compare the cyclic topological changes with those of the hair follicle, a convergently evolved follicle configuration. This work presents a model for analyzing homeostasis and tissue remodeling of mesenchymal progenitors.

Development of Physiologically Relevant Skin Organoids from Human Induced Pluripotent Stem Cells.

The development of skin organs for studying developmental pathways, modeling diseases, or regenerative medicine purposes is a major endeavor in the field. Human induced pluripotent stem cells (hiPSCs) are successfully used to derive skin cells, but the field is still far from meeting the goal of creating skin containing appendages, such as hair follicles and sweat glands. Here, the goal is to generate skin organoids (SKOs) from human skin fibroblast or placental CD34+ cell-derived hiPSCs. With all three hiPSC lines, complex SKOs with stratified skin layers and pigmented hair follicles are generated with different efficacies. In addition, the hiPSC-derived SKOs develop sebaceous glands, touch-receptive Merkel cells, and more importantly eccrine sweat glands. Together, physiologically relevant skin organoids are developed by direct induction of embryoid body formation, along with simultaneous inactivation of transforming growth factor beta signaling, activation of fibroblast growth factor signaling, and inhibition of bone morphogenetic protein signaling pathways. The skin organoids created in this study can be used as valuable platforms for further research into human skin development, disease modeling, or reconstructive surgeries.

Melanocytes in regenerative medicine applications and disease modeling.

Melanocytes are dendritic cells localized in skin, eyes, hair follicles, ears, heart and central nervous system. They are characterized by the presence of melanosomes enriched in melanin which are responsible for skin, eye and hair pigmentation. They also have different functions in photoprotection, immunity and sound perception. Melanocyte dysfunction can cause pigmentary disorders, hearing and vision impairments or increased cancer susceptibility. This review focuses on the role of melanocytes in homeostasis and disease, before discussing their potential in regenerative medicine applications, such as for disease modeling, drug testing or therapy development using stem cell technologies, tissue engineering and extracellular vesicles.

Blue light inhibits cell viability and proliferation in hair follicle stem cells and dermal papilla cells.

Hair loss is a prevalent issue worldwide, which, though not life-threatening, can result in psychological problems, low self-esteem, and social anxiety. Previous studies have shown that ultraviolet radiation can have negative effects on hair follicle cells, leading to hair loss, while the impact of blue light on hair and hair follicle has largely been overlooked. This study aimed to examine the effects of blue light on hair follicle stem cells (HFSCs) and primary dermal papilla cells (DPCs), which are essential components of hair follicles. Human HFSCs and primary DPCs were exposed to blue light (457 nm) at various intensities (1, 4, 8, and 16 mW/cm2) for 3 days. Subsequently, cell viability, cell proliferation, and intracellular reactive oxygen species (ROS) were assessed. The results showed that blue light (457 nm) significantly reduced the cell viability and proliferation of HFSCs and DPCs in vitro, with the inhibition being intensity-dependent. Additionally, blue light triggered the overproduction of ROS in the DPCs. While the exact mechanisms by which blue light affects hair follicle cells remain unclear, these findings suggest that blue light could impede the growth of these cells. This insight may offer a new approach to protecting hair by avoiding exposure to high-intensity blue light.

EGFR marks a subpopulation of dermal mesenchymal cells highly expressing IGF1 which enhances hair follicle regeneration.

The skin harbours transcriptionally and functionally heterogeneous mesenchymal cells that participate in various physiological activities by secreting biochemical cues. In this study, we aimed to identify a new subpopulation of dermal mesenchymal cells that enhance hair follicle regeneration through a paracrine mechanism. Integrated single-cell RNA sequencing (scRNA-seq) data analysis revealed epidermal growth factor receptor (EGFR) as a marker of distinct fibroblast subpopulation in the neonatal murine dermis. Immunofluorescence staining and fluorescence-activated cell sorting (FACS) were used to validate the existence of the cell population in Krt14-rtTA-H2BGFP mouse. The difference of gene expression between separated cell subpopulation was examined by real-time PCR. Potential effect of the designated factor on hair follicle regeneration was observed after the application on excisional wounds in Krt14-rtTA-H2BGFP mouse. Immunofluorescence staining demonstrated the existence of dermal EGFR+ cells in neonatal and adult mouse dermis. The EGFR+ mesenchymal population, sorted by FACS, displayed a higher expression level of Igf1 (insulin-like growth factor 1). Co-localisation of IGF1 with EGFR in the mouse dermis and upregulated numbers of hair follicles in healed wounds following the application of exogenous IGF1 illustrated the contribution of EGFR+ cells in promoting wound-induced hair follicle neogenesis. Our results indicate that EGFR identifies a subpopulation of dermal fibroblasts that contribute to IGF1 promotion of hair follicle neogenesis. It broadens the understanding of heterogeneity and the mesenchymal cell function in skin and may facilitate the potential translational application of these cells.

