Advances in Understanding Hair Growth

In this short review, I introduce an integrated vision of human hair follicle behavior and describe opposing influences that control hair follicle homeostasis, from morphogenesis to hair cycling. The interdependence and complementary roles of these influences allow us to propose that the hair follicle is a true paradigm of a “Yin Yang” type, that is a cold/slow-hot/fast duality. Moreover, a new promising field is emerging, suggesting that glycans are key elements of hair follicle growth control.


Introduction
The hair follicle is a true paradigm of mesenchymal-epithelial interaction. From early morphogenesis to a fully formed organ, the hair follicle life-cycle is controlled by a dialog between mesenchymal and epithelial compartments 1 . However, this dialog relies on a delicate balance between conflicting and/or opposing influences.
With respect to hair follicle morphogenesis, the reaction-diffusion model explains how slowly diffusing inducers and rapidly diffusing inhibitors orchestrate, through local activation and at distance inhibition, the hair follicle patterned formation. Indeed, the seminal work of A. Turing 2 has been recently confirmed through a formal identification of morphogen activator-inhibitor couples, such as Wnt/DKK1 3 ( Figure 1) and EDAR/BMP 4 .
Considering its dual mesenchymal and epithelial origin, the hair follicle can be considered a composite organ, with a concentric structure. Dermal and epithelial compartments interact with each other and are characterized by specific differentiation programs. Opposing signaling pathways concur to control the unique behavior of human hair follicle and maintain its unique intrinsic homeostasis. As the activity of diffusible factors, such as growth factors and morphogens, can be modulated by glycans, their possible role in hair growth control must be taken into account.

Hair follicle behavior
The hair follicle is the only organ in mammals that sequentially and repeatedly transits from a phase of active fiber production (anagen) to a resting phase (telogen), through rapid phases of tissue regression (catagen) and regeneration (neogen). A recently published comprehensive guide describes most of the morphological and immunohistological markers that characterize the different stages of the human hair follicle cycle and the intense tissue remodeling events which take place 5 . Of note, hair follicle regeneration relies on the cyclical activation of stem cells 6 . In the human hair follicle, these stem cells are harbored within two distinct reservoirs 7,8 , one of them bathing in a hypoxic environment 9 . Instead of a cyclical behavior with an intrinsic automaton, the human hair follicle exhibits a stochastic behavior, the probability of duration of each phase fitting with a lognormal equation 10   postulates the existence of a bi-stable equilibrium 11 which controls human hair follicle dynamics, including an active steady state (the anagen stage) and a resting steady state (the telogen stage), the transition between these two steady states involving either a degradation phase (the catagen phase) or a neo-morphogenesis phase (the neogen phase). It is now believed that mesenchymal and epithelial oscillators control the stochastic autonomous switching between these two steady states 12,13 . The transition phases are both controlled by a complex and dynamic network of interacting activators and inhibitors, diffusible morphogens, and growth factors of opposite influences 14 . Of note, however, extrapolating from results only obtained in rodents must be approached with caution, since major differences exist between human and mouse hair follicles in terms of phase duration, synchronicity, tissue remodeling, stem cell reservoirs, and so on.
During the active steady state, hair fiber production results from a finely, timely, and spatially tuned choreography of gene expression, which is highly sensitive to stimulatory and inhibitory signals. A number of signaling pathways 15 , cytokines 16,17 , neuropeptides 18 , hormones 19-22 , prostaglandins 23 , and growth factors 24 are known to modulate the duration of the active steady state of the hair follicle ( Figure 3). For example, while insulin-like growth factor (IGF)-1 is required for anagen maintenance 25,26 , fibroblast growth factor (FGF)-5 appears to be a crucial regulator of hair length in humans 27 , as a strong inducer of the catagen phase. Moreover, the human hair follicle is endowed with an autonomous androgen metabolism 28 , a strict dependence on arginine 29 , polyamines 30 , and glucose 31 for growth, and a specific immunological response 32 . The hair follicle is also endowed with a full prostaglandin metabolism and a complex network of prostaglandin (PG) receptors 33,34 . Recent data suggest that a delicate equilibrium between PGE2/PGF2a on the one hand and PGD2 on the other hand controls the duration of the active steady state. PGE2/PGF2a promotes hair growth maintenance, while PGD2 inhibits it and triggers anagen to catagen transition 35 . Finally, re-evaluating the mechanisms by which agents such as cyclosporine A 36 or JAK-STAT inhibitors 37 promote human hair growth might help to identify new key genes and pathways involved in the control of hair growth.
Besides the active steady state, new data demonstrate that the resting steady state is not as quiescent as suspected and can be divided into a refractory period and a permissive period. Indeed, during the telogen phase, the follicle is under the influence of factors that would repress the onset of the neogen phase and factors that would trigger it. Specifically, a strong expression of bone morphogenetic protein (BMP) and FGF-18 defines the refractory period, during which the neogen onset is prevented. The progressive increase in the production of BMP antagonist noggin, Wnt/Fzz/b-catenin pathway activators, and transforming growth factor (TGF)-β2 then reaches a critical threshold that shifts the telogen follicle to a competency status, receptive to FGF-7, secreted by the nearby dermal papilla, and, ultimately, triggers the onset of the neogen phase 38 .

