Restorative effect of hair follicular dermal cells on injured human hair follicles in a mouse model

No model is available for examining whether in vivo‐damaged human hair follicles (hu‐HFs) are rescued by transplanting cultured hu‐HF dermal cells (dermal papilla and dermal sheath cells). Such a model might be valuable for examining whether in vivo‐damaged hu‐HFs such as miniaturized hu‐HFs in androgenic alopecia are improvable by auto‐transplanting hu‐HF dermal cells. In this study, we first developed mice with humanized skin composed of hu‐keratinocytes and hu‐dermal fibroblasts. Then, a ‘humanized scalp model mouse’ was generated by transplanting hu‐scalp HFs into the humanized skin. To demonstrate the usability of the model, the lower halves of the hu‐HFs in the model were amputated in situ, and cultured hu‐HF dermal cells were injected around the amputated area. The results demonstrated that the transplanted cells contributed to the restoration of the damaged HFs. This model could be used to explore clinically effective technologies for hair restoration therapy by autologous cell transplantation.     

Recapitulation of the hairless mouse phenotype using catalytic oligonucleotides: implications for permanent hair removal

Ribozyme technology is widely used to target mRNA in a sequence-specific fashion and thus change the expression pattern of cells or tissues. While the goal of mRNA targeting is usually the cleavage of mutant mRNAs with the prospect of gene therapy for inherited diseases, in certain instances, targeting of wild-type genes can be used therapeutically. Lack of expression of the mouse hairless gene due to inherited mutations leads to the complete and irreversible loss of hair known as atrichia. We designed this study to recapitulate the hairless phenotype in a restricted manner by topical application of deoxyribozyme-targeting molecules to specifically cleave the mouse hairless mRNA. Histological samples taken from treated skin at different times demonstrated a decreased number of hair follicles, an involution of the remaining follicles, a separation of the dermal papillae, and the presence of dermal cysts, all characteristics of the hairless phenotype, but not normally present in the skin of C57Bl/6 J mice. In this study, we successfully recapitulated the hairless phenotype using topically applied target-specific catalytic oligonucleotides designed to cleave the mouse hairless mRNA. Our results demonstrate the feasibility of using ribozyme technology to alter the gene expression in the skin via topical application and provide proof of principle for the development of this strategy for permanent hair removal.     

An advanced mouse model for human skin wound healing

Here, we report a model for studying wound repair based on skin regenerated from human tissue culture‐expanded cells. The reconstituted skin (hRSK) responds to injury similar to that of intact human skin, and its constituent cells contribute to the healing process. As we have demonstrated that hRSK composed of GFP‐labelled cells also heals “normally,” we believe this model will be useful in analysing the wound repair process using genetically modified human cells.     

Establishment of a mouse xenograft model for mycosis fungoides

Mycosis fungoides (MF) is the most frequent variant of cutaneous T-cell lymphomas (CTCLs). MF primarily involves the skin initially with patches and plaques. In later stages, cutaneous tumors develop and tumor cells may spread to lymph nodes and finally to visceral sites. Here, we describe an animal model for MF in immune-deficient nude mice, using the CTCL cell line MyLa. Subcutaneous transplantation of MyLa cells leads to the formation of cutaneous tumors in 80% of the mice (50/60 total). Spread of tumor cells to visceral sites was detected by immunohistochemistry and polymerase chain reaction (PCR)-based detection of specific T-cell receptor-gamma rearrangement. MyLa cells were found circulating in the blood, lymph nodes, and in blood vessels of heart, kidney, lung, and liver. In lung and liver tissue, tumor cells presented perivascular invasion, but no large secondary tumors developed. The nude mouse model described here will be a valuable test system for new therapeutic approaches for the treatment of MF and opens the unique opportunity to study the disease in vivo.     

