Dynamic regulation of retinoic acid-binding proteins in developing, adult and neoplastic skin reveals roles for beta-catenin and Notch signalling

Retinoic acid (RA) signalling is essential for epidermal differentiation; however, the mechanisms by which it acts are largely unexplored. Partitioning of RA between different nuclear receptors is regulated by RA-binding proteins. We show that cellular RA-binding proteins CRABP1 and CRABP2 and the fatty acid-binding protein FABP5 are dynamically expressed during skin development and in adult tissue. CRABP1 is expressed in embryonic dermis and in the stroma of skin tumours, but confined to the hair follicle dermal papilla in normal postnatal skin. CRABP2 and FABP5 are expressed in the differentiating cells of sebaceous gland, interfollicular epidermis and hair follicles, with FABP5 being a prominent marker of sebaceous glands and anagen follicle bulbs. All three proteins are upregulated in response to RA treatment or Notch activation and are negatively regulated by Wnt/β-catenin signalling. Ectopic follicles induced by β-catenin arise from areas of the sebaceous gland that have lost CRABP2 and FABP5; conversely, inhibition of hair follicle formation by N-terminally truncated Lef1 results in upregulation of CRABP2 and FABP5. Our findings demonstrate that there is dynamic regulation of RA signalling in different regions of the skin and provide evidence for interactions between the RA, β-catenin and Notch pathways.     

Activation of RXR and RAR signaling promotes myogenic differentiation of myoblastic C2C12 cells

Differentiation of embryonic and adult myogenic progenitors undergoes a complex series of cell rearrangements and specification events which are controlled by distinct gene regulatory networks. Delineation of the molecular mechanisms that regulate skeletal muscle specification and formation should be important for understanding congenital myopathies and muscular degenerative diseases. Retinoic acid (RA) signaling plays an important role in development. However, the role of RA signaling in adult myogenic progenitors is poorly understood. Here, we investigate the role of RA signaling in regulating myogenic differentiation of myoblastic progenitor cells. Using the mouse myoblast progenitor C2C12 line as a model, we have found that the endogenous expression of most RAR and RXR isotypes is readily detected. While the nuclear receptor co-repressors are highly expressed, two of the three nuclear receptor co-activators and the enzymes involved in RA synthesis are expressed at low level or undetectable, suggesting that the RA signaling pathway may be repressed in myogenic progenitors. Using the α-myosin heavy chain promoter-driven reporter (MyHC-GLuc), we have demonstrated that either ATRA or 9CRA is able to effectively induce myogenic differentiation, which can be synergistically enhanced when both ATRA and 9CRA are used. Upon ATRA and 9CRA treatment of C2C12 cells the expression of late myogenic markers significantly increases. We have further shown that adenovirus-mediated exogenous expression of RARα and/or RXRα is able to effectively induce myogenic differentiation in a ligand-independent fashion. Morphologically, ATRA- and 9CRA-treated C2C12 cells exhibit elongated cell body and become multi-nucleated myoblasts, and even form myoblast fusion. Ultrastructural analysis under transmission electron microscope reveals that RA-treated myogenic progenitor cells exhibit an abundant presence of muscle fibers. Therefore, our results strongly suggest that RA signaling may play an important role in regulating myogenic differentiation.     

毛囊分子调控机制

毛囊是产生毛被的组织,具有周期性,经历生长期、衰退期和休止期三个时期,毛囊与皮脂腺组成毛囊皮脂腺单位。本文从分子层面分别对调控毛囊周期性变化、毛囊皮脂腺发育以及毛囊中影响毛被光泽度及毡结度的因子等给予了详细阐述,这些均为提高山羊绒产量和质量提供了重要思路。     

Dietary vitamin A regulates wingless-related MMTV integration site signaling to alter the hair cycle

