珊瑚姜提取物对C3H/He小鼠毛发生长周期及其对小鼠触须毛囊体外培养的影响

目的：探讨中药珊瑚姜对C3H／He小鼠毛发生长周期及其对小鼠触须毛囊体外培养的作用。方法：采用体外局部直接给药的方式，观察珊瑚姜提取物对C3H／He小鼠毛发生长的影响。结果：肉眼观察19d以后，2．0％珊瑚姜用药组小鼠毛已全部长满，0．5％珊瑚姜用药组小鼠也有长毛，较空白组多而长，毛色较空白对照组深，生长速度明显加快，提前进入生长期（P＜0．05）。病理切片显示用药15d后，用药组动物皮肤真皮层黑素分布较多，毛发处于生长期，而空白对照组黑素分布较少，毛发大多处于休止期。结论：珊瑚姜提取物具有明显的促毛发生长作用。     

Hair cycle-dependent changes in adrenergic skin innervation, and hair growth modulation by adrenergic drugs

Skin nerves may exert "trophic" functions during hair follicle development, growth, and/or cycling. Here, we demonstrate hair cycle-related plasticity in the sympathetic innervation of skin and hair follicle in C57BL/6 mice. Compared with telogen skin, the number of nerve fibers containing norepinephrine or immunoreactive for tyrosine hydroxylase increased during the early growth phase of the hair cycle (anagen) in dermis and subcutis. The number of these fibers declined again during late anagen. beta2-adrenoreceptor-positive keratinocytes were transiently detectable in the noncycling hair follicle epithelium, especially in the isthmus and bulge region, but only during early anagen. In early anagen skin organ culture, the beta2-adrenoreceptor agonist isoproterenol promoted hair cycle progression from anagen III to anagen IV. The observed hair cycle-dependent changes in adrenergic skin innervation on the one hand, and hair growth modulation by isoproterenol, accompanied by changes in beta2-adrenoreceptor expression of selected regions of the hair follicle epithelium on the other, further support the concept that bi-directional interactions between the hair follicle and its innervation play a part in hair growth control. This invites one to systematically explore the neuropharmacologic manipulation of follicular neuroepithelial interactions as a novel therapeutic strategy for managing hair growth disorders.     

Intact hair follicle innervation is not essential for anagen induction and development

Abstract Neuropeptides produced, stored and secreted by the unusually dense sensory and autonomic innervation of hair follicles (HFs) can induce hair growth (anagen) and may be involved in hair growth control. To test the role of follicle innervation of HF cycling in vivo, we generated innervation-deficient HFs by unilateral surgical denervation of a defined region of back skin in C57BL/6 mice and assessed its effect on spontaneous and induced anagen development. Successful denervation was demonstrated by the absence of PGP 9.5+ or tyrosine hydroxylase+ nerves and nerve-associated neuropeptides (substance P, CGRP). By quantitative histomorphometry, no significant difference in spontaneous or cyclosporin A-induced anagen development could be detected between sham-operated control skin and denervated skin. Only after hair growth induction by depilation, a discrete, marginally significant retardation of anagen development was apparent in denervated HFs. Thus, even though cutaneous nerves may exert a minor modulatory role in depilation-induced hair growth, they are not essential for normal murine anagen development. Received: 4 March 1998     

PDGF isoforms induce and maintain anagen phase of murine hair follicles

BACKGROUND: It is known that platelet-derived growth factor (PDGF) receptors are expressed in hair follicle (HF) epithelium. OBJECTIVES: The aim of the present study was to clarify the effects of PDGF-AA and -BB on the cyclic growth of HFs. METHODS: PDGF-AA or -BB was injected into the dorsal skin of C3H mice during the second telogen phase once daily for five consecutive days, or PDGF-AA or -BB dissolved in hyaluronic acid was injected only once. In order to confirm the effects of different PDGF isoforms, anti-PDGF-AA antibody or anti-PDGF-BB antibody was injected just after each injection of PDGF-AA or -BB. In addition, anti-PDGF antibodies were injected into the skin of C3H mice during the second anagen phase once daily for 5 days. We studied expression of signaling molecules in the skin where anagen phase had been induced by PDGF injection by real-time RT-PCR. RESULTS: Both PDGF-AA and -BB injection experiments immediately induced the anagen phase of the hair growth cycle at the injection sites. The induction of anagen was interfered by anti-PDGF antibody treatment. Real-time RT-PCR using extracted RNA from the PDGF injected sites of skin samples showed upregulated expression of HF differentiation-related key signaling molecules, Sonic hedgehog (Shh), Lef-1 and Wnt5a. CONCLUSIONS: These results indicate that both PDGF-AA and -BB are involved in the induction and maintenance of the anagen phase in the mouse hair cycle. Local application of PDGF-AA and -BB might therefore prove to be an effective treatment option for alopecia associated with early catagen induction and elongated telogen phase.     