The impact of perceived stress on the hair follicle: Towards solving a psychoneuroendocrine and neuroimmunological puzzle.

While popular belief harbors little doubt that perceived stress can cause hair loss and premature graying, the scientific evidence for this is arguably much thinner. Here, we investigate whether these phenomena are real, and show that the cyclic growth and pigmentation of the hair follicle (HF) provides a tractable model system for dissecting how perceived stress modulates aspects of human physiology. Local production of stress-associated neurohormones and neurotrophins coalesces with neurotransmitters and neuropeptides released from HF-associated sensory and autonomic nerve endings, forming a complex local stress-response system that regulates perifollicular neurogenic inflammation, interacts with the HF microbiome and controls mitochondrial function. This local system integrates into the central stress response systems, allowing the study of systemic stress responses affecting organ function by quantifying stress mediator content of hair. Focusing on selected mediators in this "brain-HF axis" under stress conditions, we distill general principles of HF dysfunction induced by perceived stress.

An exclusive review of melatonin effects on mammalian melanocytes and melanoma.

Melatonin influences mammalian coat colour and hair follicle pigmentation and also weakly alters the electrical stimulation of retinal cells in the eyes. A direct melanocytic response to melatonin is still uncertain in mammals and human skin pigmentation. Melatonin acts as a free radical scavenger and thus inhibits the initiation of cancer cell growth. Treatment of melanoma sees perspective features in the administration of melatonin along with known chemotherapeutic molecules to improve the efficacy of conventional cytotoxic agents. Being richly supplied with a variety of receptors, melanocytes and melanoma cells can be used as in vitro test models for pharmacological applications of known and novel drugs.

NFATc1 balances quiescence and proliferation of skin stem cells.

Quiescent adult stem cells reside in specialized niches where they become activated to proliferate and differentiate during tissue homeostasis and injury. How stem cell quiescence is governed is poorly understood. We report here that NFATc1 is preferentially expressed by hair follicle stem cells in their niche, where its expression is activated by BMP signaling upstream and it acts downstream to transcriptionally repress CDK4 and maintain stem cell quiescence. As stem cells become activated during hair growth, NFATc1 is downregulated, relieving CDK4 repression and activating proliferation. When calcineurin/NFATc1 signaling is suppressed, pharmacologically or via complete or conditional NFATc1 gene ablation, stem cells are activated prematurely, resulting in precocious follicular growth. Our findings may explain why patients receiving cyclosporine A for immunosuppressive therapy display excessive hair growth, and unveil a functional role for calcium-NFATc1-CDK4 circuitry in governing stem cell quiescence.

The aging skin microenvironment dictates stem cell behavior.

Aging manifests with architectural alteration and functional decline of multiple organs throughout an organism. In mammals, aged skin is accompanied by a marked reduction in hair cycling and appearance of bald patches, leading researchers to propose that hair follicle stem cells (HFSCs) are either lost, differentiate, or change to an epidermal fate during aging. Here, we employed single-cell RNA-sequencing to interrogate aging-related changes in the HFSCs. Surprisingly, although numbers declined, aging HFSCs were present, maintained their identity, and showed no overt signs of shifting to an epidermal fate. However, they did exhibit prevalent transcriptional changes particularly in extracellular matrix genes, and this was accompanied by profound structural perturbations in the aging SC niche. Moreover, marked age-related changes occurred in many nonepithelial cell types, including resident immune cells, sensory neurons, and arrector pili muscles. Each of these SC niche components has been shown to influence HF regeneration. When we performed skin injuries that are known to mobilize young HFSCs to exit their niche and regenerate HFs, we discovered that aged skin is defective at doing so. Interestingly, however, in transplantation assays in vivo, aged HFSCs regenerated HFs when supported with young dermis, while young HFSCs failed to regenerate HFs when combined with aged dermis. Together, our findings highlight the importance of SC:niche interactions and favor a model where youthfulness of the niche microenvironment plays a dominant role in dictating the properties of its SCs and tissue health and fitness.