Alopecia areata (AA) is an autoimmune hair loss disease caused by a cell-mediated immune attack of the lower portion of the cycling hair follicle. Feeding mice 3–7 times the recommended level of dietary vitamin A accelerated the progression of AA in the graft-induced C3H/HeJ mouse model of AA. In this study, we also found that dietary vitamin A, in a dose dependent manner, activated the hair follicle stem cells (SCs) to induce the development and growth phase of the hair cycle (anagen), which may have made the hair follicle more susceptible to autoimmune attack. Our purpose here is to determine the mechanism by which dietary vitamin A regulates the hair cycle. We found that vitamin A in a dose-dependent manner increased nuclear localized beta-catenin (CTNNB1; a marker of canonical wingless-type Mouse Mammary Tumor Virus integration site family (WNT) signaling) and levels of WNT7A within the hair follicle bulge in these C3H/HeJ mice. These findings suggest that feeding mice high levels of dietary vitamin A increases WNT signaling to activate hair follicle SCs.     

Normal fur development and sebum production depends on fatty acid 2-hydroxylase expression in sebaceous glands

2-Hydroxylated fatty acid (HFA)-containing sphingolipids are abundant in mammalian skin and are believed to play a role in the formation of the epidermal barrier. Fatty acid 2-hydroxylase (FA2H), required for the synthesis of 2-hydroxylated sphingolipids in various organs, is highly expressed in skin, and previous in vitro studies demonstrated its role in the synthesis of HFA sphingolipids in human keratinocytes. Unexpectedly, however, mice deficient in FA2H did not show significant changes in their epidermal HFA sphingolipids. Expression of FA2H in murine skin was restricted to the sebaceous glands, where it was required for synthesis of 2-hydroxylated glucosylceramide and a fraction of type II wax diesters. Absence of FA2H resulted in hyperproliferation of sebocytes and enlarged sebaceous glands during hair follicle morphogenesis and anagen (active growth phase) in adult mice. This was accompanied by a significant up-regulation of the epidermal growth factor receptor ligand epigen in sebocytes. Loss of FA2H significantly altered the composition and physicochemical properties of sebum, which often blocked the hair canal, apparently causing a delay in the hair fiber exit. Furthermore, mice lacking FA2H displayed a cycling alopecia with hair loss in telogen. These results underline the importance of the sebaceous glands and suggest a role of specific sebaceous gland or sebum lipids, synthesized by FA2H, in the hair follicle homeostasis.     

Retinoic acid activates human inducible nitric oxide synthase gene through binding of RARalpha/RXRalpha heterodimer to a novel retinoic acid response element in the promoter

Human inducible nitric oxide synthase (hiNOS) catalyzes nitric oxide (NO) which has a significant effect on tumor suppression and cancer therapy. Here we revealed the detailed molecular mechanism involved in the regulation of hiNOS expression induced by retinoic acid (RA). We showed that RARalpha/RXRalpha heterodimer was important in hiNOS promoter activation, hiNOS protein expression, and NO production. Serial deletion and site-directed mutation analysis revealed two half-sites of retinoic acid response element (RARE) spaced by 5bp located at -172 to -156 in the hiNOS promoter. EMSA and ChIP assays demonstrated that RARalpha/RXRalpha directly bound to this RARE of hiNOS promoter. Our results suggested the identification of a novel RARE in the hiNOS promoter and the roles of the nuclear receptors (RARalpha/RXRalpha) in the induction of hiNOS by RA.     

Vitamin D3 (soltriol) nuclear receptors in abdominal scent gland and skin of Siberian hamster (Phodopus sungorus) localized by autoradiography and immunohistochemistry

In vivo autoradiography with [³H]1,25-dihydroxycholecalciferol (vitamin D, soltriol) and immuno-staining with antibodies to vitamin D receptor were applied to identify specific binding sites in the abdominal scent gland of male Siberian hamster (Phodopus sungorus). Nuclear concentration of radiolabeled hormone and receptor antibodies was observed in the corresponding cell types including basal cells of sebaceous glands, cells of the outer hair sheaths and hair bulbs, and also keratinocytes in the epidermis. Cells of the hair dermal papillae and fibroblasts of the dermis did not show nuclear labeling. There was good correspondence between the autoradiographic and immunohistochemical data. The results indicate the presence of receptors for vitamin D-soltriol and suggest a seasonal regulation of scent gland marking activities by this steroid hormone of sunlight in cooperation with the sex steroid testosterone.     

