Modeling Edar expression reveals the hidden dynamics of tooth signaling center patterning.

When patterns are set during embryogenesis, it is expected that they are straightly established rather than subsequently modified. The patterning of the three mouse molars is, however, far from straight, likely as a result of mouse evolutionary history. The first-formed tooth signaling centers, called MS and R2, disappear before driving tooth formation and are thought to be vestiges of the premolars found in mouse ancestors. Moreover, the mature signaling center of the first molar (M1) is formed from the fusion of two signaling centers (R2 and early M1). Here, we report that broad activation of Edar expression precedes its spatial restriction to tooth signaling centers. This reveals a hidden two-step patterning process for tooth signaling centers, which was modeled with a single activator-inhibitor pair subject to reaction-diffusion (RD). The study of Edar expression also unveiled successive phases of signaling center formation, erasing, recovering, and fusion. Our model, in which R2 signaling center is not intrinsically defective but erased by the broad activation preceding M1 signaling center formation, predicted the surprising rescue of R2 in Edar mutant mice, where activation is reduced. The importance of this R2-M1 interaction was confirmed by ex vivo cultures showing that R2 is capable of forming a tooth. Finally, by introducing chemotaxis as a secondary process to RD, we recapitulated in silico different conditions in which R2 and M1 centers fuse or not. In conclusion, pattern formation in the mouse molar field relies on basic mechanisms whose dynamics produce embryonic patterns that are plastic objects rather than fixed end points.

Dominant-negative Sox18 function inhibits dermal papilla maturation and differentiation in all murine hair types.

SOX family proteins SOX2 and SOX18 have been reported as being essential in determining hair follicle type; however, the role they play during development remains unclear. Here, we demonstrate that Sox18 regulates the normal differentiation of the dermal papilla of all hair types. In guard (primary) hair dermal condensate (DC) cells, we identified transient Sox18 in addition to SOX2 expression at E14.5, which allowed fate tracing of primary DC cells until birth. Similarly, expression of Sox18 was detected in the DC cells of secondary hairs at E16.5 and in tertiary hair at E18.5. Dominant-negative Sox18 mutation (opposum) did not prevent DC formation in any hair type. However, it affected dermal papilla differentiation, restricting hair formation especially in secondary and tertiary hairs. This Sox18 mutation also prevented neonatal dermal cells or dermal papilla spheres from inducing hair in regeneration assays. Microarray expression studies identified WNT5A and TNC as potential downstream effectors of SOX18 that are important for epidermal WNT signalling. In conclusion, SOX18 acts as a mesenchymal molecular switch necessary for the formation and function of the dermal papilla in all hair types.

Getting to the root of scales, feather and hair: As deep as odontodes?

While every jawed vertebrate, or its recent ancestor, possesses teeth, skin appendages are characteristic of the living clades: skin denticles (odontodes) in chondrichthyans, dermal scales in teleosts, ducted multicellular glands in amphibians, epidermal scales in squamates, feathers in birds and hair-gland complexes in mammals, all of them showing a dense periodic patterning. While the odontode origin of teleost scales is generally accepted, the origin of both feather and hair is still debated. They appear long before mammals and birds, at least in the Jurassic in mammaliaforms and in ornithodires (pterosaurs and dinosaurs), and are contemporary to scales of early squamates. Epidermal scales might have appeared several times in evolution, and basal amniotes could not have developed a scaled dry integument, as the function of hair follicle requires its association with glands. In areas such as amnion, cornea or plantar pads, the formation of feather and hair is prevented early in embryogenesis, but can be easily reverted by playing with the Wnt/BMP/Shh pathways, which both imply the plasticity and the default competence of ectoderm. Conserved ectodermal/mesenchymal signalling pathways lead to placode formation, while later the crosstalk differs, as well as the final performing tissue(s): both epidermis and dermis for teeth and odontodes, mostly dermis for teleosts scales and only epidermis for squamate scale, feather and hair. We therefore suggest that tooth, dermal scale, epidermal scale, feather and hair evolved in parallel from a shared placode/dermal cell unit, which was present in a common ancestor, an early vertebrate gnathostome with odontodes, ca. 420 million years ago.