Glyco-biology of the human hair follicle
It is clear from the above that the complex and rhythmic behavior of the human hair follicle is under the control of multiple, intricate pathways with opposing influences. In this respect, the interdependence and complementary roles of these influences allow us to propose that the hair follicle is a true paradigm of a "Yin Yang" type duality and harmony. However, in our opinion, the fine tuning of these influences cannot solely rely on the timely and spatially controlled gene expression, but also on glycans, "the third revolution in evolution" 39 . Glycans are endowed with such a huge molecular diversity that they can be considered the third language of life, after DNA and proteins.
Linear or branched oligosaccharides can be attached to a protein backbone via O-(serine/threonine) or N-(asparagine) linkages. They form the large class of N-Complex type glycans. Glycosaminoglycans are linear copolymers of 6-O-sulfated disaccharide units which define them as chondroitin, dermatan, keratin, or heparin sulfates. Proteoglycans have one or more glycosaminoglycan side chains attached to a core protein. Glycosaminoglycans, proteoglycans, and glycan moieties of glycoproteins have long been known to play important roles in the maintenance of protein conformation and solubility, protection against proteolytic degradation, mediation of biological activity, intracellular sorting and externalization, and embryonic development and differentiation [40][41][42][43][44][45] . The distribution of proteoglycans in the human hair follicle was originally described in the early 1990s, namely for chondroitin sulfate, dermatan sulfate, and heparin sulfate proteoglycans 46 , for syndecan 1, perlecan and decorin 47 , and for versican 48 . Thanks to the availability of new immunological tools, the distribution of proteoglycans in the human hair follicle has been further refined 49 (Figure 4), highlighting a complex, dynamic, and regionalized network of proteoglycans. With respect to cell surface complex type N-glycans, the use of specific fluorescently labeled lectins (saccharide-binding proteins) revealed a differential N-glycan composition among the different hair follicle compartments 50-52 ( Figure 5).
What could be the role of these glycans? It has been known for quite a long time that growth factor activation could be regulated by proteoglycans 53,54 and that heparan sulfate proteoglycans were involved in fine-tuning mammalian physiology 55 and in cell signaling during development 56 . With respect to key regulators of hair follicle growth and cycling, syndecans modulate Wnt signaling cascades 57 , the glycosaminoglycan chains of proteoglycans shape Hedgehog gradients and signal transduction 58 , and O-linked glycosylation controls Notch1 interaction with its cognate Delta-like 4 receptor 59 . Decorin, a small leucine-rich proteoglycan, directly modulates TGF-β, epidermal growth factor (EGF), IGF-1 and hepatocyte growth factor (HGF) signaling, all known actors of hair follicle cycling 60 , and appears to act as an anagen inducer 61 . Altogether, these recent results designate glycans as long time ignored key players in hair growth control. But, on top of that, enzymes can further modulate the biological activity of these glycans. For example, fucosyl transferase is absolutely required for Notch activity, and disruption of fucosyl transferase expression in murine hair follicle lineages results in aberrant telogen morphology, a decrease of bulge stem cell markers, a delay in anagen re-entry, and dysregulation of proliferation and apoptosis during the hair cycle transition 62 . With respect to proteoglycans, heparanase (an endoglycosidase that cleaves heparin sulfate) was found expressed in the outer root sheath of murine hair follicles and identified as an important regulator of hair growth through its ability to release heparin-bound growth factors 63 . In the human hair follicle, however, heparanase was found located in the inner root sheath. Its inhibition provoked an immediate transition from anagen to catagen 64 . In this case, the HPSG/heparanase network appears to be a key controller of internal hair follicle homeostasis.
Finally, extracellular sulfatases appear to be critical regulators of heparin sulfate activities. Sulf1 and Sulf2, by removing glucosamine-6S groups from specific regions of heparan sulfate chain, modulate (a) Wnt interaction with its cognate receptor Frizzled, (b) BMP signaling by releasing BMP antagonist Noggin, and (c) FGF-2 ability to form the functional FGF-2-HS-FGFR ternary complex 65,66 . Of note, TGF-β1, by inducing Sulf1 expression 67 , might indirectly modulate Wnt, BMP, and FGF-2 activities, which could explain its inhibitory effect on hair growth. From a clinical point of view, alterations of glycosaminoglycan degradation provoke mucopolysaccharidoses and abnormalities in hair morphology 68 , which can be reversed by appropriate enzyme replacement therapy 69 .

Conclusion
The hair follicle is clearly endowed with a unique behavior. Its bi-stability and the intense remodeling processes that it provokes rely on the permanent dialog between opposing and complementary influences, impacting all follicle compartments. From this interdependent duality, one can easily understand that an optimal way to describe the complex equilibrium which controls hair follicle homeostasis is the concept of "Yin Yang". Until recently, the understanding of hair growth mainly relied on deciphering the patterns of gene expression within the different hair follicle compartments throughout the hair cycle 70,71 . From now on, the fine-tuning of the activities of growth factors and morphogens by the modulating effects of glycans will also have to be taken into consideration.
From a prospective point of view, it is likely that a better understanding of hair diseases, and more specifically the role of inflammation and immune response in the development of alopecia areata 72 and androgenetic alopecia 73 , will likely provide further insights into the role of the so-called immune privilege 74 in hair growth control. Moreover, with the advent of mature metabolomics technologies 75 coupled with in vitro human hair growth technology 76 , one can predict that this integrative approach will permit us to identify these key metabolic pathways sustaining normal hair growth.

Competing interests
The author is an employee of L'Oréal company.

Grant information
The author(s) declared that no grants were involved in supporting this work. Editorial Note on the Review Process are commissioned from members of the prestigious and are edited as a F1000 Faculty Reviews F1000 Faculty service to readers. In order to make these reviews as comprehensive and accessible as possible, the referees provide input before publication and only the final, revised version is published. The referees who approved the final version are listed with their names and affiliations but without their reports on earlier versions (any comments will already have been addressed in the published version).