Inhibition of BMP signaling in P-Cadherin positive hair progenitor cells leads to trichofolliculoma-like hair follicle neoplasias

Background  

Skin stem cells contribute to all three major lineages of epidermal appendages, i.e., the epidermis, the hair follicle, and the sebaceous gland. In hair follicles, highly proliferative committed progenitor cells, called matrix cells, are located at the base of the follicle in the hair bulb. The differentiation of these early progenitor cells leads to specification of a central hair shaft surrounded by an inner root sheath (IRS) and a companion layer. Multiple signaling molecules, including bone morphogenetic proteins (BMPs), have been implicated in this process.     

Mechanisms regulating epidermal stem cells

The skin epidermis contains different appendages such as the hair follicle and the sebaceous glands. Recent studies demonstrated that several types of stem cells (SCs) exist in different niches within the epidermis and maintain discrete epidermal compartments, but the exact contribution of each SC populations under physiological conditions is still unclear. In addition, the precise mechanisms controlling the balance between proliferation and differentiation of epidermal SC still remain elusive. Recent studies provide new insights into these important questions by showing the contribution of hair follicle SC to the sebaceous lineage and the importance of chromatin modifications and micro-RNAs (miRs) in regulating epidermal SCs renewal and differentiation. In this review, we will discuss the importance of these papers to our understanding of the mechanisms that control epidermal SC functions.     

Hair growth in mouse mutants affecting coat texture

The genetic control of hair growth has been studied in mice carrying the following coat texture genes: fz (fuzzy), soc (soft coat), hid (hair interior defect), sa (satin), It (lustrous), Ve (velvet), wa-1 (waved-1), Re (rex), Re ^wc (wavy coat) and pk (plucked).
A general effect on cells of epidermal origin, found in soc/soc and Ve /+ skin samples illustrates how common factors control developmental potential in both the stratum germinativum and the follicle bulb. A direct influence on follicle bulb development is also seen in fz/fz homozygotes in which the dermal papilla functions abnormally. The role of the bulb cells and the dermal papilla in the control of hair shaft calibre is discussed.
hid is a new gene, found in homozygous condition in all mice of the AKR inbred strain. hid and sa appear primarily to be concerned in the differentiation of the medulla.
In the hair waving mutants, waved-1, rex and wavy coat, the processes controlling hair movement within the follicle are disturbed. These genes appear to regulate internal root sheath function. When the normal relationship between internal root sheath and developing hair shaft is disturbed, shaft movement slows, with the subsequent development of shaft calibre abnormalities.
pk acts at the level of the sebaceous gland, disturbing the normal process of hair eruption. The roles of the internal root sheath, external root sheath and the sebaceous gland in hair eruption are discussed.
The abnormal epidermal layer in soc/soc and Ve /+ skin also disturbs hair eruption to a small extent. The resulting abnormalities this causes in hair shaft formation are compared with those found pk/pk samples and also with the similar effects of faulty hair movement in the hair waving mutants. An effect on pigmentation is also described.
The chemistry of keratinization appears to be normal in all these mutants.     

Spatial patterns produced by a reaction-diffusion system in primary hair follicles

This paper is the third in a series examining the role of a reaction-diffusion (RD) system as the principal mechanism providing spatial information for cell differentiation during hair follicle initiation and development and hair fibre formation. A theoretical mechanism is described by which the RD system supplies positional information during hair follicle development. Solutions of the RD system within the primordial follicle are described as well as the sequence of spatial patterns provides the follicle/epidermis boundary conditions required to account for the density and grouping of follicles during initiation. At the same time the spatial patterns are also shown to be capable of providing the positional information which determines various geometrical aspects of follicle development; in particular the development of follicles at an angle to the skin surface and the initiation and location of sweat glands and sebaceous glands on the follicle.     