Ectopic Atoh1 expression drives Merkel cell production in embryonic, postnatal and adult mouse epidermis.

Merkel cells are mechanosensitive skin cells whose production requires the basic helix-loop-helix transcription factor Atoh1. We induced ectopic Atoh1 expression in the skin of transgenic mice to determine whether Atoh1 was sufficient to create additional Merkel cells. In embryos, ectopic Atoh1 expression drove ectopic expression of the Merkel cell marker keratin 8 (K8) throughout the epidermis. Epidermal Atoh1 induction in adolescent mice similarly drove widespread K8 expression in glabrous skin of the paws, but in the whisker pads and body skin ectopic K8+ cells were confined to hair follicles and absent from interfollicular regions. Ectopic K8+ cells acquired several characteristics of mature Merkel cells in a time frame similar to that seen during postnatal development of normal Merkel cells. Although ectopic K8+ cell numbers decreased over time, small numbers of these cells remained in deep regions of body skin hair follicles at 3 months post-induction. In adult mice, greater numbers of ectopic K8+ cells were created by Atoh1 induction during anagen versus telogen and following disruption of Notch signaling by conditional deletion of Rbpj in the epidermis. Our data demonstrate that Atoh1 expression is sufficient to produce new Merkel cells in the epidermis, that epidermal cell competency to respond to Atoh1 varies by skin location, developmental age and hair cycle stage, and that the Notch pathway plays a key role in limiting epidermal cell competency to respond to Atoh1 expression.

Keratin 79 identifies a novel population of migratory epithelial cells that initiates hair canal morphogenesis and regeneration.

The formation of epithelial tubes underlies the development of diverse organs. In the skin, hair follicles resemble tube-like structures with lumens that are generated through poorly understood cellular rearrangements. Here, we show that creation of the hair follicle lumen is mediated by early outward movement of keratinocytes from within the cores of developing hair buds. These migratory keratinocytes express keratin 79 (K79) and stream out of the hair germ and into the epidermis prior to lumen formation in the embryo. Remarkably, this process is recapitulated during hair regeneration in the adult mouse, when K79(+) cells migrate out of the reactivated secondary hair germ prior to formation of a new hair canal. During homeostasis, K79(+) cells line the hair follicle infundibulum, a domain we show to be multilayered, biochemically distinct and maintained by Lrig1(+) stem cell-derived progeny. Upward movement of these cells sustains the infundibulum, while perturbation of this domain during acne progression is often accompanied by loss of K79. Our findings uncover previously unappreciated long-distance cell movements throughout the life cycle of the hair follicle, and suggest a novel mechanism by which the follicle generates its hollow core through outward cell migration.

Two frizzled planar cell polarity signals in the Drosophila wing are differentially organized by the Fat/Dachsous pathway.

The regular array of distally pointing hairs on the mature Drosophila wing is evidence for the fine control of Planar Cell Polarity (PCP) during wing development. Normal wing PCP requires both the Frizzled (Fz) PCP pathway and the Fat/Dachsous (Ft/Ds) pathway, although the functional relationship between these pathways remains under debate. There is strong evidence that the Fz PCP pathway signals twice during wing development, and we have previously presented a Bidirectional-Biphasic Fz PCP signaling model which proposes that the Early and Late Fz PCP signals are in different directions and employ different isoforms of the Prickle protein. The goal of this study was to investigate the role of the Ft/Ds pathway in the context of our Fz PCP signaling model. Our results allow us to draw the following conclusions: (1) The Early Fz PCP signals are in opposing directions in the anterior and posterior wing and converge precisely at the site of the L3 wing vein. (2) Increased or decreased expression of Ft/Ds pathway genes can alter the direction of the Early Fz PCP signal without affecting the Late Fz PCP signal. (3) Lowfat, a Ft/Ds pathway regulator, is required for the normal orientation of the Early Fz PCP signal but not the Late Fz PCP signal. (4) At the time of the Early Fz PCP signal there are symmetric gradients of dachsous (ds) expression centered on the L3 wing vein, suggesting Ds activity gradients may orient the Fz signal. (5) Localized knockdown or over-expression of Ft/Ds pathway genes shows that boundaries/gradients of Ft/Ds pathway gene expression can redirect the Early Fz PCP signal specifically. (6) Altering the timing of ds knockdown during wing development can separate the role of the Ft/Ds pathway in wing morphogenesis from its role in Early Fz PCP signaling.