Human scalp skin and hair follicles express neurotrophin-3 and its high-affinity receptor tyrosine kinase C, and show hair cycle-dependent alterations in expression

BACKGROUND: Neurotrophin (NT)-3 and its high-affinity receptor tyrosine kinase C (Trk C) are essential for nervous system development. These members of the NT family are also involved in murine hair morphogenesis and cycling. However, their role in human hair follicle (HF) biology remains to be elucidated. OBJECTIVES: To explore the role of NTs in human skin and HF biology. METHODS: The immunoreactivity (IR) of NT-3 and Trk C was studied in human scalp skin and HFs by immunofluorescent and light microscopic immunohistology. Skin biopsies were obtained from normal human scalp containing mainly anagen VI HFs from women (age 53-57 years) undergoing elective plastic surgery. RESULTS: Both NT-3 and Trk C showed prominent, yet distinct, IR patterns in human scalp anagen HFs (anagen VI), whereas they were weakly expressed in catagen and increased again in telogen HFs. Within HF compartments, NT-3 IR was prominent in the outer root sheath, inner root sheath, dermal papilla and connective tissue sheath. Trk C IR was prominent in all HF epithelial and mesenchymal compartments. Outside the HF, both NT-3 and Trk C showed prominent IR in the epidermis, sebaceous glands and sweat glands. CONCLUSIONS: These observations provide the first indication that NT-3 and Trk C are expressed in human scalp skin and HFs, and suggest that Trk C-mediated signalling is involved not only in murine but also in human HF biology. They may be useful in determining therapeutic strategies for the treatment of hair cycle and skin-related disorders.     

Hair follicle is a target of stress hormone and autoimmune reactions

Interest in the hair follicle (HF) has recently increased, yet the detailed mechanisms of HF function and immune privilege (IP) have not yet been elucidated. This review discusses the critical points of immunobiology and hormonal aspects of HFs. The HF is a unique mini-organ because it has its own immune system and hormonal milieu. In addition, the HF immune and hormonal systems may greatly affect skin immunobiology. Therefore, knowledge of HF immunobiology and hormonal aspects will lead to a better understanding of skin biology. The HF has a unique hair cycle (anagen, catagen and telogen) and contains stem cells in the bulge area. The HF is closely related to sebaceous glands and the nervous system. This article reviews the interaction between the endocrine/immune system and HFs, including the pathogenesis of alopecia areata associated with stress.     

Expression of decorin throughout the murine hair follicle cycle: hair cycle dependence and anagen phase prolongation

Decorin is a prototypical member of the small leucine‐rich proteoglycan (SLRP) family, which is involved in numerous biological processes. The role of decorin, as a representative SLRP, in hair follicle morphogenesis has not been elucidated. We present our initial findings on decorin expression patterns during induced murine hair follicle (HF) cycles. It was found that decorin expression is exclusively restricted to the epidermis, outer root sheath and sebaceous glands during the anagen phase, which correlates with the upregulation of decorin mRNA and protein expression in depilated murine dorsal skin. Furthermore, we used a functional approach to investigate the effects of recombinant human decorin (rhDecorin) via cutaneous injection into HFs at various murine hair cycle stages. The local injection of rhDecorin (100 μg/ml) into the hypodermis of depilated C57BL/6 mice at anagen delayed catagen progression. In contrast, rhDecorin injection during the telogen phase caused the premature onset of anagen, as demonstrated by the assessment of the following parameters: (i) hair shaft length, (ii) follicular bulbar diameter, (iii) hair follicle cycling score and (iv) follicular phase percentage. Taken together, our results suggest that decorin may modulate follicular cycling and morphogenesis. In addition, this study also provides insight into the molecular control mechanisms governing hair follicular epithelial–mesenchymal interactions.     