Morphogenetic effects of 9-cis-retinoic acid on the regenerating limbs of the axolotl

9-cis-retinoic acid has recently been found to be a high affinity ligand for the retinoic X receptor (RXR). RXRs are believed to be involved in metabolic activities rather than in morphogenetic ones. Interestingly, RXR has been found to form heterodimers involving other receptors from the steroid family, such as the thyroid hormone receptor, vitamin D receptor or retinoic acid receptors (RARs). In this paper we examined whether or not 9-cis-retinoic acid had any morphogenetic properties on the regenerating axolotl limb. It is shown that 9-cis-retinoic acid proximalized regenerating limbs and was somewhat more potent in this action than all-trans-retinoic acid. Based on these observations, the possible roles of other receptors during pattern formation is discussed. Correspondence to: P.A. Tsonis     

Manipulation of stem cell proliferation and lineage commitment: visualisation of label-retaining cells in wholemounts of mouse epidermis

Mammalian epidermis is maintained by stem cells that have the ability to self-renew and generate daughter cells that differentiate along the lineages of the hair follicles, interfollicular epidermis and sebaceous gland. As stem cells divide infrequently in adult mouse epidermis, they can be visualised as DNA label-retaining cells (LRC). With whole-mount labelling, we can examine large areas of interfollicular epidermis and many hair follicles simultaneously, enabling us to evaluate stem cell markers and examine the effects of different stimuli on the LRC population. LRC are not confined to the hair follicle, but also lie in sebaceous glands and interfollicular epidermis. LRC reside throughout the permanent region of the hair follicle, where they express keratin 15 and lie in a region of high alpha6beta4 integrin expression. LRC are not significantly depleted by successive hair growth cycles. They can, nevertheless, be stimulated to divide by treatment with phorbol ester, resulting in near complete loss of LRC within 12 days. Activation of Myc stimulates epidermal proliferation without depleting LRC and induces differentiation of sebocytes within the interfollicular epidermis. Expression of N-terminally truncated Lef1 to block beta-catenin signalling induces transdifferentiation of hair follicles into interfollicular epidermis and sebocytes and causes loss of LRC primarily through proliferation. We conclude that LRC are more sensitive to some proliferative stimuli than others and that changes in lineage can occur with or without recruitment of LRC into cycle.     

Topographical and developmental studies on target sites of 1,25 (OH2) vitamin D3 in skin

Summary Tritium-labeled 1,25 (OH₂) vitamin D₃, when injected into vitamin D-deficient adult and pregnant rats is concentrated and retained strongest in nuclei of cells in the outer root sheath of the hair, followed by the stratum granulosum, spinosum, and basale of the epidermis. In the hair follicle, in addition to the most heavily labeled outer root sheath, nuclear labeling exists also in cells of the hair bulb and of the inner root sheath, as well as in basal cells of the sebaceous gland. In contrast, cells of the dermal papilla and the connective tissue of the dermis are generally unlabeled, except for labeled cells in the outer connective tissue sheath at the infundibulum of vibrissae of 20-day fetal rats and a few scattered labeled cells in the dermis, probably macrophages. In the developing hair, in 18- and 20-day fetal rats, a distinct topographic pattern of labeled cells can be seen, which is characteristic of the different stages of hair follicle development. In the hair germ, heavily labeled cells appear first in the stratum spinosum. In the hair peg, they remain in this position in its juxtaepidermal portion; however, when a dermal papilla develops, heavily labeled cells assume a marginal position. This suggests a sequential epidermal-epidermal and mesenchymal-epidermal receptor induction. Injection of tritium labeled 25 (OH) vitamin D₃ did not show nuclear concentration in these tissues and excess unlabeled 25 (OH) vitamin D₃ — unlike excess 1,25 (OH₂) vitamin D₃ — did not prevent nuclear uptake of tritium labeled 1,25 (OH₂) vitamin D₃. The results indicate differential effects of 1,25 (OH₂) vitamin D₃ on different structures in the epidermis and dermis.Supported by US PHS grant PCM8200569     