Expression of Foxi3 is regulated by ectodysplasin in skin appendage placodes.

BACKGROUND: Foxi3 is a member of the large forkhead box family of transcriptional regulators, which have a wide range of biological activities including manifold developmental processes. Heterozygous mutation in Foxi3 was identified in several hairless dog breeds characterized by sparse fur coat and missing teeth. A related phenotype called hypohidrotic ectodermal dysplasia (HED) is caused by mutations in the ectodysplasin (Eda) pathway genes. RESULTS: Expression of Foxi3 was strictly confined to the epithelium in developing ectodermal appendages in mouse embryos, but no expression was detected in the epidermis. Foxi3 was expressed in teeth and hair follicles throughout embryogenesis, but in mammary glands only during the earliest stages of development. Foxi3 expression was decreased and increased in Eda loss- and gain-of-function embryos, respectively, and was highly induced by Eda protein in embryonic skin explants. Also activin A treatment up-regulated Foxi3 mRNA levels in vitro. CONCLUSIONS: Eda and activin A were identified as upstream regulators of Foxi3. Foxi3 is a likely transcriptional target of Eda in ectodermal appendage placodes suggesting that HED phenotype may in part be produced by compromised Foxi3 activity. In addition to hair and teeth, Foxi3 may have a role in nail, eye, and mammary, sweat, and salivary gland development.

Spatial and temporal control of laminin-332 and -511 expressions during hair morphogenesis.

Hair is one of the smallest organs, but has many important functions to mammals. Hair morphogenesis occurs through the reciprocal exchange of epithelial and mesenchymal signals. There are some reports about the expression of laminin-511 and -332 during hair morphogenesis, but are no reports of the chronological expression and function of laminin-511 and its counter regulator laminin-332 during hair morphogenesis. Our results of immunoblotting revealed that laminin-332 proteins were detected at stage 0 and downregulated during stage 1 to stage 2, and then recovered at stage 3. However, laminin α5 expression was constant throughout stages 0-3. According to the results of semi-quantitative RT-PCR, the mRNA expression of all laminin-332 subunits increased gradually from stage 0 to stage 2, while the mRNA expression of all laminin-511 subunits remained constant from stage 0 to stage 3. Our results suggest that the proper expression of laminin-332 and laminin-511 may regulate appropriate hair morphogenesis.

Hair loss in infancy.

Hair diseases represent a significant portion of cases seen by pediatric dermatologists although hair has always been a secondary aspect in pediatricians and dermatologists training, on the erroneous basis that there is not much information extractable from it. Dermatologists are in the enviable situation of being able to study many disorders with simple diagnostic techniques. The hair is easily accessible to examination but, paradoxically, this approach is often disregarded by non-dermatologist. This paper has been written on the purpose of trying to serve in the diagnostic process of daily practice, and trying to help, for example, to distinguish between certain acquired and some genetically determined hair diseases. We will focus on all the data that can be obtained from our patients' hair and try to help on using the messages given by hair for each patient. Quite often it is extremely hard to distinguish between abnormality and normality in neonatal hair aspects. We will specially focus in the most common physiological changes that may mislead to an incorrect diagnosis. Specific treatment for those hair diseases that do have one, and basic general approach to improve the cosmetic appearance of hair, will be also be discussed for those hair disturbances that do not have a specific treatment.

Cyclic expression of lhx2 regulates hair formation.