Foot shock stress prolongs the telogen stage of the spontaneous hair cycle in a non-depilated mouse model

BACKGROUND: There is an increasing evidence to indicate that stress can influence skin disease and cutaneous functions. Previous studies have shown that stress alters the murine hair cycle; however, these studies have been carried out by using mouse models in which the hair cycle is forcibly synchronized after depilation. OBJECTIVE: To examine whether foot shock stress (FS) changes the spontaneous hair cycle in a non-depilated animal model, and to evaluate the role of mast cells and substance P (SP) in the influence of stress on the hair cycle. METHODS: Changes in the spontaneous hair cycle and the inhibitory effects of a specific SP NK1 receptor antagonist were examined in non-depilated mice during 3-4 weeks of FS. RESULTS: Foot shock stress prolonged the telogen stage of the hair cycle and delayed the induction of the subsequent anagen stage in the animal model. FS caused an increase in the ratio of de-granulated mast cells in the skin, an increase in the number of TUNEL-positive cells, and a decrease in the number of Ki67-positive cells. The NK1 receptor antagonist, WIN 62577, inhibited these stress responses. CONCLUSION: Our results strongly support previous work, demonstrating that stress alters active hair-cycling in vivo through the action of SP.     

A novel model of wound healing in the SCID mouse using a cultured human skin substitute

Studies of skin graft behaviour in rodent excisional wound models are limited by the dominance of wound contracture and graft sloughing as primary healing responses. To slow skin contraction, polytetrafluoroethylene (Teflon) rings were inserted into dorso-lateral full-thickness wounds in SCID mice. Cultured skin substitutes (OrCel), composed of cultured human keratinocytes and fibroblasts in a bovine collagen sponge, were implanted within the rings. Examination and histology of grafts 14 days later showed graft take in four of six recipients, with 90% epithelialization and wound contraction of 31–47%. Immunohistochemical studies, using human-specific antisera to distinguish graft from host tissues, showed that regenerated tissue was predominantly human. Staining with anticytokeratin, revealed a multilayered, stratified neoepidermis. HBG were identified in keratinocytes in all epidermal layers. Langerhans cells were absent. Antihuman vimentin, used as a fibroblast marker, confirmed that cells of the neodermis were primarily of human origin. Neoepidermal keratinocytes, primarily in the basal and suprabasal layers, were also stained. Results suggest that the poly(tetrafluoroethylene) ring inhibited graft sloughing and provided a more favourable environment for the skin substitute to regenerate a substantially normal human skin.     

Daily intake of Jeju groundwater improves the skin condition of the model mouse for human atopic dermatitis

Drinking water is an important nutrient for human health. The mineral ingredients included in drinking water may affect the physical condition of people. Various kinds of natural water are in circulation as bottled water in developed countries; however, its influence on clinical conditions of patients with certain diseases has not been fully evaluated. In this study, effects of the natural groundwater from Jeju Island on clinical symptoms and skin barrier function in atopic dermatitis (AD) were evaluated. NC/Tnd mice, a model for human AD, with moderate to severe dermatitis were used. Mice were given different natural groundwater or tap water for 8 weeks from 4 weeks of age. Clinical skin severity scores were recorded every week. Scratching analysis and measurement of transepidermal water loss were performed every other week. The pathological condition of the dorsal skin was evaluated histologically. Real‐time polymerase chain reaction analysis was performed for cytokine expression in the affected skin. The epidermal hyperplasia and allergic inflammation were reduced in atopic mice supplied with Jeju groundwater when compared to those supplied with tap water or other kinds of natural groundwater. The increase in scratching behavior with the aggravation of clinical severity of dermatitis was favorably controlled. Moreover, transepidermal water loss that reflects skin barrier function was recovered. The early inflammation and hypersensitivity in the atopic skin was alleviated in mice supplied with Jeju groundwater, suggesting its profitable potential on the daily care of patients with skin troubles including AD.     