Cellular dynamics of the outer layers of the hair follicle of fine-wool sheep during the phase of stable hair growth

The structure, origin, and migration of outer sheath cells of the hair follicles of domestic sheep were studied by electron microscopic, autoradiographic, and histochemical (glycogen) methods in order to understand the role of this layer in hair morphogenesis. We demonstrated that the cells of the outer layers of the outer sheath interpose into the inner “companion” layer of the outer sheath. Although this process takes place all along the hair follicle from the lower bulb up to the sebaceous glands orifices, it mainly takes place over the bulb. Labeled cells interposed into the companion layer reach sebaceous glands orifices more than 24 h faster than labeled cells of the inner sheath and hair, because these cells included the label not in the bulb cambium (as hair and inner sheath) but over the bulb, and from this point they start movement. Interposition of cells into the companion layer must cause increase of its volume and additional volume supposed to be led away into the pillar canal around the hair near the sebaceous glands orifices. This can provide the mechanism of the hair and inner sheath promotion to sebaceous gland orifices.     

The involvement of cytochrome p450 (CYP) 26 in the retinoic acid metabolism of human epidermal keratinocytes

All-trans retinoic acid (RA) levels are controlled by enzymes of the vitamin A metabolism (RDH16, RalDH2, and LRAT) and RA catabolism (CYP26 and CYP2S1). Here, the mRNA expression of these enzymes was investigated in human keratinocytes at different Ca²⁺concentrations and after exposure to RA and CYP26 inhibitors. Cellular differentiation (high Ca²⁺) increased the expression of LRAT, RDH16 and RalDH2, and decreased CYP26B1. RA (1 μM) induced CYP26A1, CYP26B1, CYP2S1, CRABPII and LRAT mRNA. The CYP26 inhibitor talarozole altered CYP26A1 and LRAT mRNA expression in a similar way as RA, increased the cellular accumulation of [³H]RA, and induced a punctate CRABPII staining, also observed after siRNA knock-down of CYP26B1 (but not after RA exposure). Furthermore, CYP26B1 siRNA increased the accumulation of [³H]RA and the CRABPII mRNA, suggesting an augmented retinoid signalling. Thus CYP26B1 appears essential for RA catabolism under physiological conditions, whereas CYP26A1 might play a greater role during RA excess.     

Glutamate transporter Slc1a3 mediates inter‐niche stem cell activation during skin growth

Tissues contain distinct stem cell niches, but whether cell turnover is coordinated between niches during growth is unknown. Here, we report that in mouse skin, hair growth is accompanied by sebaceous gland and interfollicular epidermis expansion. During hair growth, cells in the bulge and outer root sheath temporarily upregulate the glutamate transporter SLC1A3, and the number of SLC1A3⁺ basal cells in interfollicular epidermis and sebaceous gland increases. Fate mapping of SLC1A3⁺ cells in mice revealed transient expression in proliferating stem/progenitor cells in all three niches. Deletion of slc1a3 delays hair follicle anagen entry, uncouples interfollicular epidermis and sebaceous gland expansion from the hair cycle, and leads to reduced fur density in aged mice, indicating a role of SLC1A3 in stem/progenitor cell activation. Modulation of metabotropic glutamate receptor 5 activity mimics the effects of SLC1A3 deletion or inhibition. These data reveal that stem/progenitor cell activation is synchronized over distinct niches during growth and identify SLC1A3 as a general marker and effector of activated epithelial stem/progenitor cells throughout the skin.