Hair is important for thermoregulation, physical protection, sensory activity, seasonal camouflage, and social interactions. Hair is generated in hair follicles (HFs) and, following morphogenesis, HFs undergo cyclic phases of active growth (anagen), regression (catagen), and inactivity (telogen) throughout life. The transcriptional regulation of this process is not well understood. We show that the transcription factor Lhx2 is expressed in cells of the outer root sheath and a subpopulation of matrix cells during both morphogenesis and anagen. As the HFs enter telogen, expression becomes undetectable and reappears prior to initiation of anagen in the secondary hair germ. In contrast to previously published results, we find that Lhx2 is primarily expressed by precursor cells outside of the bulge region where the HF stem cells are located. This developmental, stage- and cell-specific expression suggests that Lhx2 regulates the generation and regeneration of hair. In support of this hypothesis, we show that Lhx2 is required for anagen progression and HF morphogenesis. Moreover, transgenic expression of Lhx2 in postnatal HFs is sufficient to induce anagen. Thus, our results reveal an alternative interpretation of Lhx2 function in HFs compared to previously published results, since Lhx2 is periodically expressed, primarily in precursor cells distinct from those in the bulge region, and is an essential positive regulator of hair formation.

Sphere formation restores and confers hair-inducing capacity in cultured mesenchymal cells.

Interactions between epithelial and dermal cells are essential for hair follicle morphogenesis and maintenance. In experimental trials of hair regeneration, isolated dermal cells have been shown to possess hair-inducing capacity. However, dermal cells lose this potential immediately after cultivation. Sphere-forming multipotent cells derived from the dermis possess hair-inducing capacity. These previous findings raise the question of whether hair-inducing capacity depends on the identity as dermal cells or the process of sphere formation. To address this issue, we compared the in vitro and in vivo characteristics of two-dimensionally cultured or thereafter sphere formation-induced dermal and lung mesenchymal cells. We show that sphere-forming mesenchymal cells exhibited higher expression of Wnt signalling genes. Sphere-forming cells but not two-dimensionally cultured cells possessed in vivo hair-inducing capacity. These data suggest that various mesenchymal cells attain hair-inducing capacity through the process of sphere formation.

Frizzled signaling and cell-cell interactions in planar polarity.

The function of the Frizzled pathway is essential for the formation of the array of distally pointing hairs found on the Drosophila wing. Previous research found that regulating the subcellular location for hair initiation controlled hair polarity. Recent work argues a graded Frizzled-dependent signal results in the accumulation of the Frizzled, Dishevelled and Flamingo proteins along the distal edge of the wing cells. This cortical mark leads to the local activation of downstream gene products and the subsequent activation of the cytoskeleton to form a hair.

TNF superfamily in skin appendage development.

The development of skin appendages such as hairs, teeth, and mammary glands is regulated by signaling molecules of the Wnt, FGF, TGFbeta, and Hedgehog pathways. Last decade has also revealed a pivotal role for the TNF family ligand ectodysplasin (Eda) in multiple steps of epithelial appendage morphogenesis, from initiation to differentiation. Surprisingly, other members of the TNF superfamily such as Rank ligand, lymphotoxins, and TNF have recently been linked with specific aspects of skin appendage biology including branching of the mammary gland, hair shaft formation, and hair follicle cycling. This review focuses on the novel discoveries of Eda and other TNF related cytokines in skin appendage development made since the previous review on this topic in Cytokine and Growth Factor reviews in 2003.

Does D matter? The role of vitamin D in hair disorders and hair follicle cycling.

BACKGROUND: The role of vitamin D in the proliferation and differentiation of keratinocytes is well known within the field of dermatology. OBJECTIVE: We sought to evaluate the role that vitamin D and the vitamin D receptor play in the hair cycle and assess how this can be clinically applied to the treatment of hair disorders. METHODS: A MEDLINE search (1955-July 2009) was preformed to find relevant articles pertaining to vitamin D, the vitamin D receptor, and hair loss. RESULTS: The vitamin D receptor, independent of vitamin D, plays an important role in hair cycling, specifically anagen initiation. The role of vitamin D in hair follicle cycling is not as well understood. LIMITATIONS: The review is broad and there are limited human studies available to date. CONCLUSION: Additional studies to evaluate the role of vitamin D in the hair cycle should be done. Treatments that up regulate the vitamin D receptor may be successful in treating hair disorders and are a potential area of further study.