Substance P as an immunomodulatory neuropeptide in a mouse model for autoimmune hair loss (alopecia areata)

Alopecia areata (AA) is an autoimmune disorder of the hair follicle characterized by inflammatory cell infiltrates around actively growing (anagen) hair follicles. Substance P (SP) plays a critical role in the cutaneous neuroimmune network and influences immune cell functions through the neurokinin-1 receptor (NK-1R). To better understand the role of SP as an immunomodulatory neuropeptide in AA, we studied its expression and effects on immune cells in a C3H/HeJ mouse model for AA. During early stages of AA development, the number of SP-immunoreactive nerve fibers in skin is increased, compared to non-affected mice. However, during advanced stages of AA, the number of SP-immunoreactive nerves and SP protein levels in skin are decreased, whereas the expression of the SP-degrading enzyme neutral endopeptidase (NEP) is increased, compared to control skin. In AA, NK-1R is expressed on CD8+ lymphocytes and macrophages accumulating around affected hair follicles. Additional SP supply to the skin of AA-affected mice leads to a significant increase of mast cell degranulation and to accelerated hair follicle regression (catagen), accompanied by an increase of CD8+ cells-expressing granzyme B. These data suggest that SP, NEP, and NK-1R serve as important regulators in the molecular signaling network modulating inflammatory response in autoimmune hair loss.     

Pemphigus mouse model as a tool to evaluate various immunosuppressive therapies

Abstract:  Pemphigus vulgaris (PV) is an autoimmune bullous disease caused by immunoglobulin G (IgG) autoantibodies against desmoglein 3 (Dsg3). We have generated an active disease mouse model for PV by adoptive transfer of Dsg3^−/− lymphocytes. In this study, we investigated the benefits and limitations of this model as a tool to evaluate various immunosuppressive therapeutic strategies. We used the following three measurements to evaluate the effects of the drugs during the time course: Dsg3 enzyme-linked immunosorbent assay scores that represent the level of production of anti-Dsg3 IgG, body weight loss that reflects the severity of oral erosions and PV score that reflects the extent of skin lesions. We examined various immunosuppressive agents currently used to treat patients with PV model mice in preventive protocol. Cyclophosphamide almost completely suppressed the production of anti-Dsg3 IgG, development of body weight loss and the appearance of the PV phenotype in contrast with the control group without the drug. Azathioprine, cyclosporin A and tacrolimus hydrate also showed suppressive effects to various degrees. However, methylprednisolone and dexamethasone failed to show significant effects in contrast to the findings reported in humans. Knowing the advantages and limitations of this model will provide an important foundation for the future evaluation and development of novel therapeutic strategies.     

Identifying mouse models for skin cancer using the Mouse Tumor Biology Database

In recent years, the scientific community has generated an ever‐increasing amount of data from a growing number of animal models of human cancers. Much of these data come from genetically engineered mouse models. Identifying appropriate models for skin cancer and related relevant genetic data sets from an expanding pool of widely disseminated data can be a daunting task. The Mouse Tumor Biology Database (MTB) provides an electronic archive, search and analysis system that can be used to identify dermatological mouse models of cancer, retrieve model‐specific data and analyse these data. In this report, we detail MTB's contents and capabilities, together with instructions on how to use MTB to search for skin‐related tumor models and associated data.     

Refinement of the relative alkylation index (RAI) model for skin sensitization and application to mouse and guinea-pig test data for alkyl alkanesulphonates

Summary A derivation, more rigorous than hitherto, of the Relative Alkylation Index (RAI) as a quantifier of carrier protein haptenation in skin sensitization tests is presented. It is shown that the RAI, which is a composite parameter made up of dose, reactivity and lipophilicity terms, is likely to require a higher weighting for the reactivity term in the case of non-adjuvant tests than in the case of Freund's adjuvant-based tests. Methyl alkanesulphonates, RSO₃Me with R ranging from n-C₆H₁₃ to n-C₁₆H₃₃, were found to be skin sensitizers in a mouse ear swelling test, in agreement with published findings in a guinea-pig adjuvant model. A structure-activity relationship consistent with the published RAI model was observed whereby, in tests at fixed molar induction (0.1 mM) and challenge concentrations (0.025 mM), the level of sensitization response at first increased with increasing chain length of R, then showed a reversal of this trend at the highest chain length (R=n-C₁₆H₃₃). That this is a genuine overload effect, as reported for several other series of compounds examined in guinea-pig adjuvant models, is indicated by the finding that on reducing the induction concentration for the R=n-C₁₆H₃₃ compound the sensitization response was increased. Alkyl and alkenyl methanesulphonates, MeSO₃ R (R=n-C₁₂H₂₅, n-C₁₈H₃₇ and R=oleyl) did not give significant sensitization in the mouse ear test. Although they are chemically less reactive than methyl alkanesulphonates, these compounds are reported to be strong sensitizers in guinea-pig adjuvant tests and to fit a common quantitative sensitization — structure — dose relationship with the methyl alkanesulphonates. This difference between the murine and the guinea-pig adjuvant tests is attributed to the sensitization potential being more sensitive in the murine test to differences in chemical reactivity, as predicted by the more rigorously derived RAI model.Visiting Professor at the Université Louis Pasteur of Strasbourg, on leave of absence from Unilever, UK     