Dysregulated behaviour of hair follicle stem cells triggers alopecia and provides potential therapeutic targets.

Due to a steady increase in the number of individuals suffering from alopecia, this condition has recently received increasing attention. Alopecia can be caused by various pathological, environmental or psychological factors, eventually resulting in abnormalities in hair follicle (HF) structures or HF regeneration disorders, especially dysregulated hair follicle stem cell (HFSC) behaviour. HFSC behaviour includes activation, proliferation and differentiation. Appropriate HFSC behaviour sustains a persistent hair cycle (HC). HFSC behaviour is mainly influenced by HFSC metabolism, ageing and the microenvironment. In this review, we summarize recent findings on how HFSC metabolism, ageing and the microenvironment give rise to hair growth disorders, as well as related genes and signalling pathways. Recent research on the application of stem cell-based hair tissue engineering and regenerative medicine to treat alopecia is also summarized. Determining how dysregulated HFSC behaviour underlies alopecia would be helpful in identifying potential therapeutic targets.

Constraints on the deformation of the vibrissa within the follicle.

Nearly all mammals have a vibrissal system specialized for tactile sensation, composed of whiskers growing from sensor-rich follicles in the skin. When a whisker deflects against an object, it deforms within the follicle and exerts forces on the mechanoreceptors inside. In addition, during active whisking behavior, muscle contractions around the follicle and increases in blood pressure in the ring sinus will affect the whisker deformation profile. To date, however, it is not yet possible to experimentally measure how the whisker deforms in an intact follicle or its effects on different groups of mechanoreceptors. The present study develops a novel model to predict vibrissal deformation within the follicle sinus complex. The model is based on experimental results from a previous ex vivo study on whisker deformation within the follicle, and on a new histological analysis of follicle tissue. It is then used to simulate whisker deformation within the follicle during passive touch and active whisking. Results suggest that the most likely whisker deformation profile is "S-shaped," crossing the midline of the follicle right below the ring sinus. Simulations of active whisking indicate that an increase in overall muscle stiffness, an increase in the ratio between deep and superficial intrinsic muscle stiffness, and an increase in sinus blood pressure will all enhance tactile sensitivity. Finally, we discuss how the deformation profiles might map to the responses of primary afferents of each mechanoreceptor type. The mechanical model presented in this study is an important first step in simulating mechanical interactions within whisker follicles.

CD34 defines melanocyte stem cell subpopulations with distinct regenerative properties.

Melanocyte stem cells (McSCs) are the undifferentiated melanocytic cells of the mammalian hair follicle (HF) responsible for recurrent generation of a large number of differentiated melanocytes during each HF cycle. HF McSCs reside in both the CD34+ bulge/lower permanent portion (LPP) and the CD34- secondary hair germ (SHG) regions of the HF during telogen. Using Dct-H2BGFP mice, we separate bulge/LPP and SHG McSCs using FACS with GFP and anti-CD34 to show that these two subsets of McSCs are functionally distinct. Genome-wide expression profiling results support the distinct nature of these populations, with CD34- McSCs exhibiting higher expression of melanocyte differentiation genes and with CD34+ McSCs demonstrating a profile more consistent with a neural crest stem cell. In culture and in vivo, CD34- McSCs regenerate pigmentation more efficiently whereas CD34+ McSCs selectively exhibit the ability to myelinate neurons. CD34+ McSCs, and their counterparts in human skin, may be useful for myelinating neurons in vivo, leading to new therapeutic opportunities for demyelinating diseases and traumatic nerve injury.

Phenotypical Conversions of Dermal Adipocytes as Pathophysiological Steps in Inflammatory Cutaneous Disorders.

Adipocytes from the superficial layer of subcutaneous adipose tissue undergo cyclic de- and re-differentiation, which can significantly influence the development of skin inflammation under different cutaneous conditions. This inflammation can be connected with local loading of the reticular dermis with lipids released due to de-differentiation of adipocytes during the catagen phase of the hair follicle cycle. Alternatively, the inflammation parallels a widespread release of cathelicidin, which typically takes place in the anagen phase (especially in the presence of pathogens). Additionally, trans-differentiation of dermal adipocytes into myofibroblasts, which can occur under some pathological conditions, can be responsible for the development of collateral scarring in acne. Here, we provide an overview of such cellular conversions in the skin and discuss their possible involvement in the pathophysiology of inflammatory skin conditions, such as acne and psoriasis.