Clues that mitochondria are involved in the hair cycle clock: MPZL3 regulates entry into and progression of murine hair follicle cycling

The molecular nature of the hair cycle clock (HCC), the intrinsic oscillator system that drives hair follicle (HF) cycling, remains incompletely understood; therefore, all relevant key players need to be identified. Here, we present evidence that implicates myelin protein zero-like 3 (MPZL3), a multifunctional nuclear-encoded mitochondrial protein known to be involved in epidermal differentiation, in HCC regulation. By analysing global Mpzl3 knockout (−/−) mice, we show that in the absence of functional MPZL3, mice commence HF cycling with retarded first catagen-telogen transition after normal postnatal HF morphogenesis. However, Mpzl3 −/− mice subsequently display strikingly accelerated HF cycling, i.e. a precocious telogen-to-anagen transition during the second hair cycle, compared to controls, suggesting that MPZL3 inhibits anagen entry. We also show that intrafollicular MPZL3 protein expression fluctuates in a hair cycle-dependent manner. In telogen HFs, MPZL3 is localized to the secondary hair germ, an epicentre of hair cycle regulation, where it partially co-localizes with P-cadherin. In early anagen HF, MPZL3 is localized immediately distal to the proximal hair matrix. These findings introduce the novel concept that mitochondria are more actively involved in hair cycle control than previously recognized and that MPZL3 plays a central role in the HCC.     

Telogen skin contains an inhibitor of hair growth

We have investigated whether C57B1-6 mouse skin with all its follicles in the telogen stage of the hair cycle contains a hair-growth inhibitory activity, as opposed to skin with anagen follicles. Crude aqueous extracts of whole telogen mouse skin (TE), anagen skin (AE) or vehicle alone (V) were injected intraperitoneally into mice in which anagen had previously been induced by plucking of telogen hair follicles. Injection of TE, but not AE or V, significantly retarded the development of anagen follicles, as measured by macroscopic and quantitative microscopic hair growth parameters (skin pigmentation and thickness, appearance of trichohyaline granules) and the incorporation of tritiated thymidine into mouse skin from animals previously treated with either TE or V (skin organ culture). This inhibitory activity seemed to be localized to the epidermis and was also present in rat epidermis. We suggest that this apparently non-species-specific inhibitor present in telogen skin may play a role in regulating the hair cycle in rodents.     

Comprehensive analysis of FGF and FGFR expression in skin: FGF18 is highly expressed in hair follicles and capable of inducing anagen from telogen stage hair follicles

We quantified the mRNA expression of all 22 fibroblast growth factor family members (FGF) and their four receptors (FGFR) in adult mouse full-thickness skin at various stages of the hair growth cycle. We found that in addition to mRNA encoding FGF previously identified in skin (FGF1, 2, 5, 7, 10, 13, and 22), FGF18 mRNA was also strongly expressed. Expression of these FGF varied throughout hair growth cycle: mRNA expression of FGF18 and 13 peaked at telogen; FGF7 and 10 at anagen V; and FGF5 and 22 at anagen VI. In situ hybridization revealed that FGF18 mRNA is mainly expressed in the anagen inner root sheath and telogen bulge of hair follicles. In culture, FGF18 stimulated DNA synthesis in human dermal fibroblasts, dermal papilla cells, epidermal keratinocytes and vascular endothelial cells. When FGF18 was administered subcutaneously to mice in a uniform telogen state, anagen hair growth was observed. Our findings suggest that FGF18 is important for the regulation of hair growth and the maintenance of skin in adult mice.     