球形马拉色菌过增殖特应性皮炎小鼠模型的建立

 目的:探讨球形马拉色菌过增殖小鼠特应性皮炎(AD)模型的建立方法。方法：将18只BALB/c小鼠随机分为6组：对照组、AD组、NS/M组、AD+NS/M组、Oil/M组和AD+Oil/M组，每组3只。通过耳皮肤涂布MC903溶液诱导小鼠特应性皮炎模型后，再分别使用橄榄油和生理盐水作为基质配置球形马拉色菌混悬液涂抹小鼠耳皮肤，以构建真菌过增殖模型，并通过观察其临床表现、皮损评分、真菌学和组织病理学变化，扫描/透射电镜观察毛发结果等评估。结果：与AD组相比：AD+Oil/M组小鼠耳部湿疹样皮炎加重，耳周毛发有明显脱落，皮损评分升高（q=5.95, P<0.01）；组织病理学结果显示AD+Oil/M组表皮明显增厚，真皮浅层致密的炎症细胞浸润，伴部分毛囊受损，毛干退化；PAS染色和真菌荧光染色可见角质层内大量球形马拉色菌；扫描/透射电镜下见毛干表面及毛干内均有大量球形马拉色菌。NS/M组和AD+NS/M组小鼠耳组织PAS染色和真菌荧光染色仅见散在数个马拉色菌。结论：应用橄榄油混悬球形马拉色菌液涂布皮肤可成功建立马拉色菌过增殖小鼠特应性皮炎模型。     

Lack of evidence for allergenic properties of coumarin in a fragrance allergy mouse model

BACKGROUND: There is controversy as to whether coumarin, an ingredient in cosmetics and fragrances, is a contact allergen involved in fragrance allergy. We recently showed that the purity of coumarin is a critical parameter for its allergenicity because coumarin preparations containing trace amounts of contaminants induced cell proliferation in the local lymph node (LN) assay whereas pure coumarin did not. OBJECTIVE/METHOD: In the present study, we analyzed the sensitizing properties of coumarin (purity > 99.9) and of dihydrocoumarin (DHC), in a recently developed model of fragrance allergy in mice. RESULTS: DHC was able to prime T cells in LNs draining the sensitization skin site and to induce a typical allergic contact dermatitis (ACD) reaction upon challenge, confirming that DHC is endowed with moderate sensitizing properties. In contrast, no T-cell activation and no ACD responses were obtained following sensitization and challenge with coumarin. CONCLUSION: These results confirm that pure coumarin is endowed with very weak sensitizing capacities, if any, and suggest that the presence of contaminants in coumarin preparations may account for the previously reported allergenic properties of coumarin.     

Comparison between cold plasma,electrochemotherapy and combined therapy in a melanoma mouse model

The study was undertaken to compare antitumor efficacy of electrochemotherapy (ECT) with cold plasma therapy (CP) in a melanoma mouse model. After melanoma implantation into the flank of C57BL/6N mice, CP by two different plasma sources (APPJ and DBD) was applied directly to the tumor surface. ECT was performed with bleomycin intravenously at a field strength of 1000 V/cm without or combined with CP. Primary endpoints were tumor growth acceleration (TGA), daily volume progression (DVP) and survival after treatment. Both plasma sources as single treatment showed a significant TGA delay, which proved less effective than ECT. CP (APPJ) combined with ECT (ECJ) significantly improved per cent mouse survival, with significant superiority compared with ECT. Plasma therapy alone albeit less effective seems a potential alternative to ECT in patients with melanoma and can be applied manifold in a session without general anaesthesia. Accordingly, CP alone and combined with ECT may serve as new option in palliative skin melanoma therapy.     