Winging it--actin on the fly.

How actin dynamics might be regulated to generate specific cellular structures during development remains something of a mystery. New insights may be gained from the recent identification of a conserved cofilin phosphatase, Slingshot, which modulates actin dynamics to help control Drosophila wing hair morphogenesis.

Defect of hepatocyte growth factor activator inhibitor type 1/serine protease inhibitor, Kunitz type 1 (Hai-1/Spint1) leads to ichthyosis-like condition and abnormal hair development in mice.

Hepatocyte growth factor activator inhibitor type 1 (HAI-1)/serine protease inhibitor, Kunitz type 1 (SPINT1) is a membrane-bound, serine proteinase inhibitor initially identified as an inhibitor of hepatocyte growth factor activator. It also inhibits matriptase and prostasin, both of which are membrane-bound serine proteinases that have critical roles in epidermal differentiation and function. In this study, skin and hair phenotypes of mice lacking the Hai-1/Spint1 gene were characterized. Previously, we reported that the homozygous deletion of Hai-1/Spint1 in mice resulted in embryonic lethality attributable to impaired placental development. To test the role of Hai-1/Spint1 in mice, the placental function of Hai-1/Spint1-mutant mice was rescued. Injection of Hai-1/Spint1(+/+) blastocysts with Hai-1/Spint1(-/-) embryonic stem cells successfully generated high-chimeric Hai-1/Spint1(-/-) embryos (B6Hai-1(-/-High)) with normal placentas. These embryos were delivered without apparent developmental abnormalities, confirming that embryonic lethality of Hai-1/Spint1(-/-) mice was caused by placental dysfunction. However, newborn B6Hai-1(-/-High) mice showed growth retardation and died by 16 days. These mice developed scaly skin because of hyperkeratinization, reminiscent of ichthyosis, and abnormal hair shafts that showed loss of regular cuticular septation. The interfollicular epidermis showed acanthosis with enhanced Akt phosphorylation. Immunoblot analysis revealed altered proteolytic processing of profilaggrin in Hai-1/Spint1-deleted skin with impaired generation of filaggrin monomers. These findings indicate that Hai-1/Spint1 has critical roles in the regulated keratinization of the epidermis and hair development.

Mouse notch: expression in hair follicles correlates with cell fate determination.

Many vertebrate tissues, including skin, are known to develop as a consequence of epithelial-mesenchymal interactions. Much less is known about the role of cell-cell interaction within the epithelial or the mesenchymal compartments in morphogenesis. To investigate cell-cell interactions during skin development, and the potential role of the Notch homolog in this process, we cloned the mouse homolog of Notch (mNotch) and studied its expression pattern, starting as early as mesoderm formation. The novel application of double-labeled in situ hybridization in vertebrates allowed high resolution analysis to follow the fate of mNotch expressing cells directly. In comparison with the distribution of Id mRNA, analysis confirmed that in the hair follicle high levels of mNotch are expressed exclusively in the epithelial compartment. Hair follicle matrix cells start expressing mNotch as different cell types become distinguishable in the developing follicle. mNotch mRNA expression persists throughout the growth phase of the follicle and maintains the same expression profile in the second hair cycle. The cells in the follicle that undergo a phase of high level mNotch expression are in transition from mitotic precursors to several discreet, differentiating cell types. Our observations point out that both in time (during development) and in space (by being removed one cell layer from the dermal papilla) mNotch expression is clearly separated from the inductive interactions. This is a novel finding and suggests that mNotch is important for follicular differentiation and possibly cell fate selection within the follicle.

Expression of epidermal keratins and filaggrin during human fetal skin development.