Methods in hair research: how to objectively distinguish between anagen and catagen in human hair follicle organ culture

Please cite this paper as: Methods in hair research: how to objectively distinguish between anagen and catagen in human hair follicle organ culture. Experimental Dermatology 2010; 19: 305–312. Abstract: The organ culture of human scalp hair follicles (HFs) is the best currently available assay for hair research in the human system. In order to determine the hair growth‐modulatory effects of agents in this assay, one critical read‐out parameter is the assessment of whether the test agent has prolonged anagen duration or induced catagen in vitro. However, objective criteria to distinguish between anagen VI HFs and early catagen in human HF organ culture, two hair cycle stages with a deceptively similar morphology, remain to be established. Here, we develop, document and test an objective classification system that allows to distinguish between anagen VI and early catagen in organ‐cultured human HFs, using both qualitative and quantitative parameters that can be generated by light microscopy or immunofluorescence. Seven qualitative classification criteria are defined that are based on assessing the morphology of the hair matrix, the dermal papilla and the distribution of pigmentary markers (melanin, gp100). These are complemented by ten quantitative parameters. We have tested this classification system by employing the clinically used topical hair growth inhibitor, eflornithine, and show that eflornithine indeed produces the expected premature catagen induction, as identified by the novel classification criteria reported here. Therefore, this classification system offers a standardized, objective and reproducible new experimental method to reliably distinguish between human anagen VI and early catagen HFs in organ culture.     

Characterization of dermal type I collagen of C3H mouse at different stages of the hair cycle

Hair follicles develop or regress in accordance with the hair cycle. In this study, we partially characterized fibrillar type I collagen, the predominant component in the dermis, at two stages of the hair cycle: anagen and telogen. Skin samples were obtained from the backs of two groups of 11-week-old C3H mice: one at anagen stage induced by shaving and the other at telogen stage. The amount of neutral salt-soluble (newly synthesized) collagen obtained from anagen skin was about twofold that from telogen skin, while the level of acid-soluble collagen was not significantly different between the two groups. The degree of lysine hydroxylation of pepsinized type I collagen obtained from anagen skin was significantly higher than that in telogen (5.0% higher in alpha1 chain, and 15.6% higher in alpha2 chain). Proline hydroxylation at the anagen stage was also slightly higher than in the telogen stage. Two major collagen cross-links were found in both groups of skin; dehydro-hydroxylysinonorleucine and dehydro-histidinohydroxymerodesmosine. The concentration of the latter, a complex tetravalent cross-link, was significantly lower in anagen skin when compared with telogen skin (mean +/- SD 0.64 +/- 0. 07 vs. 0.78 +/- 0.06 mol/mol collagen). The former showed no significant difference between the two groups. In addition, a significant amount of lysyl-aldehyde (a cross-link precursor) was found in anagen (0.16 +/- 0.02 mol/mol collagen), while it was 0.12 mol/mol collagen in telogen. These results indicate that the remodelling of collagen is more active in anagen skin than in telogen, and that characteristic post-translational modifications of dermal collagen seen in anagen may play a part in facilitating an environment around hair follicles for their migration and growth.     

Expression of nerve growth factor and its high-affinity receptor, tyrosine kinase A proteins, in the human scalp skin

BACKGROUND: Nerve growth factor (NGF) and its high-affinity receptor, tyrosine kinase A (TrkA), are members of the neurotrophin family. NGF-TrkA are involved in murine hair morphogenesis and cycling. To date, their expression in human hair follicle (HF) is unknown. In this investigation, we hypothesize that NGF-TrkA proteins are expressed in the human scalp skin. Moreover, NGF-TrkA expression in HF changes with the transitions from anagen-->catagen-->telogen stages. MATERIALS AND METHODS: To test our hypothesis and to fill this existing gap in literature, the immunostaining values (semiquantitative evaluation of protein expression: SI, staining intensity; PP, percentage of positive cells; and IR score, immunoreactivity score) of NGF and TrkA proteins were examined in human scalp skin by immunofluorescent and immunoperoxidase staining methods. Fifty normal human scalp skin biopsy specimens were examined (healthy females, 53-57 years). In each case, 50 HFs were analyzed (35, 10, and five follicles in anagen, catagen, and telogen, respectively). RESULTS: The IR scores were statistically significantly higher (p < 0.001) in anagen as compared with either catagen or telogen HF (9.61 +/- 0.12 vs. 1.4 +/- 0.10 vs. 0.6 +/- 0.10 for NGF and 3.31 +/- 0.02 vs. 0.5 +/- 0.10 vs. 0.2 +/- 0.10 for TrkA). In the anagen HF, high expression values were seen in the distal region, followed by upper distal, lower distal, and bulb regions for both NGF (10.6 +/- 0.21 vs. 10.3 +/- 0.21 vs. 9.2 +/- 0.40 vs. 8.1 +/- 0.30) and TrkA (3.54 +/- 0.07 vs. 3.45 +/- 0.07 vs. 3.31 +/- 0.06 vs. 3.13 +/- 0.04). Both NGF and TrkA proteins showed prominent expression in the melanocytes (7.6 +/- 0.15 vs. 2.50 +/- 0.07), keratinocytes (10.2 +/- 0.40 vs. 2.71 +/- 0.06), sebaceous glands (10.2 +/- 0.40 vs. 2.72 +/- 0.06), and sweat glands (10.4 +/- 0.40 vs. 2.84 +/- 0.05). CONCLUSIONS: Our findings report, for the first time, the expression pattern of NGF and TrkA proteins in human scalp skin and HF. The differential expression of these proteins during HF cycling suggests their possible roles in human HF biology. The clinical ramifications of these observations mandate further investigations.     