Highly pigmented Tg(Grm1) mouse melanoma develops non‐pigmented melanoma cells in distant metastases

Murine model systems are critically required tools for the investigation of unknown mechanisms of melanoma development and metastasis and for developing more efficient therapies. The Tg(Grm1)EPv melanoma mouse model is characterized by spontaneous development of pigmented cutaneous melanomas at hairless skin regions, with a short latency and 100% penetrance. Local metastasis was described in initial analyses; however, melanoma cells were not observed in distant organs. Here, we demonstrate that the established Tg(Grm1)EPv melanoma mouse model exhibits more extensive metastasis into distant organs than previously described. Disseminated cells undergo phenotypic changes, as we observed high numbers of non‐pigmented Grm1‐expressing melanoma cells within distant organs. As such changes during metastasis are common in human melanoma, our findings demonstrate that this mouse model represents an even more useful tool to study unknown mechanisms of metastasis in human melanoma than previously assumed.     

Hair growth inhibition by psychoemotional stress: a mouse model for neural mechanisms in hair growth control

Stress has long been discussed controversially as a cause of hair loss. However, solid proof of stress-induced hair growth inhibition had long been missing. If psychoemotional stress can affect hair growth, this must be mediated via definable neurorendocrine and/or neuroimmunological signaling pathways. Revisiting and up-dating relevant background data on neural mechanisms of hair growth control, we sketch essentials of hair follicle (HF) neurobiology and discuss the modulation of murine hair growth by neuropeptides, neurotransmitters, neurotrophins, and mast cells. Exploiting an established mouse model for stress, we summarize recent evidence that sonic stress triggers a cascade of molecular events including plasticity of the peptidergic peri- and interfollicular innervation and neuroimmune crosstalk. Substance P (SP) and NGF (nerve growth factor) are recruited as key mediators of stress-induced hair growth-inhibitory effects. These effects include perifollicular neurogenic inflammation, HF keratinocyte apoptosis, inhibition of proliferation within the HF epithelium, and premature HF regression (catagen induction). Intriguingly, most of these effects can be abrogated by treatment of stressed mice with SP-receptor neurokinin-1 receptor (NK-1) antagonists or NGF-neutralizing antibodies - as well as, surprisingly, by topical minoxidil. Thus there is now solid in vivo-evidence for the existence of a defined brain- HF axis. This axis can be utilized by psychoemotional and other stressors to prematurely terminate hair growth. Stress-induced hair growth inhibition can therefore serve as a highly instructive model for exploring the brain-skin connection and provides a unique experimental model for dissecting general principles of skin neuroendocrinology and neuroimmunology well beyond the HF.     

Targeted silencing of DEFB4 in a bioengineered skin‐humanized mouse model for psoriasis: development of siRNA SECosome‐based novel therapies

Psoriasis is a complex inflammatory skin disease that presents a wide variety of clinical manifestations. Human β defensin‐2 (hBD‐2) is highly up‐regulated in psoriatic lesions and has been defined as a biomarker for disease activity. We explored the potential benefits of targeting hBD‐2 by topical application of DEFB4‐siRNA‐containing SECosomes in a bioengineered skin‐humanized mouse model for psoriasis. A significant improvement in the psoriatic phenotype was observed by histological examination, with a normalization of the skin architecture and a reduction in the number and size of blood vessels in the dermal compartment. Treatment leads to the recovery of transglutaminase activity, filaggrin expression and stratum corneum appearance to the levels similar to those found in normal regenerated human skin. The availability of a reliable skin‐humanized mouse model for psoriasis in conjunction with the use of the SECosome technology may provide a valuable preclinical tool for identifying potential therapeutic targets for this disease.     