The major structural proteins of epithelia, the keratins, and the keratin filament-associated protein, filaggrin, were analyzed in more than 50 samples of human embryonic and fetal skin by one-dimensional SDS PAGE and immunoblotting with monoclonal and polyclonal antibodies. Companion samples were examined by immunohistochemistry and electron microscopy. Based on structural characteristics of the epidermis, four periods of human epidermal development were identified. The first is the embryonic period (before 9 wk estimated gestational age), and the others are within the fetal period: stratification (9-14 wk), follicular keratinization (14-24 wk), and interfollicular keratinization (beginning at approximately 24 wk). Keratin proteins of both the acidic (AE1-reactive, type I) and the basic (AE3-reactive, type II) subfamilies were present throughout development. Keratin intermediate filaments were recognized in the tissue by electron microscopy and immunohistochemical staining. Keratins of 50 and 58 kD were present in the epidermis at all ages studied (8 wk to birth), and those of 56.5 and 67 kD were expressed at the time of stratification and increased in abundance as development proceeded. 40- and 52-kD keratins were present early in development but disappeared with keratinization. Immunohistochemical staining suggested the presence of keratins of 50 and 58 kD in basal cells, 56.5 and 67 kD in intermediate cells, and 40 and 52 kD in the periderm as well as in the basal cells between the time of stratification and birth. Filaggrin was first detected biochemically at approximately 15 wk and was localized immunohistochemically in the keratinizing cells that surround hair follicles. It was identified 8-10 wk later in the granular and cornified cell layers of keratinized interfollicular epidermis. These results demonstrate the following. An intimate relationship exists between expression of structural proteins and morphologic changes during development of the epidermis. The order of expression of individual keratins is consistent with the known expression of keratins in simple vs. stratified vs. keratinized epithelia. Expression of keratins typical of stratified epithelia (50 and 58 kD) precedes stratification, and expression of keratins typical of keratinization (56.5 and 67 kD) precedes keratinization, which suggests that their expression marks the tissue commitment to those processes. Because only keratins that have been demonstrated in various adult tissues are expressed during fetal development, we conclude that there are no "fetal" keratins per se.(ABSTRACT TRUNCATED AT 400 WORDS)

Onset of keratin 17 expression coincides with the definition of major epithelial lineages during skin development.

The type I keratin 17 (K17) shows a peculiar localization in human epithelial appendages including hair follicles, which undergo a growth cycle throughout adult life. Additionally K17 is induced, along with K6 and K16, early after acute injury to human skin. To gain further insights into its potential function(s), we cloned the mouse K17 gene and investigated its expression during skin development. Synthesis of K17 protein first occurs in a subset of epithelial cells within the single-layered, undifferentiated ectoderm of embryonic day 10.5 mouse fetuses. In the ensuing 48 h, K17-expressing cells give rise to placodes, the precursors of ectoderm-derived appendages (hair, glands, and tooth), and to periderm. During early development, there is a spatial correspondence in the distribution of K17 and that of lymphoid-enhancer factor (lef-1), a DNA-bending protein involved in inductive epithelial-mesenchymal interactions. We demonstrate that ectopic lef-1 expression induces K17 protein in the skin of adult transgenic mice. The pattern of K17 gene expression during development has direct implications for the morphogenesis of skin epithelia, and points to the existence of a molecular relationship between development and wound repair.

Fish Scales Dictate the Pattern of Adult Skin Innervation and Vascularization.

As animals mature from embryonic to adult stages, the skin grows and acquires specialized appendages, like hairs, feathers, and scales. How cutaneous blood vessels and sensory axons adapt to these dramatic changes is poorly understood. By characterizing skin maturation in zebrafish, we discovered that sensory axons are delivered to the adult epidermis in organized nerves patterned by features in bony scales. These nerves associate with blood vessels and osteoblasts above scales. Osteoblasts create paths in scales that independently guide nerves and blood vessels during both development and regeneration. By preventing scale regeneration and examining mutants lacking scales, we found that scales recruit, organize, and polarize axons and blood vessels to evenly distribute them in the skin. These studies uncover mechanisms for achieving comprehensive innervation and vascularization of the adult skin and suggest that scales coordinate a metamorphosis-like transformation of the skin with sensory axon and vascular remodeling.