Active hair growth (anagen) is associated with angiogenesis

After the completion of skin development, angiogenesis, i.e., the growth of new capillaries from pre-existing blood vessels, is held to occur in the skin only under pathologic conditions. It has long been noted, however, that hair follicle cycling is associated with prominent changes in skin perfusion, that the epithelial hair bulbs of anagen follicles display angiogenic properties, and that the follicular dermal papilla can produce angiogenic factors. Despite these suggestive observations, no formal proof is as yet available for the concept that angiogenesis is a physiologic event that occurs all over the mature mammalian integument whenever hair follicles switch from resting (telogen) to active growth (anagen). This study uses quantitative histomorphometry and double-immunohistologic detection techniques for the demarcation of proliferating endothelial cells, to show that synchronized hair follicle cycling in adolescent C57BL/6 mice is associated with substantial angiogenesis, and that inhibiting angiogenesis in vivo by the intraperitoneal application of a fumagillin derivative retards experimentally induced anagen development in these mice. Thus, angiogenesis is a physiologic event in normal postnatal murine skin, apparently is dictated by the hair follicle, and appears to be required for normal anagen development. Anagen-associated angiogenesis offers an attractive model for identifying the physiologic controls of cutaneous angiogenesis, and an interesting system for screening the effects of potential antiangiogenic drugs in vivo.     

Injury modifies the fate of hair follicle dermal stem cell progeny in a hair cycle‐dependent manner

The dermal papilla (DP) is one of two principal mesenchymal compartments of the hair follicle (HF). We previously reported that a population of HF dermal stem cells (hfDSCs) function to regenerate the dermal sheath (DS), but intriguingly also contribute new cells to the adult DP at the onset of anagen hair growth to maintain normal cycling of HFs and support the production of large hair fibres. Here, we asked whether injury altered the behaviour of hfDSCs and their progeny in order to support wound‐induced hair growth (WIHG) and if the response was modulated by hair cycle stage. αSMACreER^T2:ROSA^YFP mice received tamoxifen to label the DS, including hfDSCs. Full‐thickness excisions were made on the dorsal skin during various stages of the hair cycle. The skin was harvested at the subsequent anagen. Interestingly, there was an increase in the magnitude of recruitment of hfDSC progeny into the DP after injury compared to follicles entering natural second anagen. This bias towards a DP fate only occurred when a wound was induced during certain stages of the HC. In summary, injury modifies the fate of hfDSCs progeny, biasing them towards recruitment into the DP, with the hair cycle stage also influencing this response.     

Evidence that the bulge region is a site of relative immune privilege in human hair follicles