茶多酚、吡美莫司、他克莫司外用对莫诺苯宗诱导白癜风样模型小鼠的疗效比较

目的 探讨茶多酚、吡美莫司、他克莫司对莫诺苯宗诱导的白癜风样模型小鼠的疗效差异。 方法 45%莫诺苯宗诱导C57BL/6三周龄雌小鼠脱色,建立白癜风样动物模型。并研究他克莫司、吡美莫司、茶多酚对白癜风样模型小鼠脱色的治疗效果。通过肉眼观察毛发脱色及脱色面积,实验结束后取非用药部位脱色皮肤行组织学检查,HE染色检测淋巴细胞浸润情况,共聚焦激光扫描显微镜（RCM）观察小鼠皮肤的黑素和黑素细胞,免疫荧光检测CD8+ T细胞 。 结果 模型组小鼠在用药部位及非用药部位均有脱色现象。他克莫司组、吡美莫司组、茶多酚组小鼠脱色减少,出现时间晚和面积指数均较模型组低;且用药部位脱色斑局部淋巴细胞和CD8+ T细胞浸润减少。其中他克莫司组疗效优于其他组。 结论 他克莫司、吡美莫司、茶多酚对白癜风样模型小鼠均有疗效。     

Clinical comparison of human and canine atopic dermatitis using human diagnostic criteria (Japanese Dermatological Association, 2009): proposal of provisional diagnostic criteria for canine atopic dermatitis

Atopic dermatitis (AD) is a common skin disease encountered in both humans and dogs. Canine AD can be used in the analysis of naturally occurring AD; however, details of clinical comparison have been lacking. The purpose of this study is to compare those clinical features using the human diagnostic criteria (Japanese Dermatological Association, 2009). Fifty-one dogs with canine AD were evaluated by the human criteria. Prior to this study, canine AD was basically diagnosed by the fulfillment of two authentic canine AD criteria and a positive reaction against Dermatophagoides farinae in serum immunoglobulin E levels and/or in intradermal tests. Among the human AD criteria items, behavior corresponding to pruritus was observed in all 51 dogs. Skin lesions corresponding to eczematous dermatitis were seen in 50 dogs, and symmetrical distribution of skin lesions was noted in all 51 dogs. A chronic or chronically relapsing course was observed in 50 dogs. Based on these results, the concordance rate for the criteria was 96% (49/51). Differential diagnoses of AD were also investigated in the same manner. The concordance rate for the criteria was 0% (0/69) in scabies, 2% (1/50) in pyoderma, 0% (0/50) in demodicosis, 0% (0/9) in cutaneous lymphoma, 0% (0/2) in ichthyosis, 25% (2/7) in flea allergy, 48% (24/50) in seborrheic dermatitis and 75% (3/4) in food allergy. Canine AD is thus indicated as a valuable counterpart to human AD in clinical aspects. In addition, the human AD criteria could be applicable, with some modification, as provisional diagnostic criteria for canine AD.     

不同血清型沙眼衣原体小鼠生殖道感染模型的建立

目的 建立不同血清型沙眼衣原体小鼠生殖道感染模型,为研究型别与毒力的关系奠定基础。方法 将6周龄BALB/c雌性小鼠分为4组,实验组经孕酮处理后接种4.0 × 107沙眼衣原体,孕酮对照组接种McCoy细胞培养液,无孕酮对照组直接接种4.0 × 107沙眼衣原体,以及单纯小鼠的空白对照。分别于接种后4 d起观察小鼠外阴变化,阴道分泌物细胞培养、直接免疫荧光法和实时荧光PCR检测沙眼衣原体。结果 实验组接种沙眼衣原体E、F、J和K型,4 d后小鼠生殖道表现出轻微的感染迹象,7 d后阴道分泌物培养、免疫荧光和PCR检测均检出沙眼衣原体,而对照组为阴性。感染持续时间以K型最长（21 d,1/11只阳性）,其次为J型（17 d,1/11只阳性）、F型（14 d,5/11只阳性）和E型（14 d,2/11只阳性）。结论 适龄小鼠经孕酮处理后,接种一定量的沙眼衣原体,可建立不同血清型沙眼衣原体感染模型。