Background Recent gene profiling data suggest that, besides the anagen hair bulb, the epithelial stem cell region in the outer root sheath of hair follicles (HFs), termed the bulge, may also represent an area of relative immune privilege (IP). Objectives To investigate whether the human HF bulge is a site of relative IP within anagen VI HFs. Methods Anagen VI HFs from normal human scalp skin were analysed using immunohistological staining techniques, quantitative histomorphometry and statistical analysis. For functional evidence we performed full‐thickness human scalp skin organ cultures to investigate whether interferon (IFN)‐γ, a key inducer of IP collapse in hair bulbs, has a similar effect on the putative bulge IP. Results Major histocompatibility complex (MHC) class Ia, β₂‐microglobulin and MHC class II immunoreactivity are downregulated in the human bulge. The immunosuppressants α‐melanocyte stimulating hormone, transforming growth factor‐β2, macrophage migration inhibitory factor and indoleamine‐2,3‐dioxygenase (IDO) are upregulated in the CD200+, stem cell‐rich bulge region. These CD200+ cells also co‐express HLA‐E. Furthermore, IFN‐γ induces significant ectopic MHC class Ia expression in bulge cells of organ‐cultured human scalp skin. Conclusions These data suggest that the bulge of human anagen HFs represents a hitherto unrecognized site of relative IP in human skin. Simultaneously, we present the first evidence of IDO and HLA‐E protein expression in normal human HFs. Bulge IP presumably protects the HF epithelial stem cell reservoir from autoaggressive immune attack whereas a loss of bulge IP may play a central role in the pathogenesis of cicatricial alopecias.     

Do human dermal adipocytes switch from lipogenesis in anagen to lipophagy and lipolysis during catagen in the human hair cycle?

In murine skin, dermal white adipose tissue (DWAT) undergoes fluctuations in size across the hair cycle, whereas changes in size, function and metabolism of dermal adipocytes (DAs) during the human scalp hair cycle remain unexplored. Transmission electron microscopy results suggest that during anagen‐catagen transition, human DAs co‐opt the autophagy machinery to undergo lipophagy within their lipid droplets. Whole‐mount staining of hair follicles (HFs) and surrounding DWAT for the autophagy marker LC3B confirms the increased presence of LC3B+ lipid droplets adjacent to catagen HFs; moreover, DWAT around catagen HFs engages in greater glycerol release compared to DWAT surrounding anagen HFs. Thus, we hypothesize that human DAs switch from lipogenesis during anagen to lipophagy together with lipolysis during catagen. We propose various experiments to further prove this hypothesis, whose systematic exploration should help to better characterize the functions of human DWAT and its communication with the HF.     

Overexpression of skin protein kinase C-alpha in anagen hair follicles during induced growth of mouse hair

A role for protein kinase C (PKC)-alpha has been implicated in the growth of mouse hair. Topical application of PKC activators, hair plucking, allergic contact dermatitis and skin irritation can all enhance growth of mouse hair, and a significant increase in PKC-alpha level in whole mouse skin in mature anagen has been demonstrated in these processes. Overexpression of PKC-alpha in anagen hair follicles has also been reported in natural growth of mouse hair. It is known that overexpression of PKC-alpha is associated with the acceleration of cell growth. Therefore, we postulated that overexpression of PKC-alpha in mature anagen may relate to enhancement of hair growth. The distribution of PKC-alpha in hair follicles during induced growth of mouse hair has not previously been studied. In this study, hair growth in C57BL/6 mice was induced by plucking the telogen hairs on one side of the back. The undepilated contralateral side served as a control. Expression of PKC-alpha in hair follicles during the hair growth cycle induced was evaluated by immunohistochemistry using cryosections and a specific polyclonal anti-PKC-alpha immunoglobulin G (IgG) antibody. No PKC-alpha was detected in telogen hair follicles or in the hair follicles at 1 day post-depilation, when the induced hair cycle was in early anagen. At 4 days after plucking, when the induced hair cycle was in mid-anagen, intense staining for PKC-alpha was found in hair papillae. At 10 and 17 days after depilation, when the induced hair cycle was in mature anagen and early catagen, respectively, all outer root sheath (ORS) cells and outer connective sheaths of hair follicles were stained positive. Because no PKC-alpha was detected in telogen hair follicles in this study, down-regulation of PKC-alpha in early anagen could not be observed. However, consistent with our previous findings, overexpression of PKC-alpha was found in mid-anagen and mature anagen. As overexpression of PKC-alpha has been shown to be associated with acceleration of cell growth, our results support the notion that PKC-alpha may play an important role in growth of hair follicle cells in induced growth of hair. As PKC levels are known to increase in hyperglycaemia, overexpressed PKC-alpha in mature anagen hair follicles may be related to the putative function of the ORS in mobilizing glycogen stores for anagen growth.